PrimaTB STAT-PAK Assay, a Novel, Rapid Lateral-Flow Test for Tuberculosis in Nonhuman Primates▿

PubMed Central

Lyashchenko, Konstantin P.; Greenwald, Rena; Esfandiari, Javan; Greenwald, David; Nacy, Carol A.; Gibson, Susan; Didier, Peter J.; Washington, Marc; Szczerba, Peter; Motzel, Sherri; Handt, Larry; Pollock, John M.; McNair, James; Andersen, Peter; Langermans, Jan A. M.; Verreck, Frank; Ervin, Sean; Ervin, Frank; McCombs, Candace

2007-01-01

Tuberculosis (TB) is the most important zoonotic bacterial disease in nonhuman primates (NHP). The current diagnostic method, the intradermal palpebral tuberculin test, has serious shortcomings. We characterized antibody responses in NHP against Mycobacterium tuberculosis to identify immunodominant antigens and develop a rapid serodiagnostic test for TB. A total of 422 NHP were evaluated, including 243 rhesus (Macaca mulatta), 46 cynomolgus (Macaca fascicularis), and 133 African green (Cercopithecus aethiops sabaeus) monkeys at five collaborative centers. Of those, 50 monkeys of the three species were experimentally inoculated with M. tuberculosis. Antibody responses were monitored every 2 to 4 weeks for up to 8 months postinfection by MultiAntigen Print ImmunoAssay with a panel of 12 recombinant antigens. All of the infected monkeys produced antibodies at various levels and with different antigen recognition patterns. ESAT-6 and MPB83 were the most frequently recognized proteins during infection. A combination of selected antigens which detected antibodies in all of the infected monkeys was designed to develop the PrimaTB STAT-PAK assay by lateral-flow technology. Serological evaluation demonstrated high diagnostic sensitivity (90%) and specificity (99%). The highest rate of TB detection was achieved when the skin test was combined with the PrimaTB STAT-PAK kit. This novel immunoassay provides a simple, rapid, and accurate test for TB in NHP. PMID:17652522

Rapid antigen tests for dengue virus serotypes and Zika virus in patient serum.

PubMed

Bosch, Irene; de Puig, Helena; Hiley, Megan; Carré-Camps, Marc; Perdomo-Celis, Federico; Narváez, Carlos F; Salgado, Doris M; Senthoor, Dewahar; O'Grady, Madeline; Phillips, Elizabeth; Durbin, Ann; Fandos, Diana; Miyazaki, Hikaru; Yen, Chun-Wan; Gélvez-Ramírez, Margarita; Warke, Rajas V; Ribeiro, Lucas S; Teixeira, Mauro M; Almeida, Roque P; Muñóz-Medina, José E; Ludert, Juan E; Nogueira, Mauricio L; Colombo, Tatiana E; Terzian, Ana C B; Bozza, Patricia T; Calheiros, Andrea S; Vieira, Yasmine R; Barbosa-Lima, Giselle; Vizzoni, Alexandre; Cerbino-Neto, José; Bozza, Fernando A; Souza, Thiago M L; Trugilho, Monique R O; de Filippis, Ana M B; de Sequeira, Patricia C; Marques, Ernesto T A; Magalhaes, Tereza; Díaz, Francisco J; Restrepo, Berta N; Marín, Katerine; Mattar, Salim; Olson, Daniel; Asturias, Edwin J; Lucera, Mark; Singla, Mohit; Medigeshi, Guruprasad R; de Bosch, Norma; Tam, Justina; Gómez-Márquez, Jose; Clavet, Charles; Villar, Luis; Hamad-Schifferli, Kimberly; Gehrke, Lee

2017-09-27

The recent Zika virus (ZIKV) outbreak demonstrates that cost-effective clinical diagnostics are urgently needed to detect and distinguish viral infections to improve patient care. Unlike dengue virus (DENV), ZIKV infections during pregnancy correlate with severe birth defects, including microcephaly and neurological disorders. Because ZIKV and DENV are related flaviviruses, their homologous proteins and nucleic acids can cause cross-reactions and false-positive results in molecular, antigenic, and serologic diagnostics. We report the characterization of monoclonal antibody pairs that have been translated into rapid immunochromatography tests to specifically detect the viral nonstructural 1 (NS1) protein antigen and distinguish the four DENV serotypes (DENV1-4) and ZIKV without cross-reaction. To complement visual test analysis and remove user subjectivity in reading test results, we used image processing and data analysis for data capture and test result quantification. Using a 30-μl serum sample, the sensitivity and specificity values of the DENV1-4 tests and the pan-DENV test, which detects all four dengue serotypes, ranged from 0.76 to 1.00. Sensitivity/specificity for the ZIKV rapid test was 0.81/0.86, respectively, using a 150-μl serum input. Serum ZIKV NS1 protein concentrations were about 10-fold lower than corresponding DENV NS1 concentrations in infected patients; moreover, ZIKV NS1 protein was not detected in polymerase chain reaction-positive patient urine samples. Our rapid immunochromatography approach and reagents have immediate application in differential clinical diagnosis of acute ZIKV and DENV cases, and the platform can be applied toward developing rapid antigen diagnostics for emerging viruses. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Performance Characteristics and Utilization of Rapid Antigen Test, DNA Probe, and Culture for Detection of Group A Streptococci in an Acute Care Clinic

PubMed Central

Chapin, Kimberle C.; Blake, Patricia; Wilson, Claire D.

2002-01-01

Group A streptococcus (GAS) antigen testing has become a routine point-of-care (POC) test in acute care settings. Concern about performance parameters (PP) of these tests as well as inappropriate antibiotic use has resulted in various recommendations regarding diagnosis of GAS. There were two objectives in this study. The first was to evaluate the rapid GAS antigen test presently in use (Thermo BioStar, Boulder, Colo.) and the GAS Direct probe test (Gen-Probe, San Diego, Calif.) compared to culture. The second was to define the optimal use of these technologies in a large acute care pediatric clinic. A total of 520 consecutive pediatric patients presenting with symptoms of pharyngitis at any of three Lahey Clinic acute care facilities were evaluated. Pharyngeal specimens were collected using a double-swab collection device (Copan, Corona, Calif.). One swab was used for the antigen test, the second was used for the probe test, and the pledget was placed in the collection device for culture on 5% sheep blood agar, incubated for 48 h anaerobically, and subsequently placed in Todd-Hewitt broth. After discrepant analysis, sensitivity, specificity, and positive and negative predictive values were as follows: 94.8, 100, 100, and 96.9% for the probe test and 86.1, 97.1, 93.7, and 93.4% for the antigen test, respectively. Sensitivity using an enhanced culture technique was 99.4% (163 of 164). False-positive (FP) antigen results were often seen from patients previously diagnosed and/or treated for GAS. No FP results were seen with the probe test. Colony counts for the false-negative (FN) antigen tests were higher than those for the FN probe tests. Compared to culture and DNA probe, the rapid antigen test (RAT) offered a result at the time of the patient's visit, with acceptable PP when prevalence of disease is high. Follow-up testing with the RAT of GAS patients who previously tested as positive should be avoided due to increased FP results. The probe test was comparable to culture in performance. Results indicate the probe test can be used as the primary test or as a backup to negative antigen tests. The probe test offers the advantage over culture of same-day reporting of a final result but, in contrast to a POC test, necessitates follow-up communication to the patient. Preliminary data show the specificity of the probe test to be greater than that of the RAT for patients previously diagnosed with GAS. PMID:12409399

Rapid detection of EBOLA VP40 in microchip immunofiltration assay

NASA Astrophysics Data System (ADS)

Miethe, Peter; Gary, Dominik; Hlawatsch, Nadine; Gad, Anne-Marie

2015-05-01

In the spring of 2014, the Ebola virus (EBOV) strain Zaire caused a dramatic outbreak in several regions of West Africa. The RT-PCR and antigen capture diagnostic proved to be effective for detecting EBOV in blood and serum. In this paper, we present data of a rapid antigen capture test for the detection of VP40. The test was performed in a microfluidic chip for immunofiltration analysis. The chip integrates all necessary assay components. The analytical sensitivity of the rapid test was 8 ng/ml for recombinant VP40. In serum and whole blood samples spiked with virus culture material, the detection limit was 2.2 x 102 PFU/ml. The performance data of the rapid test (15 min) are comparable to that of the VP40 laboratory ELISA.

9 CFR 145.14 - Testing.

Code of Federal Regulations, 2014 CFR

2014-01-01

... immunosorbent assay test (ELISA), or the rapid serum test for all poultry; and the stained antigen, rapid whole... test or enzyme-labeled immunosorbent assay test (ELISA), or blood from birds that react on the stained... enzyme-linked immunosorbent assay (ELISA) test,3 a polymerase chain reaction (PCR)-based test, or a...

9 CFR 145.14 - Testing.

Code of Federal Regulations, 2012 CFR

2012-01-01

... immunosorbent assay test (ELISA), or the rapid serum test for all poultry; and the stained antigen, rapid whole... test or enzyme-labeled immunosorbent assay test (ELISA), or blood from birds that react on the stained... enzyme-linked immunosorbent assay (ELISA) test,3 a polymerase chain reaction (PCR)-based test, or a...

9 CFR 145.14 - Testing.

Code of Federal Regulations, 2013 CFR

2013-01-01

... immunosorbent assay test (ELISA), or the rapid serum test for all poultry; and the stained antigen, rapid whole... test or enzyme-labeled immunosorbent assay test (ELISA), or blood from birds that react on the stained... enzyme-linked immunosorbent assay (ELISA) test,3 a polymerase chain reaction (PCR)-based test, or a...

Rapid diagnosis of equine influenza by highly sensitive silver amplification immunochromatography system.

PubMed

Yamanaka, Takashi; Nemoto, Manabu; Bannai, Hiroshi; Tsujimura, Koji; Kondo, Takashi; Matsumura, Tomio; Fu, Tao Qi Huang; Fernandez, Charlene Judith; Gildea, Sarah; Cullinane, Ann

2017-06-16

Equine influenza (EI) is a respiratory disease caused by equine influenza A virus (EIV, H3N8) infection. Rapid diagnosis is essential to limit the disease spread. We previously reported that some rapid antigen detection (RAD) tests are fit for diagnosing EI although their sensitivity is not optimal. Here, we evaluated the performance of the newly developed RAD test using silver amplification immunochromatography (Quick Chaser Auto Flu A, B: QCA) to diagnose EI. The detection limits of QCA for EIVs were five-fold lower than the conventional RAD tests. The duration of virus antigen detection in the infected horses was longer than the conventional RAD tests. We conclude that QCA could be a valuable diagnostic method for EI.

Multicentre evaluations of two new rapid IgG4 tests (WB rapid and panLF rapid) for detection of lymphatic filariasis

PubMed Central

Noordin, Rahmah; Itoh, Makoto; Kimura, Eisaku; Abdul Rahman, Rohana; Ravindran, Balachandran; Mahmud, Rohela; Supali, Taniawati; Weerasooriya, Mirani

2007-01-01

In the global effort to eliminate lymphatic filariasis (LF), rapid field-applicable tests are useful tools that will allow on-site testing to be performed in remote places and the results to be obtained rapidly. Exclusive reliance on the few existing tests may jeopardize the progress of the LF elimination program, thus the introduction of other rapid tests would be useful to address this issue. Two new rapid immunochromatographic IgG4 cassette tests have been produced, namely WB rapid and panLF rapid, for detection of bancroftian filariasis and all three species of lymphatic filaria respectively. WB rapid was developed using BmSXP recombinant antigen, while PanLF rapid was developed using BmR1 and BmSXP recombinant antigens. A total of 165 WB rapid and 276 panLF rapid tests respectively were evaluated at USM and the rest were couriered to another university in Malaysia (98 WB rapid, 129 panLF rapid) and to universities in Indonesia (56 WB rapid, 62 panLF rapid), Japan (152 of each test) and India (18 of each test) where each of the tests underwent independent evaluations in a blinded manner. The average sensitivities of WB rapid and panLF rapid were found to be 97.6% (94%–100%) and 96.5% (94%–100%) respectively; while their average specificities were both 99.6% (99%–100%). Thus this study demonstrated that both the IgG4 rapid tests were highly sensitive and specific, and would be useful additional tests to facilitate the global drive to eliminate this disease. PMID:17961262

Evaluation of a New and Rapid Serologic Test for Detecting Brucellosis: Brucella Coombs Gel Test.

PubMed

Hanci, Hayrunisa; Igan, Hakan; Uyanik, Muhammet Hamidullah

2017-01-01

Many serological tests have been used for the diagnosis of human brucellosis. A new serological method is identified as Brucella Coombs gel test based on the principle of centrifugation gel system similar to the gel system used in blood group determination. In this system, if Brucella antibodies were present in the serum, antigen and antibody would remain as a pink complex on the gel. Otherwise, the pink Brucella antigens would precipitate at the bottom of the gel card system. In this study, we aimed to compare the Brucella Coombs gel test, a new, rapid screen and titration method for detection of non-agglutinating IgG with the Brucella Coombs test. For this study, a total of 88 serum samples were obtained from 45 healthy persons and 43 individuals who had clinical signs and symptoms of brucellosis. For each specimen, Rose Bengal test, standard agglutination test, Coombs test and Brucella Coombs gel test were carried out. Sensitivity and specificity of Brucella Coombs gel test were found as 100.0 and 82.2%, respectively. Brucella Coombs gel test can be used as a screening test with high sensitivity. By the help of pink Brucella antigen precipitation, the tests' evaluation is simple and objective. In addition, determination of Brucella antibody by rapid titration offers another important advantage.

RAPID DIAGNOSIS OF PNEUMOCOCCAL PNEUMONIA AMONG HIV-INFECTED ADULTS WITH URINE ANTIGEN DETECTION

PubMed Central

Boulware, David R; Daley, Charles L.; Merrifield, Cynthia; Hopewell, Philip C.; Janoff, Edward N.

2007-01-01

Objectives Streptococcus pneumoniae is the leading cause of bacterial pneumonia and associated bacteremia during HIV infection. Rapid diagnostic assays may limit inappropriate therapy. Methods Clinical signs and symptoms and sera and urine were collected prospectively from 70 adults with pneumococcal pneumonia, including 47 with HIV co-infection. Pneumococcal C-polysaccharide antigen was detected in urine using the Binax® immunochromatographic test (ICT). A systematic review of 24 published studies was conducted. Results Clinical symptoms, signs, and laboratory parameters except leukocytosis, were similar in HIV-infected and HIV-seronegative pneumonia. The performance of the urine antigen ICT was independent of HIV-status (sensitivity 81%, specificity 98%, positive (PPV) and negative predictive values (NPV) 98%, and 82%, respectively). The sensitivity of sputum Gram’s stain was 58% [34/59] with sputum unable to be provided by 16%. The CRP response was identical in HIV-infected (mean ± SD) 133 ± 88 vs. seronegative 135 ± 104 mg/L (p=0.9). In the systematic review, the ICT performance revealed 74% sensitivity (95% CI: 72% to 77%) and 94% specificity (95% CI: 93% to 95%). Urine antigen testing increases etiologic diagnosis by 23% (Range: 10% –59%) when testing adults with community acquired pneumonia of unknown etiology. Conclusions Urinary antigen detection provides a credible rapid diagnostic test for pneumococcal pneumonia regardless of HIV-status. CRP response to acute infection is similar in HIV co-infection and increases diagnostic certainty. PMID:17692384

A comparative study of the diagnostic methods for Group A streptococcal sore throat in two reference hospitals in Yaounde, Cameroon

PubMed Central

Gonsu, Hortense Kamga; Bomki, Cynthia Mbimenyuy; Djomou, François; Toukam, Michel; Ndze, Valantine Ngum; Lyonga, Emilia Enjema; Mbakop, Calixte Didier; Koulla-Shiro, Sinata

2015-01-01

Introduction Sore throat is a common complaint in general practice which is more frequent in children. The most frequent pathogenic bacteria associated with this infection is Streptococcus pyogenes. Rapid Antigen Diagnostic Test (RADT) facilitates the rapid identification and consequently prompt treatment of patients, prevents complications, and also reduces the risk of spread of Group A Streptococcus (GAS). The main objective of this study was to assess the diagnostic value of a rapid streptococcal antigen detection test in patients with sore throat. Methods A cross-sectional descriptive study was carried out from January to April 2011 on patients aged 3 to 72 years consulting for pharyngitis or sore throat at the paediatric and Ear, Nose and Throat units of the University Teaching Hospital Yaounde and the Central Hospital Yaounde. Two throat swabs were collected per patient. One was used for the rapid test and the other for standard bacteriological analysis. Results The prevalence of GAS in the study population was 22.5%. Out of the 71 samples collected, the RADT detected group A streptococcal antigens in 12 of 16 positive cultures giving a sensitivity of 75%. The specificity of the rapid test was 96%, with positive predictive value of 85.7%, and negative predictive value of 93% respectively. Conclusion Rapid test may have an additional value in the management of patients with high risk of having GAS infection. However, tests with a higher sensitivity are needed for accurate and reliable results for early diagnosis of patients with sore throat caused by GAS. PMID:27386017

A comparative study of the diagnostic methods for Group A streptococcal sore throat in two reference hospitals in Yaounde, Cameroon.

PubMed

Gonsu, Hortense Kamga; Bomki, Cynthia Mbimenyuy; Djomou, François; Toukam, Michel; Ndze, Valantine Ngum; Lyonga, Emilia Enjema; Mbakop, Calixte Didier; Koulla-Shiro, Sinata

2015-01-01

Sore throat is a common complaint in general practice which is more frequent in children. The most frequent pathogenic bacteria associated with this infection is Streptococcus pyogenes. Rapid Antigen Diagnostic Test (RADT) facilitates the rapid identification and consequently prompt treatment of patients, prevents complications, and also reduces the risk of spread of Group A Streptococcus (GAS). The main objective of this study was to assess the diagnostic value of a rapid streptococcal antigen detection test in patients with sore throat. A cross-sectional descriptive study was carried out from January to April 2011 on patients aged 3 to 72 years consulting for pharyngitis or sore throat at the paediatric and Ear, Nose and Throat units of the University Teaching Hospital Yaounde and the Central Hospital Yaounde. Two throat swabs were collected per patient. One was used for the rapid test and the other for standard bacteriological analysis. The prevalence of GAS in the study population was 22.5%. Out of the 71 samples collected, the RADT detected group A streptococcal antigens in 12 of 16 positive cultures giving a sensitivity of 75%. The specificity of the rapid test was 96%, with positive predictive value of 85.7%, and negative predictive value of 93% respectively. Rapid test may have an additional value in the management of patients with high risk of having GAS infection. However, tests with a higher sensitivity are needed for accurate and reliable results for early diagnosis of patients with sore throat caused by GAS.

A Novel Technique for Detecting Antibiotic-Resistant Typhoid from Rapid Diagnostic Tests

PubMed Central

Nic Fhogartaigh, Caoimhe; Dance, David A. B.; Davong, Viengmon; Tann, Pisey; Phetsouvanh, Rattanaphone; Turner, Paul; Newton, Paul N.

2015-01-01

Fluoroquinolone-resistant typhoid is increasing. An antigen-detecting rapid diagnotic test (RDT) can rapidly diagnose typhoid from blood cultures. A simple, inexpensive molecular technique performed with DNA from positive RDTs accurately identified gyrA mutations consistent with phenotypic susceptibility testing results. Field diagnosis combined with centralized molecular resistance testing could improve typhoid management and surveillance in low-resource settings. PMID:25762768

The latex agglutination test versus counterimmunoelectrophoresis for rapid diagnosis of bacterial meningitis

PubMed Central

Bortolussi, Robert; Wort, Arthur J.; Casey, Stephanie

1982-01-01

A modified latex agglutination (LA) test was compared with Gram-staining and counterimmunoelectrophoresis (CIE) for the rapid detection in the cerebrospinal fluid (CSF) of antigen to Haemophilus influenzae type b, Neisseria meningitidis groups A, B and C, Escherichia coli K1, Streptococcus pneumoniae and group B streptococci, seven frequent causes of bacterial meningitis in children. Of 50 CSF samples from patients with culture-proven bacterial meningitis 90% were correctly shown by the LA test to contain antigen of the responsible organism. Gram-staining revealed organisms in 80% of 45 of these samples. In 75% of the 40 samples that were of sufficient volume for CIE, positive results for the appropriate antigen were obtained. The concentration of antigen detected in the CSF by the LA test varied from undetectable to 800 000 ng/ml. Patients with a high concentration (more than 2000 ng/ml or a positive result at dilutions of CSF over 1/8) were significantly more likely to have a poor response to therapy (two died and two had persistent pleocytosis or bacteria in the CSF) than patients with a lower concentration (4/16 v. 0/18, P < 0.05). After appropriate therapy was begun the concentration of antigen fell dramatically, but measurable amounts of antigen persisted in the CSF for up to 6 days. The LA test detected bacterial antigen at concentrations 2 to 70 times below the lower limit detected by CIE. In seven additional patients who had received antibiotics before lumbar puncture was performed the LA test detected antigen from meningitis-causing bacteria even though cultures of the CSF were sterile. In another 145 patients who did not have meningitis the results of the LA test were negative. The LA test, done as described in this article, is easier to perform than CIE and should be a useful addition to the diagnostic tests carried out on the CSF of any patient suspected of having meningitis. PMID:6749272

Simple Tests for Rapid Detection of Canine Parvovirus Antigen and Canine Parvovirus-Specific Antibodies▿ †

PubMed Central

Marulappa, Shashidhara Y.; Kapil, Sanjay

2009-01-01

Canine parvovirus (CPV) is the number one viral cause of enteritis, morbidity, and mortality in 8-week-old young puppies. We have developed twin assays (slide agglutination test [SAT] for CPV antigen and slide inhibition test [SIT] for CPV antibody) that are sensitive, specific, cost-effective, generic for all genotypes of CPV, and provide instant results for CPV antigen detection in feces and antibody quantification in serum. We found these assays to be useful for routine applications in kennels with large numbers of puppies at risk. The results of these assays are available in 1 min and do not require any special instrumentation. SAT-SIT technology will find applications in rapid screening of samples for other hemagglutinating emerging viruses of animals and humans (influenza virus and severe acute respiratory syndrome coronavirus). PMID:18987166

Selective testing strategies for diagnosing group A streptococcal infection in children with pharyngitis: a systematic review and prospective multicentre external validation study

PubMed Central

Cohen, Jérémie F.; Cohen, Robert; Levy, Corinne; Thollot, Franck; Benani, Mohamed; Bidet, Philippe; Chalumeau, Martin

2015-01-01

Background: Several clinical prediction rules for diagnosing group A streptococcal infection in children with pharyngitis are available. We aimed to compare the diagnostic accuracy of rules-based selective testing strategies in a prospective cohort of children with pharyngitis. Methods: We identified clinical prediction rules through a systematic search of MEDLINE and Embase (1975–2014), which we then validated in a prospective cohort involving French children who presented with pharyngitis during a 1-year period (2010–2011). We diagnosed infection with group A streptococcus using two throat swabs: one obtained for a rapid antigen detection test (StreptAtest, Dectrapharm) and one obtained for culture (reference standard). We validated rules-based selective testing strategies as follows: low risk of group A streptococcal infection, no further testing or antibiotic therapy needed; intermediate risk of infection, rapid antigen detection for all patients and antibiotic therapy for those with a positive test result; and high risk of infection, empiric antibiotic treatment. Results: We identified 8 clinical prediction rules, 6 of which could be prospectively validated. Sensitivity and specificity of rules-based selective testing strategies ranged from 66% (95% confidence interval [CI] 61–72) to 94% (95% CI 92–97) and from 40% (95% CI 35–45) to 88% (95% CI 85–91), respectively. Use of rapid antigen detection testing following the clinical prediction rule ranged from 24% (95% CI 21–27) to 86% (95% CI 84–89). None of the rules-based selective testing strategies achieved our diagnostic accuracy target (sensitivity and specificity > 85%). Interpretation: Rules-based selective testing strategies did not show sufficient diagnostic accuracy in this study population. The relevance of clinical prediction rules for determining which children with pharyngitis should undergo a rapid antigen detection test remains questionable. PMID:25487666

Performances of fourth generation HIV antigen/antibody assays on filter paper for detection of early HIV infections.

PubMed

Kania, Dramane; Truong, Tam Nguyen; Montoya, Ana; Nagot, Nicolas; Van de Perre, Philippe; Tuaillon, Edouard

2015-01-01

Point-of-care testing and diagnosis of HIV acute infections play important roles in preventing transmission, but HIV rapid diagnosis tests have poor capacity to detect early infections. Filter paper can be used for capillary blood collection and HIV testing using 4th generation immunoassays. Antigen/antibody combined immunoassays were evaluated for their capacity to identify early HIV infections using filter paper in comparison with rapid test. Thirty nine serum samples collected from HIV seroconverters were spotted onto filter paper and tested by the Roche Elecsys(®) HIV Combi PT test and the DiaSorin Liaison XL Murex HIV Ab/Ag assay. Fourth generation immunoassays identified 34 out of 39 HIV early infections using dried serum spot, whereas the Determine™ HIV-1/2 rapid test detected 24 out of 39 HIV positive serum (87.2% vs 61.5% respectively, p = 0.009). p24 antigen was detected by the Liaison XL in 19 dried serum samples (48.7%). In the group characterized by a negative western blot, 7 out of 8 (87.5%) and 6 out of 8 (75.0%) samples were found positive for HIV using the Elecsys and the Liaison XL, respectively. None of these eight samples classified in this group of early acute infections were found positive by the rapid test. Fourth generation Ag/Ab immunoassays performed on dried serum spot had good performance for HIV testing during the early phases of HIV infection. This method may be useful to detect HIV early infections in hard-to-reach populations and individuals living in remote areas before rapid tests become positive. Copyright © 2014 Elsevier B.V. All rights reserved.

Structural and functional studies of a 50 kDa antigenic protein from Salmonella enterica serovar Typhi.

PubMed

Choong, Yee Siew; Lim, Theam Soon; Chew, Ai Lan; Aziah, Ismail; Ismail, Asma

2011-04-01

The high typhoid incidence rate in developing and under-developed countries emphasizes the need for a rapid, affordable and accessible diagnostic test for effective therapy and disease management. TYPHIDOT®, a rapid dot enzyme immunoassay test for typhoid, was developed from the discovery of a ∼50 kDa protein specific for Salmonella enterica serovar Typhi. However, the structure of this antigen remains unknown till today. Studies on the structure of this antigen are important to elucidate its function, which will in turn increase the efficiency of the development and improvement of the typhoid detection test. This paper described the predictive structure and function of the antigenically specific protein. The homology modeling approach was employed to construct the three-dimensional structure of the antigen. The built structure possesses the features of TolC-like outer membrane protein. Molecular docking simulation was also performed to further probe the functionality of the antigen. Docking results showed that hexamminecobalt, Co(NH(3))(6)(3+), as an inhibitor of TolC protein, formed favorable hydrogen bonds with D368 and D371 of the antigen. The single point (D368A, D371A) and double point (D368A and D371A) mutations of the antigen showed a decrease (single point mutation) and loss (double point mutations) of binding affinity towards hexamminecobalt. The architecture features of the built model and the docking simulation reinforced and supported that this antigen is indeed the variant of outer membrane protein, TolC. As channel proteins are important for the virulence and survival of bacteria, therefore this ∼50 kDa channel protein is a good specific target for typhoid detection test. Copyright © 2011 Elsevier Inc. All rights reserved.

Integrated Magneto-Chemical Sensor For On-Site Food Allergen Detection.

PubMed

Lin, Hsing-Ying; Huang, Chen-Han; Park, Jongmin; Pathania, Divya; Castro, Cesar M; Fasano, Alessio; Weissleder, Ralph; Lee, Hakho

2017-10-24

Adverse food reactions, including food allergies, food sensitivities, and autoimmune reaction (e.g., celiac disease) affect 5-15% of the population and remain a considerable public health problem requiring stringent food avoidance and epinephrine availability for emergency events. Avoiding problematic foods is practically difficult, given current reliance on prepared foods and out-of-home meals. In response, we developed a portable, point-of-use detection technology, termed integrated exogenous antigen testing (iEAT). The system consists of a disposable antigen extraction device coupled with an electronic keychain reader for rapid sensing and communication. We optimized the prototype iEAT system to detect five major food antigens in peanuts, hazelnuts, wheat, milk, and eggs. Antigen extraction and detection with iEAT requires <10 min and achieves high-detection sensitivities (e.g., 0.1 mg/kg for gluten, lower than regulatory limits of 20 mg/kg). When testing under restaurant conditions, we were able to detect hidden food antigens such as gluten within "gluten-free" food items. The small size and rapid, simple testing of the iEAT system should help not only consumers but also other key stakeholders such as clinicians, food industries, and regulators to enhance food safety.

Comparison of Xpert Flu rapid nucleic acid testing with rapid antigen testing for the diagnosis of influenza A and B.

PubMed

DiMaio, Michael A; Sahoo, Malaya K; Waggoner, Jesse; Pinsky, Benjamin A

2012-12-01

Influenza infections are associated with thousands of hospital admissions and deaths each year. Rapid detection of influenza is important for prompt initiation of antiviral therapy and appropriate patient triage. In this study the Cepheid Xpert Flu assay was compared with two rapid antigen tests, BinaxNOW Influenza A & B and BD Directigen EZ Flu A+B, as well as direct fluorescent antibody testing for the rapid detection of influenza A and B. Using real-time, hydrolysis probe-based, reverse transcriptase PCR as the reference method, influenza A sensitivity was 97.3% for Xpert Flu, 95.9% for direct fluorescent antibody testing, 62.2% for BinaxNOW, and 71.6% for BD Directigen. Influenza B sensitivity was 100% for Xpert Flu and direct fluorescent antibody testing, 54.5% for BinaxNOW, and 48.5% for BD Directigen. Specificity for influenza A was 100% for Xpert Flu, BinaxNOW, and BD Directigen, and 99.2% for direct fluorescent antibody testing. All methods demonstrated 100% specificity for influenza B. These findings support the use of the Xpert Flu assay in settings requiring urgent diagnosis of influenza A and B. Copyright © 2012 Elsevier B.V. All rights reserved.

Rapid Serodiagnosis of Active Pulmonary Mycobacterium tuberculosis by Analysis of Results from Multiple Antigen-Specific Tests

PubMed Central

Okuda, Yoshinari; Maekura, Ryoji; Hirotani, Atsusi; Kitada, Seigo; Yoshimura, Kenji; Hiraga, Touru; Yamamoto, Yuoko; Itou, Masami; Ogura, Takeshi; Ogihara, Toshio

2004-01-01

We have prospectively analyzed three antigens for serodiagnosis of tuberculosis (TB). These antigens were tuberculous glycolipid antigen, lypoarabinomannan polysaccharide antigen, and antigen 60 (A60), which was derived from purified protein derivatives. Of the 131 patients with active pulmonary TB, 57 were both smear and culture negative and 14 had chronic active pulmonary TB that remained smear positive for >12 months of chemotherapy. One hundred twenty healthy adults were controls. The percentages of patients positive in all three tests were 58.8% for smear-positive active pulmonary TB and 71.4% for chronic active pulmonary TB. When the results of the three serodiagnostic tests were evaluated in combination, the sensitivity increased to 91.5% in patients with active pulmonary TB and to 86.0% in smear- and culture-negative patients. The false-positive rate of the three-test combination was 12.5% in the healthy control groups. In conclusion, it was not possible to detect all of the antibodies against antigenic substances in the cell walls of the tuberculous bacilli in the sera of all TB patients by using available serodiagnostic tests. However, the combined use of tests with three separate antigens maximizes the effectiveness of serodiagnosis. PMID:15004065

Diagnostic sensitivity and specificity of a participatory disease surveillance method for highly pathogenic avian influenza in household chicken flocks in Indonesia.

PubMed

Robyn, M; Priyono, W B; Kim, L M; Brum, E

2012-06-01

A study was conducted to assess the diagnostic sensitivity and specificity of a disease surveillance method for diagnosis of highly pathogenic avian influenza (HPAI) outbreaks in household chicken flocks used by participatory disease surveillance (PDS) teams in Yogyakarta Province, Indonesia. The Government of Indonesia, in partnership with the Food and Agriculture Organization of the United Nations, has implemented a PDS method for the detection of HPAI outbreaks in poultry since 2006. The PDS method in Indonesia utilizes both a clinical case definition (CD) and the result of a commercial rapid antigen test kit Yogyakarta 55611, to diagnose HPAI outbreaks, primarily in backyard chicken flocks. The following diagnostic sensitivities and specificities were obtained relative to real-time reverse transcription-PCR as the gold standard diagnostic test: 1) 89% sensitivity (CI95: 75%-97%) and 96% specificity (CI95: 89%-99%) for the PDS CD alone; 2) 86% sensitivity (CI95: 71%-95%) and 99% specificity (CI95: 94%-100%) for the rapid antigen test alone; and 3) 84% sensitivity (CI95: 68%-94%) and 100% specificity (CI95: 96%-100%) for the PDS CD result combined with the rapid antigen test result. Based on these results, HPAI outbreaks in extensively raised household chickens can be diagnosed with sufficient sensitivity and specificity using the PDS method as implemented in Indonesia. Subject to further field evaluation, data from this study suggest that the diagnostic sensitivity of the PDS method may be improved by expanding the PDS CD to include more possible clinical presentations of HPAI and by increasing the number of rapid antigen tests to three different birds with HPAI-compatible signs of same flock.

A Lateral Flow Rapid Test for Human Toxocariasis Developed Using Three Toxocara canis Recombinant Antigens.

PubMed

Yunus, Muhammad Hafiznur; Tan Farrizam, Siti Naqiuyah; Abdul Karim, Izzati Zahidah; Noordin, Rahmah

2018-01-01

Laboratory diagnosis of toxocariasis is still a challenge especially in developing endemic countries with polyparasitism. In this study, three Toxocara canis recombinant antigens, rTES-26, rTES-30, and rTES-120, were expressed and used to prepare lateral flow immunoglobulin G4 (IgG4) dipsticks. The concordance of the results of the rapid test (comprising three dipsticks) with a commercial IgG-enzyme-linked immunosorbent assay (ELISA) (Cypress Diagnostics, Belgium) was compared against the concordance of two other commercial IgG-ELISA kits (Bordier, Switzerland and NovaTec, Germany) with the Cypress kit. Using Toxocara- positive samples, the concordance of the dipstick dotted with rTES-26, rTES-30, and rTES-120 was 41.4% (12/29), 51.7% (15/29), and 72.4% (21/29), respectively. When positivity with any dipstick was considered as an overall positive rapid test result, the concordance with the Cypress kit was 93% (27/29). Meanwhile, when compared with the results of the Cypress kit, the concordance of IgG-ELISA from NovaTec and Bordier was 100% (29/29) and 89.7% (26/29), respectively. Specific IgG4 has been recognized as a marker of active infection for several helminthic diseases; therefore, the two non-concordant results of the rapid test when compared with the NovaTec IgG-ELISA kit may be from samples of people with non-active infection. All the three dipsticks showed 100% (50/50) concordance with the Cypress kit when tested with serum from individuals who were healthy and with other infections. In conclusion, the lateral flow rapid test is potentially a good, fast, and easy test for toxocariasis. Next, further validation studies and development of a test with the three antigens in one dipstick will be performed.

Novel Antigen Identification Method for Discovery of Protective Malaria Antigens by Rapid Testing of DNA Vaccines Encoding Exons from the Parasite Genome

PubMed Central

Haddad, Diana; Bilcikova, Erika; Witney, Adam A.; Carlton, Jane M.; White, Charles E.; Blair, Peter L.; Chattopadhyay, Rana; Russell, Joshua; Abot, Esteban; Charoenvit, Yupin; Aguiar, Joao C.; Carucci, Daniel J.; Weiss, Walter R.

2004-01-01

We describe a novel approach for identifying target antigens for preerythrocytic malaria vaccines. Our strategy is to rapidly test hundreds of DNA vaccines encoding exons from the Plasmodium yoelii yoelii genomic sequence. In this antigen identification method, we measure reduction in parasite burden in the liver after sporozoite challenge in mice. Orthologs of protective P. y. yoelii genes can then be identified in the genomic databases of Plasmodium falciparum and Plasmodium vivax and investigated as candidate antigens for a human vaccine. A pilot study to develop the antigen identification method approach used 192 P. y. yoelii exons from genes expressed during the sporozoite stage of the life cycle. A total of 182 (94%) exons were successfully cloned into a DNA immunization vector with the Gateway cloning technology. To assess immunization strategies, mice were vaccinated with 19 of the new DNA plasmids in addition to the well-characterized protective plasmid encoding P. y. yoelii circumsporozoite protein. Single plasmid immunization by gene gun identified a novel vaccine target antigen which decreased liver parasite burden by 95% and which has orthologs in P. vivax and P. knowlesi but not P. falciparum. Intramuscular injection of DNA plasmids produced a different pattern of protective responses from those seen with gene gun immunization. Intramuscular immunization with plasmid pools could reduce liver parasite burden in mice despite the fact that none of the plasmids was protective when given individually. We conclude that high-throughput cloning of exons into DNA vaccines and their screening is feasible and can rapidly identify new malaria vaccine candidate antigens. PMID:14977966

Comparison of Sofia Legionella FIA and BinaxNOW® Legionella urinary antigen card in two national reference centers.

PubMed

Beraud, L; Gervasoni, K; Freydiere, A M; Descours, G; Ranc, A G; Vandenesch, F; Lina, G; Gaia, V; Jarraud, S

2015-09-01

The Sofia Legionella Fluorescence Immunoassay (FIA; Quidel) is a recently introduced rapid immunochromatographic diagnostic test for Legionnaires' disease using immunofluorescence technology designed to enhance its sensitivity. The aim of this study was to evaluate its performance for the detection of urinary antigens for Legionella pneumophila serogroup 1 in two National Reference Centers for Legionella. The sensitivity and specificity of the Sofia Legionella FIA test were determined in concentrated and nonconcentrated urine samples, before and after boiling, in comparison with the BinaxNOW® Legionella Urinary Antigen Card (UAC; Alere). Compared with BinaxNOW® Legionella UAC, the sensitivity of the Sofia Legionella test was slightly higher in nonconcentrated urine samples and was identical in concentrated urine samples. The specificity of the Sofia Legionella FIA test was highly reduced by the concentration of urine samples. In nonconcentrated samples, a lack of specificity was observed in 2.3 % of samples, all of them resolved by heat treatment. The Sofia Legionella FIA is a sensitive test for detecting Legionella urinary antigens with no previous urine concentration. However, all positive samples have to be re-tested after boiling to reach a high specificity. The reading is automatized on the Sofia analyzer, which can be connected to laboratory information systems, facilitating accurate and rapid reporting of results.

Plasmodium knowlesi malaria in a traveller returning from the Philippines to Italy, 2016.

PubMed

De Canale, Ettore; Sgarabotto, Dino; Marini, Giulia; Menegotto, Nicola; Masiero, Serena; Akkouche, Wassim; Biasolo, Maria Angela; Barzon, Luisa; Palù, Giorgio

2017-10-01

Plasmodium knowlesi is a simian parasite responsible for most human cases of malaria in Malaysian Borneo. A timely recognition of infection is crucial because of the risk of severe disease due to the rapid increase in parasitemia. We report a case of P. knowlesi infection in a traveller who developed fever and thrombocytopenia after returning from the Philippines in 2016. Rapid antigen test was negative, microscopy examination showed parasites similar to Plasmodium malariae, with a parasite count of 10,000 parasites per μL blood, while molecular testing identified P. knowlesi infection. Treatment with atovaquone-proguanil led to resolution of fever and restoration of platelet count in two days. P. knowlesi infection should be suspected in febrile travellers returning from South East Asia. Due to the low sensitivity of rapid antigen tests and the low specificity of microscopy, confirmation by molecular tests is recommended.

Clinical score and rapid antigen detection test to guide antibiotic use for sore throats: randomised controlled trial of PRISM (primary care streptococcal management).

PubMed

Little, Paul; Hobbs, F D Richard; Moore, Michael; Mant, David; Williamson, Ian; McNulty, Cliodna; Cheng, Ying Edith; Leydon, Geraldine; McManus, Richard; Kelly, Joanne; Barnett, Jane; Glasziou, Paul; Mullee, Mark

2013-10-10

To determine the effect of clinical scores that predict streptococcal infection or rapid streptococcal antigen detection tests compared with delayed antibiotic prescribing. Open adaptive pragmatic parallel group randomised controlled trial. Primary care in United Kingdom. Patients aged ≥ 3 with acute sore throat. An internet programme randomised patients to targeted antibiotic use according to: delayed antibiotics (the comparator group for analyses), clinical score, or antigen test used according to clinical score. During the trial a preliminary streptococcal score (score 1, n=1129) was replaced by a more consistent score (score 2, n=631; features: fever during previous 24 hours; purulence; attends rapidly (within three days after onset of symptoms); inflamed tonsils; no cough/coryza (acronym FeverPAIN). Symptom severity reported by patients on a 7 point Likert scale (mean severity of sore throat/difficulty swallowing for days two to four after the consultation (primary outcome)), duration of symptoms, use of antibiotics. For score 1 there were no significant differences between groups. For score 2, symptom severity was documented in 80% (168/207 (81%) in delayed antibiotics group; 168/211 (80%) in clinical score group; 166/213 (78%) in antigen test group). Reported severity of symptoms was lower in the clinical score group (-0.33, 95% confidence interval -0.64 to -0.02; P=0.04), equivalent to one in three rating sore throat a slight versus moderate problem, with a similar reduction for the antigen test group (-0.30, -0.61 to -0.00; P=0.05). Symptoms rated moderately bad or worse resolved significantly faster in the clinical score group (hazard ratio 1.30, 95% confidence interval 1.03 to 1.63) but not the antigen test group (1.11, 0.88 to 1.40). In the delayed antibiotics group, 75/164 (46%) used antibiotics. Use of antibiotics in the clinical score group (60/161) was 29% lower (adjusted risk ratio 0.71, 95% confidence interval 0.50 to 0.95; P=0.02) and in the antigen test group (58/164) was 27% lower (0.73, 0.52 to 0.98; P=0.03). There were no significant differences in complications or reconsultations. Targeted use of antibiotics for acute sore throat with a clinical score improves reported symptoms and reduces antibiotic use. Antigen tests used according to a clinical score provide similar benefits but with no clear advantages over a clinical score alone. ISRCTN32027234.

Comparison of three feline leukaemia virus (FeLV) point-of-care antigen test kits using blood and saliva.

PubMed

Westman, Mark E; Malik, Richard; Hall, Evelyn; Sheehy, Paul A; Norris, Jacqueline M

2017-02-01

Feline leukaemia virus (FeLV) can be a challenging infection to diagnose due to a complex feline host-pathogen relationship and occasionally unreliable test results. This study compared the accuracy of three point-of-care (PoC) FeLV p27 antigen test kits commonly used in Australia and available commercially worldwide (SNAP FIV/FeLV Combo, Witness FeLV/FIV and Anigen Rapid FIV/FeLV), using detection of FeLV provirus by an in-house real-time polymerase chain reaction (qPCR) assay as the diagnostic gold standard. Blood (n=563) and saliva (n=419) specimens were collected from a population of cats determined to include 491 FeLV-uninfected and 72 FeLV-infected individuals (45 progressive infections [p27 and qPCR positive], 27 regressive infections [p27 negative, qPCR positive]). Sensitivity and specificity using whole blood was 63% and 94% for SNAP Combo, 57% and 98% for Witness, and 57% and 98% for Anigen Rapid, respectively. SNAP Combo had a significantly lower specificity using blood compared to the other two kits (P=0.004 compared to Witness, P=0.007 compared to Anigen Rapid). False-positive test results occurred with all three kits using blood, and although using any two kits in parallel increased specificity, no combination of kits completely eliminated the occurrence of false-positive results. We therefore recommend FeLV proviral PCR testing for any cat that tests positive with a PoC FeLV antigen kit, as well as for any cat that has been potentially exposed to FeLV but tests negative with a FeLV antigen kit, before final assignment of FeLV status can be made with confidence. For saliva testing, sensitivity and specificity was 54% and 100%, respectively, for all three test kits. The reduced sensitivity of saliva testing compared to blood testing, although not statistically significant, suggests saliva testing with the current generation of PoC FeLV antigen kits is unsuitable for screening large populations of cats, such as in shelters. Copyright © 2016 Elsevier Ltd. All rights reserved.

Which is the best method to trace group A streptococci in sore throat patients: culture or GAS antigen test?

PubMed

Lindbaek, Morten; Høiby, Ernst Arne; Lermark, Gro; Steinsholt, Inger Marie; Hjortdahl, Per

2004-12-01

To compare an antigen detection test (GAS antigen test) with the results from combinations of two various bacteriological test media in general practice patients with sore throat. Furthermore to assess the diagnostic properties of the chosen GAS antigen test and to compare semi-quantitative results of this test with the bacterial load found in the throat culture. Two Norwegian general practices in Stokke and Kongsberg communities. 306 patients with sore throat lasting less than 7 days; 244 were adults, 62 were children under 10 years old, mean age 23.9 years (SD 15.0), 40% were men. Results from GAS antigen test, and distribution of bacteriological findings in throat cultures, compared with the results of our GAS antigen test; semi-quantitative results of the GAS antigen test compared with the bacterial load by culture. In the primary culture 110 patients harboured group A streptococci (GAS) infection, while the second culture identified another 17, giving a total of 127 patients. Some 33 patients harboured large-colony groups C and G. The GAS antigen test used had a sensitivity of 97% and specificity of 95% regarding GAS when compared with the two cultures. We found a significant correlation between the bacterial loads by culture and the semi-quantitative results of the GAS antigen test. By using a second, different set of bacteriological media, we identified an additional 17 patients with GAS infections. This raises the question of validity of frequently used reference standards in studies related to streptococcal infections. Compared with the combined results of the two throat cultures, the GAS antigen test used showed high sensitivity and specificity. Semi-quantitative evaluations of the rapid immunological test may also be of clinical value.

Analytical Validation of the ReEBOV Antigen Rapid Test for Point-of-Care Diagnosis of Ebola Virus Infection

PubMed Central

Cross, Robert W.; Boisen, Matthew L.; Millett, Molly M.; Nelson, Diana S.; Oottamasathien, Darin; Hartnett, Jessica N.; Jones, Abigal B.; Goba, Augustine; Momoh, Mambu; Fullah, Mohamed; Bornholdt, Zachary A.; Fusco, Marnie L.; Abelson, Dafna M.; Oda, Shunichiro; Brown, Bethany L.; Pham, Ha; Rowland, Megan M.; Agans, Krystle N.; Geisbert, Joan B.; Heinrich, Megan L.; Kulakosky, Peter C.; Shaffer, Jeffrey G.; Schieffelin, John S.; Kargbo, Brima; Gbetuwa, Momoh; Gevao, Sahr M.; Wilson, Russell B.; Saphire, Erica Ollmann; Pitts, Kelly R.; Khan, Sheik Humarr; Grant, Donald S.; Geisbert, Thomas W.; Branco, Luis M.; Garry, Robert F.

2016-01-01

Background. Ebola virus disease (EVD) is a severe viral illness caused by Ebola virus (EBOV). The 2013–2016 EVD outbreak in West Africa is the largest recorded, with >11 000 deaths. Development of the ReEBOV Antigen Rapid Test (ReEBOV RDT) was expedited to provide a point-of-care test for suspected EVD cases. Methods. Recombinant EBOV viral protein 40 antigen was used to derive polyclonal antibodies for RDT and enzyme-linked immunosorbent assay development. ReEBOV RDT limits of detection (LOD), specificity, and interference were analytically validated on the basis of Food and Drug Administration (FDA) guidance. Results. The ReEBOV RDT specificity estimate was 95% for donor serum panels and 97% for donor whole-blood specimens. The RDT demonstrated sensitivity to 3 species of Ebolavirus (Zaire ebolavirus, Sudan ebolavirus, and Bundibugyo ebolavirus) associated with human disease, with no cross-reactivity by pathogens associated with non-EBOV febrile illness, including malaria parasites. Interference testing exhibited no reactivity by medications in common use. The LOD for antigen was 4.7 ng/test in serum and 9.4 ng/test in whole blood. Quantitative reverse transcription–polymerase chain reaction testing of nonhuman primate samples determined the range to be equivalent to 3.0 × 105–9.0 × 108 genomes/mL. Conclusions. The analytical validation presented here contributed to the ReEBOV RDT being the first antigen-based assay to receive FDA and World Health Organization emergency use authorization for this EVD outbreak, in February 2015. PMID:27587634

Persistent ICT Malaria P.f/P.v Panmalarial and HRP2 Antigen Reactivity after Treatment of Plasmodium falciparum Malaria Is Associated with Gametocytemia and Results in False-Positive Diagnoses of Plasmodium vivax in Convalescence

PubMed Central

Tjitra, Emiliana; Suprianto, Sri; McBroom, James; Currie, Bart J.; Anstey, Nicholas M.

2001-01-01

A problem with rapid Plasmodium falciparum-specific antigen histidine-rich protein 2 (HRP2) detection tests for malaria is the persistence of antigen in blood after the disappearance of asexual-stage parasitemia and clinical symptoms, resulting in false-positive (FP) test results following treatment. The ICT P.f/P.v immunochromatographic test detects both HRP2 and a panmalarial antigen (PMA) found in both P. falciparum and Plasmodium vivax. To examine posttreatment antigen persistence with this test and whether persistent sexual-stage forms (gametocytes) are a cause of FP tests after treatment, we compared serial antigen test results with microscopy results from patients symptomatic with P. falciparum malaria in Indonesia for 28 days following treatment with chloroquine (CQ; n = 66), sulfadoxine-pyrimethamine (SP; n = 36), and artesunate plus sulfadoxine-pyrimethamine (ART + SP; n = 15). Persistent FP antigenemia following SP treatment occurred in 29% (HRP2) and 42% (PMA) of the patients on day 7 and in 10% (HRP2) and 23% (PMA) on day 14. The high rates of persistent HRP2 and PMA antigenemia following CQ and SP treatment were strongly associated with the presence of gametocytemia, with the proportion with gametocytes on day 7 posttreatment being significantly greater in those with FP results than in those with true-negative PMA and HRP2 results. Gametocyte frequency on day 14 post-SP treatment was also greater in those with FP PMA results. Following SP treatment, PMA persisted longer than HRP2, giving an FP diagnosis of P. vivax in up to 16% of patients on day 14, with all FP P. vivax diagnoses having gametocytemia. In contrast, PMA was rapidly cleared following ART + SP treatment in association with rapid clearance of gametocytemia. Gametocytes appear to be an important cause of persistent posttreatment panmalarial antigenemia in areas of endemicity and may also contribute in part to persistent HRP2 antigenemia following treatment. PMID:11230422

[Rapid laboratory detection of antigens of infective agents of infections and technical means for their realization].

PubMed

Kal'noĭ, S M

2003-01-01

A system of new accelerated and rapid methods for the detection of the antigens of the infective agents of plague, cholera, tularemia and brucellosis were developed on the basis of solid phase immunosuspension tests: the passive hemagglutination (PHA) test and the latex agglutination (LA) test. The immunological and physico-chemical properties of suspensions in the PHA and LA tests made it possible to use extraneous sources of energy (centrifugal acceleration and the electric field) to accelerate these tests. The results of the PHA and LA tests were registered with the use of a densitometer, model Ultrascan 2202, and a tester, model C 34014.2. To apply centrifugal acceleration and the electric field, a laboratory centrifuge and an electrophoretic microchamber were designed. Densitometry was carried out on modified plates and conductometry, with the use of modified electrodes. The time of obtaining the results of the PHA and LA tests was 15-30 minutes with the use of centrifugation and 2-5 minutes in the electric field, which made it possible to regard these tests as rapid.

Diagnostic Performance of Tuberculosis-Specific IgG Antibody Profiles in Patients with Presumptive Tuberculosis from Two Continents.

PubMed

Broger, Tobias; Basu Roy, Robindra; Filomena, Angela; Greef, Charles H; Rimmele, Stefanie; Havumaki, Joshua; Danks, David; Schneiderhan-Marra, Nicole; Gray, Christen M; Singh, Mahavir; Rosenkrands, Ida; Andersen, Peter; Husar, Gregory M; Joos, Thomas O; Gennaro, Maria L; Lochhead, Michael J; Denkinger, Claudia M; Perkins, Mark D

2017-04-01

Development of rapid diagnostic tests for tuberculosis is a global priority. A whole proteome screen identified Mycobacterium tuberculosis antigens associated with serological responses in tuberculosis patients. We used World Health Organization (WHO) target product profile (TPP) criteria for a detection test and triage test to evaluate these antigens. Consecutive patients presenting to microscopy centers and district hospitals in Peru and to outpatient clinics at a tuberculosis reference center in Vietnam were recruited. We tested blood samples from 755 HIV-uninfected adults with presumptive pulmonary tuberculosis to measure IgG antibody responses to 57 M. tuberculosis antigens using a field-based multiplexed serological assay and a 132-antigen bead-based reference assay. We evaluated single antigen performance and models of all possible 3-antigen combinations and multiantigen combinations. Three-antigen and multiantigen models performed similarly and were superior to single antigens. With specificity set at 90% for a detection test, the best sensitivity of a 3-antigen model was 35% (95% confidence interval [CI], 31-40). With sensitivity set at 85% for a triage test, the specificity of the best 3-antigen model was 34% (95% CI, 29-40). The reference assay also did not meet study targets. Antigen performance differed significantly between the study sites for 7/22 of the best-performing antigens. Although M. tuberculosis antigens were recognized by the IgG response during tuberculosis, no single antigen or multiantigen set performance approached WHO TPP criteria for clinical utility among HIV-uninfected adults with presumed tuberculosis in high-volume, urban settings in tuberculosis-endemic countries. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Development of an antigen-based rapid diagnostic test for the identification of blowfly (Calliphoridae) species of forensic significance.

PubMed

McDonagh, Laura; Thornton, Chris; Wallman, James F; Stevens, Jamie R

2009-06-01

In this study we examine the limitations of currently used sequence-based approaches to blowfly (Calliphoridae) identification and evaluate the utility of an immunological approach to discriminate between blowfly species of forensic importance. By investigating antigenic similarity and dissimilarity between the first instar larval stages of four forensically important blowfly species, we have been able to identify immunoreactive proteins of potential use in the development of species-specific immuno-diagnostic tests. Here we outline our protein-based approach to species determination, and describe how it may be adapted to develop rapid diagnostic assays for the 'on-site' identification of blowfly species.

Point-Counterpoint: A Nucleic Acid Amplification Test for Streptococcus pyogenes Should Replace Antigen Detection and Culture for Detection of Bacterial Pharyngitis.

PubMed

Pritt, Bobbi S; Patel, Robin; Kirn, Thomas J; Thomson, Richard B

2016-10-01

Nucleic acid amplification tests (NAATs) have frequently been the standard diagnostic approach when specific infectious agents are sought in a clinic specimen. They can be applied for specific agents such as S. pyogenes, or commercial multiplex NAATs for detection of a variety of pathogens in gastrointestinal, bloodstream, and respiratory infections may be used. NAATs are both rapid and sensitive. For many years, S. pyogenes testing algorithms used a rapid and specific group A streptococcal antigen test to screen throat specimens, followed, in some clinical settings, by a throat culture for S. pyogenes to increase the sensitivity of its detection. Now S. pyogenes NAATs are being used with increasing frequency. Given their accuracy, rapidity, and ease of use, should they replace antigen detection and culture for the detection of bacterial pharyngitis? Bobbi Pritt and Robin Patel of the Mayo Clinic, where S. pyogenes NAATs have been used for well over a decade with great success, will explain the advantages of this approach, while Richard (Tom) Thomson and Tom Kirn of the NorthShore University HealthSystem will discuss their concerns about this approach to diagnosing bacterial pharyngitis. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Single-step colony assay for screening antibody libraries.

PubMed

Kato, Mieko; Hanyu, Yoshiro

2017-08-10

We describe a method, single-step colony assay, for simple and rapid screening of single-chain Fv fragment (scFv) libraries. Colonies of Escherichia coli expressing the scFv library are formed on a hydrophilic filter that is positioned in contact with a membrane coated with an antigen. scFv expression is triggered upon treatment of colonies with an induction reagent, following which scFvs are secreted from the cells and diffused to the antigen-coated membrane. scFvs that exhibit binding affinity for the antigen are captured by the membrane-immobilized antigen. Lastly, detection of scFv binding of the antigen on the membrane allows identification of the clones on the filter that express antigen-specific scFvs. We tested this methodology by using an anti-rabbit IgG scFv, scFv(A10B), and a rat immune scFv library. Experiments conducted using scFv(A10B) revealed that this method improves scFv expression during the colony assay. By using our method to screen an immune library of 3×10 3 scFv clones, we established several clones exhibiting affinity for the antigen. Moreover, we tested 7 other antigens, including peptides, and successfully identified positive clones. We believe that this simple procedure and controlled scFv expression of the single-step colony assay could make the antibody screening both rapid and reliable and lead to successful isolation of positive clones from antibody libraries. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of a rapid immunochromatographic ODK-0901 test for detection of pneumococcal antigen in middle ear fluids and nasopharyngeal secretions.

PubMed

Hotomi, Muneki; Togawa, Akihisa; Takei, Shin; Sugita, Gen; Sugita, Rinya; Kono, Masamitsu; Fujimaki, Yutaka; Kamide, Yosuke; Uchizono, Akihiro; Kanesada, Keiko; Sawada, Shoichi; Okitsu, Naohiro; Tanaka, Yumi; Saijo, Yoko; Yamanaka, Noboru

2012-01-01

Since the incidence of penicillin-resistant Streptococcus pneumoniae has been increasing at an astonishing rate throughout the world, the need for accurate and rapid identification of pneumococci has become increasingly important to determine the appropriate antimicrobial treatment. We have evaluated an immunochromatographic test (ODK-0901) that detects pneumococcal antigens using 264 middle ear fluids (MEFs) and 268 nasopharyngeal secretions (NPSs). A sample was defined to contain S. pneumoniae when optochin and bile sensitive alpha hemolytic streptococcal colonies were isolated by culture. The sensitivity and specificity of the ODK-0901 test were 81.4% and 80.5%, respectively, for MEFs from patients with acute otitis media (AOM). In addition, the sensitivity and specificity were 75.2% and 88.8%, respectively, for NPSs from patients with acute rhinosinusitis. The ODK-0901 test may provide a rapid and highly sensitive evaluation of the presence of S. pneumoniae and thus may be a promising method of identifying pneumococci in MEFs and NPSs.

Evaluation of a Rapid Immunochromatographic ODK-0901 Test for Detection of Pneumococcal Antigen in Middle Ear Fluids and Nasopharyngeal Secretions

PubMed Central

Hotomi, Muneki; Togawa, Akihisa; Takei, Shin; Sugita, Gen; Sugita, Rinya; Kono, Masamitsu; Fujimaki, Yutaka; Kamide, Yosuke; Uchizono, Akihiro; Kanesada, Keiko; Sawada, Shoichi; Okitsu, Naohiro; Tanaka, Yumi; Saijo, Yoko; Yamanaka, Noboru

2012-01-01

Since the incidence of penicillin-resistant Streptococcus pneumoniae has been increasing at an astonishing rate throughout the world, the need for accurate and rapid identification of pneumococci has become increasingly important to determine the appropriate antimicrobial treatment. We have evaluated an immunochromatographic test (ODK-0901) that detects pneumococcal antigens using 264 middle ear fluids (MEFs) and 268 nasopharyngeal secretions (NPSs). A sample was defined to contain S. pneumoniae when optochin and bile sensitive alpha hemolytic streptococcal colonies were isolated by culture. The sensitivity and specificity of the ODK-0901 test were 81.4% and 80.5%, respectively, for MEFs from patients with acute otitis media (AOM). In addition, the sensitivity and specificity were 75.2% and 88.8%, respectively, for NPSs from patients with acute rhinosinusitis. The ODK-0901 test may provide a rapid and highly sensitive evaluation of the presence of S. pneumoniae and thus may be a promising method of identifying pneumococci in MEFs and NPSs. PMID:22448257

[Evaluation of the new ImmunoCard STAT!® CGE test for the diagnosis of Amebiasis].

PubMed

Formenti, F; Perandin, F; Bonafini, S; Degani, M; Bisoffi, Z

2015-08-01

For many years, microscopic examination of stool samples has been considered to be the "gold standard" for diagnosis of intestinal parasites although the Polymerase Chain Reaction (PCR) analysis is increasingly utilized due to its high accuracy. Recently, PCR has been approved by the World Health Organization as the current method of choice for the diagnosis of Entamoeba histolytica infection. In this study we evaluated a novel immunochromatographic antigen detection rapid test, ImmunoCardSTAT CGE (Meridian Bioscence, Milan, Italy), which has been proposed for the diagnosis of infections caused by Cryptosporidium parvum-Giardia intestinalis-Entamoeba histolytica. There is another rapid test with a similar name, the ImmunoCard STAT! Crypto/Giardia, but it is just for Cryptosporidium and Giardia. We aimed to compare E. histolytica results obtained from the rapid test with those of a rt-PCR for the detection of E. histolytica / E. dispar DNA. The new ImmunoCard rapid antigen detection test exhibited 88% sensitivity and 92% specificity (if assessed on rt-PCR negative samples) but showed a high proportion of cross-reaction between the pathogenic E. histolytica and the non pathogenic E. dispar.

Analytical Validation of the ReEBOV Antigen Rapid Test for Point-of-Care Diagnosis of Ebola Virus Infection.

PubMed

Cross, Robert W; Boisen, Matthew L; Millett, Molly M; Nelson, Diana S; Oottamasathien, Darin; Hartnett, Jessica N; Jones, Abigal B; Goba, Augustine; Momoh, Mambu; Fullah, Mohamed; Bornholdt, Zachary A; Fusco, Marnie L; Abelson, Dafna M; Oda, Shunichiro; Brown, Bethany L; Pham, Ha; Rowland, Megan M; Agans, Krystle N; Geisbert, Joan B; Heinrich, Megan L; Kulakosky, Peter C; Shaffer, Jeffrey G; Schieffelin, John S; Kargbo, Brima; Gbetuwa, Momoh; Gevao, Sahr M; Wilson, Russell B; Saphire, Erica Ollmann; Pitts, Kelly R; Khan, Sheik Humarr; Grant, Donald S; Geisbert, Thomas W; Branco, Luis M; Garry, Robert F

2016-10-15

Ebola virus disease (EVD) is a severe viral illness caused by Ebola virus (EBOV). The 2013-2016 EVD outbreak in West Africa is the largest recorded, with >11 000 deaths. Development of the ReEBOV Antigen Rapid Test (ReEBOV RDT) was expedited to provide a point-of-care test for suspected EVD cases. Recombinant EBOV viral protein 40 antigen was used to derive polyclonal antibodies for RDT and enzyme-linked immunosorbent assay development. ReEBOV RDT limits of detection (LOD), specificity, and interference were analytically validated on the basis of Food and Drug Administration (FDA) guidance. The ReEBOV RDT specificity estimate was 95% for donor serum panels and 97% for donor whole-blood specimens. The RDT demonstrated sensitivity to 3 species of Ebolavirus (Zaire ebolavirus, Sudan ebolavirus, and Bundibugyo ebolavirus) associated with human disease, with no cross-reactivity by pathogens associated with non-EBOV febrile illness, including malaria parasites. Interference testing exhibited no reactivity by medications in common use. The LOD for antigen was 4.7 ng/test in serum and 9.4 ng/test in whole blood. Quantitative reverse transcription-polymerase chain reaction testing of nonhuman primate samples determined the range to be equivalent to 3.0 × 10 5 -9.0 × 10 8 genomes/mL. The analytical validation presented here contributed to the ReEBOV RDT being the first antigen-based assay to receive FDA and World Health Organization emergency use authorization for this EVD outbreak, in February 2015. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Rapid assessment of the antigenic integrity of tetrameric HLA complexes by human monoclonal HLA antibodies.

PubMed

Eijsink, Chantal; Kester, Michel G D; Franke, Marry E I; Franken, Kees L M C; Heemskerk, Mirjam H M; Claas, Frans H J; Mulder, Arend

2006-08-31

The ability of tetrameric major histocompatibility complex (MHC) class I-peptide complexes (tetramers) to detect antigen-specific T lymphocyte responses has yielded significant information about the generation of in vivo immunity in numerous antigenic systems. Here we present a novel method for rapid validation of tetrameric HLA molecules based on the presence of allodeterminants. Human monoclonal antibodies (mAbs) recognizing polymorphic determinants on HLA class I were immobilized on polystyrene microparticles and used to probe the structural integrity of tetrameric HLA class I molecules by flow cytometry. A total of 22 tetramers, based on HLA-A1, A2, A3, A24, B7 and B8 were reactive with their counterpart mAbs, thus confirming their antigenic integrity. A positive outcome of this mAb test ensures that tetrameric HLA class I can be used with greater confidence in subsequent functional assays.

The dual role of paramagnetic particles for integrated lysis and measurement in a rapid immunoassay for intracellular proteins.

PubMed

Sharif, Elham; Kiely, Janice; Wraith, Patrick; Luxton, Richard

2013-05-01

A novel, integrated lysis and immunoassay methodology and system for intracellular protein measurement are described. The method uses paramagnetic particles both as a lysis agent and assay label resulting in a rapid test requiring minimal operator intervention, the test being homogeneous and completed in less than 10 min. A design study highlights the critical features of the magnetic detection system used to quantify the paramagnetic particles and a novel frequency-locked loop-based magnetometer is presented. A study of paramagnetic particle enhanced lysis demonstrates that the technique is more than twice as efficient at releasing intracellular protein as ultrasonic lysis alone. Results are presented for measurements of intracellular prostate specific antigen in an LNCAP cell line. This model was selected to demonstrate the rapidity and efficiency of intracellular protein quantification. It was shown that, on average, LNCAP cells contained 0.43 fg of prostate specific antigen. This system promises an attractive solution for applications that require a rapid determination of intracellular proteins.

Using the AD12-ICT rapid-format test to detect Wuchereria bancrofti circulating antigens in comparison to Og4C3-ELISA and nucleopore membrane filtration and microscopy techniques.

PubMed

El-Moamly, Amal Abdul-Rasheed; El-Sweify, Mohamed Aly; Hafez, Mohamad Abdul

2012-09-01

Lymphatic filariasis (LF) continues to be a major source of permanent disability and an impediment to socio-economic development in 73 countries where more than 1 billion people are at risk and over 120 millions are infected. The global drive to eliminate LF necessitates an increasing demand for valid, reliable and rapid diagnostic tests. This study aimed to assess the performance of the AD12 rapid format immunochromatographic test (ICT) to detect Wuchereria bancrofti circulating antigens, against the combined gold standard: TropBio Og4C3-ELISA (enzyme-linked immunosorbent assay) which detects circulating filarial antigen (CFA) and the nucleopore membrane filtration and microscopic examination. This prospective case-control study involved 647 asymptomatic migrant workers from filariasis-endemic countries. Of these specimens, 32 were positive for microfilaremia using the membrane filtration and microscopy, 142 positive by ELISA (of which 32 had microfilaremia), and 128 positive by the ICT (of which 31 had microfilaremia). The performance of the ICT was calculated against 32 true-positive and 90 true-negative cases. For the detection of CFA, the ICT had a sensitivity of 97% (95% confidence interval [CI] 91-103), specificity 100% (95% CI 100-100), Positive Predictive Value (PPV) 100% (95% CI 100-100), Negative Predictive Value (NPV) 99% (95% CI 97-101); and the total accuracy of the test was 99% (95% CI 98-101). The agreement between ICT and ELISA in detecting W. bancrofti antigens was excellent (kappa = 0.934; p = 0.000). In conclusion, the AD12-ICT test for the detection of W. bancrofti-CFA was sensitive and specific and comparable to the performance of ELISA. The ICT would be a useful additional test to facilitate the proposed strategies for control and elimination of LF. Because it is rapid, simple to perform, and does not require the use of special equipment, the ICT may be most appropriate in screening programs and in monitoring the possible risk of introducing the disease to the non-endemic countries.

Diagnosis and treatment of streptococcal pharyngitis.

PubMed

Choby, Beth A

2009-03-01

Common signs and symptoms of streptococcal pharyngitis include sore throat, temperature greater than 100.4 degrees F (38 degrees C), tonsillar exudates, and cervical adenopathy. Cough, coryza, and diarrhea are more common with viral pharyngitis. Available diagnostic tests include throat culture and rapid antigen detection testing. Throat culture is considered the diagnostic standard, although the sensitivity and specificity of rapid antigen detection testing have improved significantly. The modified Centor score can be used to help physicians decide which patients need no testing, throat culture/rapid antigen detection testing, or empiric antibiotic therapy. Penicillin (10 days of oral therapy or one injection of intramuscular benzathine penicillin) is the treatment of choice because of cost, narrow spectrum of activity, and effectiveness. Amoxicillin is equally effective and more palatable. Erythromycin and first-generation cephalosporins are options in patients with penicillin allergy. Increased group A beta-hemolytic streptococcus (GABHS) treatment failure with penicillin has been reported. Although current guidelines recommend first-generation cephalosporins for persons with penicillin allergy, some advocate the use of cephalosporins in all nonallergic patients because of better GABHS eradication and effectiveness against chronic GABHS carriage. Chronic GABHS colonization is common despite appropriate use of antibiotic therapy. Chronic carriers are at low risk of transmitting disease or developing invasive GABHS infections, and there is generally no need to treat carriers. Whether tonsillectomy or adenoidectomy decreases the incidence of GABHS pharyngitis is poorly understood. At this time, the benefits are too small to outweigh the associated costs and surgical risks.

Operational accuracy and comparative persistent antigenicity of HRP2 rapid diagnostic tests for Plasmodium falciparum malaria in a hyperendemic region of Uganda.

PubMed

Kyabayinze, Daniel J; Tibenderana, James K; Odong, George W; Rwakimari, John B; Counihan, Helen

2008-10-29

Parasite-based diagnosis of malaria by microscopy requires laboratory skills that are generally unavailable at peripheral health facilities. Rapid diagnostic tests (RDTs) require less expertise, but accuracy under operational conditions has not been fully evaluated in Uganda. There are also concerns about RDTs that use the antigen histidine-rich protein 2 (HRP2) to detect Plasmodium falciparum, because this antigen can persist after effective treatment, giving false positive test results in the absence of infection. An assessment of the accuracy of Malaria Pf immuno-chromatographic test (ICT) and description of persistent antigenicity of HRP2 RDTs was undertaken in a hyperendemic area of Uganda. Using a cross-sectional design, a total of 357 febrile patients of all ages were tested using ICT, and compared to microscopy as the gold standard reference. Two independent RDT readings were used to assess accuracy and inter-observer reliability. With a longitudinal design to describe persistent antigenicity of ICT and Paracheck, 224 children aged 6-59 months were followed up at 7-day intervals until the HRP2 antigens where undetectable by the RDTs. Of the 357 patients tested during the cross-sectional component, 40% (139) had positive blood smears for asexual forms of P. falciparum. ICT had an overall sensitivity of 98%, a specificity of 72%, a negative predictive value (NPV) of 98% and a positive predictive value (PPV) of 69%. ICT showed a high inter-observer reliability under operational conditions, with 95% of readings having assigned the same results (kappa statistics 0.921, p < 0.001). In children followed up after successful antimalaria treatment, the mean duration of persistent antigenicity was 32 days, and this duration varied significantly depending on pre-treatment parasitaemia. In patients with parasite density >50,000/microl, the mean duration of persistent antigenicity was 37 days compared to 26 days for parasitaemia less than 1,000/microl (log rank 21.9, p < 0.001). ICT is an accurate and appropriate test for operational use as a diagnostic tool where microscopy is unavailable. However, persistent antigenicity reduces the accuracy of this and other HRP2-based RDTs. The low specificity continues to be of concern, especially in children below five years of age. These pose limitations that need consideration, such as their use for diagnosis of patients returning with symptoms within two to four weeks of treatment. Good clinical skills are essential to interpret test results.

Operational accuracy and comparative persistent antigenicity of HRP2 rapid diagnostic tests for Plasmodium falciparum malaria in a hyperendemic region of Uganda

PubMed Central

Kyabayinze, Daniel J; Tibenderana, James K; Odong, George W; Rwakimari, John B; Counihan, Helen

2008-01-01

Background Parasite-based diagnosis of malaria by microscopy requires laboratory skills that are generally unavailable at peripheral health facilities. Rapid diagnostic tests (RDTs) require less expertise, but accuracy under operational conditions has not been fully evaluated in Uganda. There are also concerns about RDTs that use the antigen histidine-rich protein 2 (HRP2) to detect Plasmodium falciparum, because this antigen can persist after effective treatment, giving false positive test results in the absence of infection. An assessment of the accuracy of Malaria Pf™ immuno-chromatographic test (ICT) and description of persistent antigenicity of HRP2 RDTs was undertaken in a hyperendemic area of Uganda. Methods Using a cross-sectional design, a total of 357 febrile patients of all ages were tested using ICT, and compared to microscopy as the gold standard reference. Two independent RDT readings were used to assess accuracy and inter-observer reliability. With a longitudinal design to describe persistent antigenicity of ICT and Paracheck, 224 children aged 6–59 months were followed up at 7-day intervals until the HRP2 antigens where undetectable by the RDTs. Results Of the 357 patients tested during the cross-sectional component, 40% (139) had positive blood smears for asexual forms of P. falciparum. ICT had an overall sensitivity of 98%, a specificity of 72%, a negative predictive value (NPV) of 98% and a positive predictive value (PPV) of 69%. ICT showed a high inter-observer reliability under operational conditions, with 95% of readings having assigned the same results (kappa statistics 0.921, p < 0.001). In children followed up after successful antimalaria treatment, the mean duration of persistent antigenicity was 32 days, and this duration varied significantly depending on pre-treatment parasitaemia. In patients with parasite density >50,000/μl, the mean duration of persistent antigenicity was 37 days compared to 26 days for parasitaemia less than 1,000/μl (log rank 21.9, p < 0.001). Conclusion ICT is an accurate and appropriate test for operational use as a diagnostic tool where microscopy is unavailable. However, persistent antigenicity reduces the accuracy of this and other HRP2-based RDTs. The low specificity continues to be of concern, especially in children below five years of age. These pose limitations that need consideration, such as their use for diagnosis of patients returning with symptoms within two to four weeks of treatment. Good clinical skills are essential to interpret test results. PMID:18959777

Development and Comparative Evaluation of a Plate Enzyme-Linked Immunosorbent Assay Based on Recombinant Outer Membrane Antigens Omp28 and Omp31 for Diagnosis of Human Brucellosis

PubMed Central

Tiwari, Sapana; Kumar, Ashu; Mangalgi, Smita; Rathod, Vedika; Prakash, Archana; Barua, Anita; Arora, Sonia; Sathyaseelan, Kannusamy

2013-01-01

Brucellosis is an important zoonotic infectious disease of humans and livestock with worldwide distribution and is caused by bacteria of the genus Brucella. The diagnosis of brucellosis always requires laboratory confirmation by either isolation of pathogens or detection of specific antibodies. The conventional serological tests available for the diagnosis of brucellosis are less specific and show cross-reactivity with other closely related organisms. These tests also necessitate the handling of Brucella species for antigen preparation. Therefore, there is a need to develop reliable, rapid, and user-friendly systems for disease diagnosis and alternatives to vaccine approaches. Keeping in mind the importance of brucellosis as an emerging infection and the prevalence in India, we carried out the present study to compare the recombinant antigens with the native antigens (cell envelope and sonicated antigen) of Brucella for diagnosis of human brucellosis by an indirect plate enzyme-linked immunosorbent assay (ELISA). Recombinant outer membrane protein 28 (rOmp28) and rOmp31 antigens were cloned, expressed, and purified in the bacterial expression system, and the purified proteins were used as antigens. Indirect plate ELISAs were then performed and standardized for comparison of the reactivities of recombinant and native antigens against the 433 clinical samples submitted for brucellosis testing, 15 culture-positive samples, and 20 healthy donor samples. The samples were separated into four groups based on their positivity to rose bengal plate agglutination tests (RBPTs), standard tube agglutination tests (STATs), and 2-mercaptoethanol (2ME) tests. The sensitivities and specificities of all the antigens were calculated, and the rOmp28 antigen was found to be more suitable for the clinical diagnosis of brucellosis than the rOmp31 antigen and native antigens. The rOmp28-based ELISA showed a very high degree of agreement with the conventional agglutination tests and promising results for further use in clinical screening and serodiagnosis of human brucellosis. PMID:23761658

[Clinical microbiology laboratory and imported parasitic diseases].

PubMed

Martín-Rabadán, Pablo; Martínez-Ruiz, Rocío; Cuadros, Juan; Cañavate, Carmen

2010-12-01

Imported parasitosis represents an increasingly frequent diagnostic challenge for microbiology laboratories. A surge in immigration and international travel has led to a rise in the number of imported cases of parasitosis, and this trend is expected to continue in the future. The present article addresses this challenge by reviewing recommended diagnostic approaches and tests. Currently, microscopy is always recommended when analysing blood samples for parasites. If malaria is suspected, rapid antigen testing (including at least HRP2 antigen) should also be performed. The work-up for suspected leishmaniasis should include serology, culture, and in selected cases detection of antigen in urine. In suspected Chagas disease, two different serological tests should be performed. PCR for blood protozoa is highly sensitive, although it cannot be used to rule out Chagas disease, since this condition may be present without parasitemia. Accurate diagnosis of intestinal amebiasis usually requires PCR or antigen detection tests. In helminthiasis, traditional microscopy may need to be complemented with other tests, such as agar plate culture for strongyloidiasis, Og4C3 antigen detection for bancroftian filariasis, and antibody detection test for filariasis and schistosomiasis. Copyright © 2010 Elsevier España, S.L. All rights reserved.

Development of a sensitive and specific indirect enzyme-linked immunosorbent assay based on a baculovirus recombinant antigen for detection of specific antibodies against Ehrlichia canis.

PubMed

López, Lissett; Venteo, Angel; Aguirre, Enara; García, Marga; Rodríguez, Majosé; Amusátegui, Inmaculada; Tesouro, Miguel A; Vela, Carmen; Sainz, Angel; Rueda, Paloma

2007-11-01

An indirect enzyme-linked immunosorbent assay (ELISA) based on baculovirus recombinant P30 protein of Ehrlichia canis and the 1BH4 anticanine IgG monoclonal antibody was developed and evaluated by examining a panel of 98 positive and 157 negative sera using the indirect fluorescent antibody (IFA) test as the reference technique. The P30-based ELISA appeared to be sensitive and specific (77.55% and 95.54%, respectively) when qualitative results (positive/negative) were compared with those of the IFA test; the coefficient of correlation (R) between the 2 tests was 0.833. Furthermore, it was possible to establish a mathematical formula for use in comparing the results of both techniques. These results indicate that recombinant P30 antigen-based ELISA is a suitable alternative of the IFA test for simple, consistent, and rapid serodiagnosis of canine ehrlichiosis. Moreover, the use of this recombinant protein as antigen offers a great advantage for antigen preparation in comparison with other techniques in which the whole E. canis organism is used as antigen.

[Methods of testing inactivated antirabies vaccines].

PubMed

Nedosekov, V V; Vishniakov, I F; Gruzdev, K N

2001-01-01

Methods for evaluating the potency of inactivated rabies vaccines are reviewed. Shortcomings of the traditional NIH method and advantages of modern rapid immunological in vitro methods (antibody binding test, radial immunodiffusion test, enzyme linked immunoadsorbent assay) for estimation of antigenic activity of vaccines are discussed.

Rapid desensitization induces internalization of antigen-specific IgE on mouse mast cells.

PubMed

Oka, Tatsuya; Rios, Eon J; Tsai, Mindy; Kalesnikoff, Janet; Galli, Stephen J

2013-10-01

Rapid desensitization transiently prevents severe allergic reactions, allowing administration of life-saving therapies in previously sensitized patients. However, the mechanisms underlying successful rapid desensitization are not fully understood. We sought to investigate whether the mast cell (MC) is an important target of rapid desensitization in mice sensitized to exhibit IgE-dependent passive systemic anaphylaxis in vivo and to investigate the antigen specificity and underlying mechanisms of rapid desensitization in our mouse model. C57BL/6 mice (in vivo) or primary isolated C57BL/6 mouse peritoneal mast cells (PMCs; in vitro) were passively sensitized with antigen-specific anti-2,4-dinitrophenyl IgE, anti-ovalbumin IgE, or both. MCs were exposed over a short period of time to increasing amounts of antigen (2,4-dinitrophenyl-human serum albumin or ovalbumin) in the presence of extracellular calcium in vitro or by means of intravenous administration to sensitized mice in vivo before challenging the mice with or exposing the PMCs to optimal amounts of specific or irrelevant antigen. Rapidly exposing mice or PMCs to progressively increasing amounts of specific antigen inhibited the development of antigen-induced hypothermia in sensitized mice in vivo and inhibited antigen-induced PMC degranulation and prostaglandin D2 synthesis in vitro. Such MC hyporesponsiveness was induced antigen-specifically and was associated with a significant reduction in antigen-specific IgE levels on MC surfaces. Rapidly exposing MCs to progressively increasing amounts of antigen can both enhance the internalization of antigen-specific IgE on the MC surface and also desensitize these cells in an antigen-specific manner in vivo and in vitro. Copyright © 2013 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Rapid desensitization induces internalization of antigen-specific IgE on mouse mast cells

PubMed Central

Oka, Tatsuya; Rios, Eon J.; Tsai, Mindy; Kalesnikoff, Janet; Galli, Stephen J.

2013-01-01

Background Rapid desensitization transiently prevents severe allergic reactions, allowing administration of life-saving therapies in previously sensitized patients. However, the mechanisms underlying successful rapid desensitization are not fully understood. Objectives We sought to investigate whether the mast cell (MC) is an important target of rapid desensitization in mice sensitized to exhibit IgE-dependent passive systemic anaphylaxis in vivo and to investigate the antigen specificity and underlying mechanisms of rapid desensitization in our mouse model. Methods C57BL/6 mice (in vivo) or primary isolated C57BL/6 mouse peritoneal mast cells (PMCs; in vitro) were passively sensitized with antigen-specific anti–2,4-dinitrophenyl IgE, anti-ovalbumin IgE, or both. MCs were exposed over a short period of time to increasing amounts of antigen (2,4-dinitrophenyl–human serum albumin or ovalbumin) in the presence of extracellular calcium in vitro or by means of intravenous administration to sensitized mice in vivo before challenging the mice with or exposing the PMCs to optimal amounts of specific or irrelevant antigen. Results Rapidly exposing mice or PMCs to progressively increasing amounts of specific antigen inhibited the development of antigen-induced hypothermia in sensitized mice in vivo and inhibited antigen-induced PMC degranulation and prostaglandin D2 synthesis in vitro. Such MC hyporesponsiveness was induced antigen-specifically and was associated with a significant reduction in antigen-specific IgE levels on MC surfaces. Conclusions Rapidly exposing MCs to progressively increasing amounts of antigen can both enhance the internalization of antigen-specific IgE on the MC surface and also desensitize these cells in an antigen-specific manner in vivo and in vitro. PMID:23810240

Detecting West Nile virus in owls and raptors by an antigen-capture assay.

PubMed

Gancz, Ady Y; Campbell, Douglas G; Barker, Ian K; Lindsay, Robbin; Hunter, Bruce

2004-12-01

We evaluated a rapid antigen-capture assay (VecTest) for detection of West Nile virus in oropharyngeal and cloacal swabs, collected at necropsy from owls (N = 93) and raptors (N = 27). Sensitivity was 93.5%-95.2% for northern owl species but <42.9% for all other species. Specificity was 100% for owls and 85.7% for raptors.

Development of solid - based paper strips for rapid diagnosis of Pseudorabies infection.

PubMed

Joon Tam, Yew; Mohd Lila, Mohd Azmi; Bahaman, Abdul Rani

2004-12-01

Pseudorabies (Aujeszky's disease) is an economically significant disease of swine known to cause central nervous disorders, respiratory disease, reproductive failure and mortality in infected pigs. In attempts to eradicate the disease from becoming endemic, early detection is important to prevent further economic losses and to allow for detection and removal of infected pigs in domestic herds. Thus, a rapid and sensitive technique is necessary for the detection of the virus. For rapid and simple examination, an immuno - chromatographic lateral - flow assay system based on immunologic recognition of specific pseudorabies virus antigen was developed by utilising, as signal generator, colloidal gold conjugated to secondary antibody to detect primary or sample antibody in the sera of pseudorabies infected animals. The pseudorabies virus used as a capture antigen in the test strip was first cultivated in VERO cell culture and then purified by sucrose gradient separation to produce the viral protein concentration of 3.8 mg/ml. The standard pseudorabies antigens reacted well with the hyperimmune serum (HIS). The antibody detection system is basically composed of colloidal gold - labelled antibodies fixed on a conjugate pad, and the complementary pseudorabies antigen immobilised onto a nitrocellulose membrane forming capture zone. If the target antibody is present in a specimen, the colloidal gold-labelled antibody will form a complex with the antibody sample. Subsequently, the formed complex will migrate to the capture zone and is then bound to the solid phase via antigen - antibody interaction. As a result, a signal marker is generated by the accumulation of colloidal gold for detection confirmation. The results obtained demonstrated that the optimum combination of pseudorabies antigen needed as the capture reagent and gold conjugate as secondary antibody recognition marker was at a concentration of 0.38mg/ml and at 1:10 dilution factor respectively. The sensitivity of the solid - based test strip towards pseudorabies antibodies was high with a detection limit of 1 to 10,000 - dilution factor. The specificity of the assay was 100% with no cross - reaction being observed with other sera or antibodies. Accurate reading time needed for confirmation of the assay can be completed in 5 min with a whole blood sample of 25 microl. The colloidal gold - labelled antibody is stable at room temperature for 6 months or more (data not shown). Findings from this study indicated that the solid - based test strip assay system provided high sensitivity and specificity for the detection of pseudorabies at low levels of antibody concentration. The assay was rapid, simple, cheap, and does not require any sophisticated equipment. Thus, the solid based test strip will be a useful serological screening technique or for rapid diagnosis of an infectious disease in target populations of animals characterised by heterogeneous antibody responses.

A novel one-step Helicobacter pylori saliva antigen test.

PubMed

Yang, Bi-Ling; Yeh, Chun; Kwong, Wei-Gang; Lee, Shou-Dong

2015-02-01

A rapid, reliable, and sufficiently accurate test for diagnosing Helicobacter pylori infection is required for screening dyspeptic patients before a referral for endoscopy. The purpose of this article is two-fold: first, to evaluate the accuracy of a one-step H. pylori saliva antigen (HPS) test; and second, to compare noninvasive and invasive H. pylori tests in Taiwanese population. A total of 104 consecutive dyspeptic patients admitted for gastroenterology into the outpatient department underwent a one-step HPS test, rapid urease test, histology, and (13)C-urea breath test (13)C-UBT (proto C-13 urea kit). The accuracy of the HPS test was compared with a gold standard defined by at least two positive H. pylori test results from three H. pylori tests (histology, rapid urease test, and (13)C-UBT). The 104 patients eligible for analysis (mean age: 58 years, range 22-87 years), 21 (20%) were gold standard positive. Among them, the positive of the one-step H. pylori saliva Ag test, rapid urease test, (13)C-UBT, histology were (52; 50%), (17; 16%), (27; 25%) and (22; 21%) respectively. The sensitivity and specificity of the HPS tests, rapid urease test, (13)C-UBTs, and histology were 71.43% and 55.42%, 76.19% and 98.80%, 100% and 92.77%, and 85.71% and 95.18%, respectively, relative to the gold standard. The one-step HPS test exhibited a sensitivity of 71.43%, nearly equivalent to that of the rapid urea test. The one-step HPS test exhibited a high sensitivity and low specificity compared with the other tests, indicating that it is not sufficiently accurate for use in a clinical setting for diagnosing H. pylori infection. However, the test is simple to use (requiring only a saliva sample), inexpensive, and noninvasive in its application, and thus appealing for use in population-based prevalence surveys of the epidemiology of H. pylori infection. Copyright © 2014. Published by Elsevier Taiwan.

Laboratory and Field Evaluation of a New Rapid Test for Detecting Wuchereria bancrofti Antigen in Human Blood

PubMed Central

Weil, Gary J.; Curtis, Kurt C.; Fakoli, Lawrence; Fischer, Kerstin; Gankpala, Lincoln; Lammie, Patrick J.; Majewski, Andrew C.; Pelletreau, Sonia; Won, Kimberly Y.; Bolay, Fatorma K.; Fischer, Peter U.

2013-01-01

Global Program to Eliminate Lymphatic Filariasis (GPELF) guidelines call for using filarial antigen testing to identify endemic areas that require mass drug administration (MDA) and for post-MDA surveillance. We compared a new filarial antigen test (the Alere Filariasis Test Strip) with the reference BinaxNOW Filariasis card test that has been used by the GPELF for more than 10 years. Laboratory testing of 227 archived serum or plasma samples showed that the two tests had similar high rates of sensitivity and specificity and > 99% agreement. However, the test strip detected 26.5% more people with filarial antigenemia (124/503 versus 98/503) and had better test result stability than the card test in a field study conducted in a filariasis-endemic area in Liberia. Based on its increased sensitivity and other practical advantages, we believe that the test strip represents a major step forward that will be welcomed by the GPELF and the filariasis research community. PMID:23690552

Performance of pfHRP2 versus pLDH antigen rapid diagnostic tests for the detection of Plasmodium falciparum: a systematic review and meta-analysis.

PubMed

Li, Bo; Sun, Zhiqiang; Li, Xiaohan; Li, Xiaoxi; Wang, Han; Chen, Weijiao; Chen, Peng; Qiao, Mengran; Mao, Yuanli

2017-04-01

There have been many inconsistent reports about the performance of histidine-rich protein 2 (HRP2) and lactate dehydrogenase (LDH) antigens as rapid diagnostic tests (RDTs) for the diagnosis of past Plasmodium falciparum infections. This meta-analysis was performed to determine the performance of pfHRP2 versus pLDH antigen RDTs in the detection of P. falciparum . After a systematic review of related studies, Meta-DiSc 1.4 software was used to calculate the pooled sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic odds ratio (DOR). Forest plots and summary receiver operating characteristic curve (SROC) analysis were used to summarize the overall test performance. Fourteen studies which met the inclusion criteria were included in the meta-analysis. The summary performances for pfHRP2- and pLDH-based tests in the diagnosis of P. falciparum infections were as follows: pooled sensitivity, 96.3% (95.8-96.7%) vs. 82.6% (81.7-83.5%); specificity, 86.1% (85.3-86.8%) vs. 95.9% (95.4-96.3%); diagnostic odds ratio (DOR), 243.31 (97.679-606.08) vs. 230.59 (114.98-462.42); and area under ROCs, 0.9822 versus 0.9849 (all p < 0.001). The two RDTs performed satisfactorily for the diagnosis of P. falciparum , but the pLDH tests had higher specificity, whereas the pfHRP2 tests had better sensitivity. The pfHRP2 tests had slightly greater accuracy compared to the pLDH tests. A combination of both antigens might be a more reliable approach for the diagnosis of malaria.

High prevalence of HIV p24 antigen among HIV antibody negative prospective blood donors in Ile-Ife, Nigeria.

PubMed

Japhet, Margaret Oluwatoyin; Adewumi, Moses Olubusuyi; Adesina, Olufisayo Adeyemi; Donbraye, Emmanuel

2016-01-01

Blood transfusion service centers in Nigeria screen donated blood for markers of HIV infection using antibody- (Ab) based rapid test and in some centers, positives are re-tested using Ab-based ELISA. Paucity of data exists on p24 antigen prevalence among HIV Ab-negative donors in Nigeria. This study aims at detecting HIV p24 antigen among prospective blood donors in Osun State, Nigeria. Prospective blood donors negative for HIV antibodies using Determine test kit were re-tested using BIORAD GENSCREEN Ultra Ag-Ab ELISA kit, a fourth-generation ELISA kit that detects HIV antibodies/p24 antigen. Of the 169 HIV Ab-negative prospective donors, 10 (5.9%) were positive for HIV p24 antigen and 70% (7/10) of them were in the age range 18-30 years. Results of this study show that blood transfusion is still one of the major routes of HIV transmission in Nigeria and a higher proportion is among youth. Inclusion of p24 antigen testing into the blood donor screening will help reduce transfusion associated HIV in Nigeria if Nucleic Acid Testing (NAT) of all blood donor samples is not affordable; also, HIV enlightenment programs tailored toward youth may help reduce this rate among donors since more young people donate blood in low/middle-income countries than in high-income countries.

A comparative laboratory diagnosis of malaria: microscopy versus rapid diagnostic test kits.

PubMed

Azikiwe, C C A; Ifezulike, C C; Siminialayi, I M; Amazu, L U; Enye, J C; Nwakwunite, O E

2012-04-01

To compare the two methods of rapid diagnostic tests (RDTs) and microscopy in the diagnosis of malaria. RDTs and microscopy were carried out to diagnose malaria. Percentage malaria parasitaemia was calculated on thin films and all non-acute cases of plasmodiasis with less than 0.001% malaria parasitaemia were regarded as negative. Results were simply presented as percentage positive of the total number of patients under study. The results of RDTs were compared to those of microscopy while those of RDTs based on antigen were compared to those of RDTs based on antibody. Patients' follow-up was made for all cases. All the 200 patients under present study tested positive to RDTs based on malaria antibodies (serum) method (100%). 128 out of 200 tested positive to RDTs based on malaria antigen (whole blood) method (64%), while 118 out of 200 patients under present study tested positive to visual microscopy of Lieshman and diluted Giemsa (59%). All patients that tested positive to microscopy also tested positive to RDTs based on antigen. All patients on the second day of follow-up were non-febrile and had antimalaria drugs. We conclude based on the present study that the RDTs based on malaria antigen (whole blood) method is as specific as the traditional microscopy and even appears more sensitive than microscopy. The RDTs based on antibody (serum) method is unspecific thus it should not be encouraged. It is most likely that Africa being an endemic region, formation of certain levels of malaria antibody may not be uncommon. The present study also supports the opinion that a good number of febrile cases is not due to malaria. We support WHO's report on cost effectiveness of RDTs but, recommend that only the antigen based method should possibly, be adopted in Africa and other malaria endemic regions of the world.

Novel Antigen Identification Method for Discovery of Protective Malaria Antigens by Rapid Testing of DNA Vaccines Encoding Exons from the Parasite Genome

DTIC Science & Technology

2004-03-01

EAA21673 1,443 — — Xeroderma pigmentosum G N&I region, helix-hairpin-helix class P.f., P.k., P.b., P.v. PY02286 EAA21722 696 — — Hypothetical protein...ND PY01828 Gene gun 0.1 2,560 640 Pos IM 0.1 Neg Neg ND CSP Gene gun 0.1 2,560 Neg Neg IM 2.7* 2,560 Neg ND a Parasite burden in liver is in...negative; Pos , positive; ND, not done. c Sera tested at a single dilution (1:80). VOL. 72, 2004 DISCOVERY OF PROTECTIVE MALARIA PARASITE ANTIGENS 1599

Development of Prototype Filovirus Recombinant Antigen Immunoassays

PubMed Central

Boisen, Matt L.; Oottamasathien, Darin; Jones, Abigail B.; Millett, Molly M.; Nelson, Diana S.; Bornholdt, Zachary A.; Fusco, Marnie L.; Abelson, Dafna M.; Oda, Shun-ichiro; Hartnett, Jessica N.; Rowland, Megan M.; Heinrich, Megan L.; Akdag, Marjan; Goba, Augustine; Momoh, Mambu; Fullah, Mohammed; Baimba, Francis; Gbakie, Michael; Safa, Sadiki; Fonnie, Richard; Kanneh, Lansana; Cross, Robert W.; Geisbert, Joan B.; Geisbert, Thomas W.; Kulakosky, Peter C.; Grant, Donald S.; Shaffer, Jeffery G.; Schieffelin, John S.; Wilson, Russell B.; Saphire, Erica Ollmann; Branco, Luis M.; Garry, Robert F.; Khan, S. Humarr; Pitts, Kelly R.

2015-01-01

Background. Throughout the 2014–2015 Ebola outbreak in West Africa, major gaps were exposed in the availability of validated rapid diagnostic platforms, protective vaccines, and effective therapeutic agents. These gaps potentiated the development of prototype rapid lateral flow immunodiagnostic (LFI) assays that are true point-of-contact platforms, for the detection of active Ebola infections in small blood samples. Methods. Recombinant Ebola and Marburg virus matrix VP40 and glycoprotein (GP) antigens were used to derive a panel of monoclonal and polyclonal antibodies. Antibodies were tested using a multivariate approach to identify antibody-antigen combinations suitable for enzyme-linked immunosorbent assay (ELISA) and LFI assay development. Results. Polyclonal antibodies generated in goats were superior reagents for capture and detection of recombinant VP40 in test sample matrices. These antibodies were optimized for use in antigen-capture ELISA and LFI assay platforms. Prototype immunoglobulin M (IgM)/immunoglobulin G (IgG) ELISAs were similarly developed that specifically detect Ebola virus–specific antibodies in the serum of experimentally infected nonhuman primates and in blood samples obtained from patients with Ebola from Sierra Leone. Conclusions. The prototype recombinant Ebola LFI assays developed in these studies have sensitivities that are useful for clinical diagnosis of acute ebolavirus infections. The antigen-capture and IgM/IgG ELISAs provide additional confirmatory assay platforms for detecting VP40 and other ebolavirus-specific immunoglobulins. PMID:26232440

Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test.

PubMed

Morioka, Ayako; Shimazaki, Yoko; Uchiyama, Mariko; Suzuki, Shoko

2016-05-03

We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2.

Serotyping reanalysis of unserotypable Actinobacillus pleuropneumoniae isolates by agar gel diffusion test

PubMed Central

MORIOKA, Ayako; SHIMAZAKI, Yoko; UCHIYAMA, Mariko; SUZUKI, Shoko

2016-01-01

We observed increasing unserotypable (UT) Actinobacillus pleuropneumoniae isolates using agar gel diffusion (AGD) test. To reanalyze their serovar, we performed rapid slide agglutination (RSA) test and multiplex PCR for 47 UT isolates. Of these, 25 were serovar 1 (UT-serovar 1), 20 were serovar 2 (UT-serovar 2) and 2 were serovar 15 (UT-serovar 15). We examined serotyping antigen extraction temperature to determine heat influence. UT-serovar 1 and 15 were influenced by heat, because their precipitation lines were observed in the case of low antigen extraction temperature. To investigate the relationship between antigenicity and genotype, we performed pulsed-field gel electrophoresis (PFGE) analysis using UT-serovar 2 and 15. The predominant PFGE pattern of UT-serovar 2 was identical to that of serovar 2. PMID:26726101

Evaluation of SD BIOLINE H. pylori Ag rapid test against double ELISA with SD H. pylori Ag ELISA and EZ-STEP H. pylori Ag ELISA tests.

PubMed

Negash, Markos; Kassu, Afework; Amare, Bemnet; Yismaw, Gizachew; Moges, Beyene

2018-01-01

Helicobacter pylori antibody titters fall very slowly even after successful treatment. Therefore, tests detecting H. pylori antibody lack specificity and sensitivity. On the other hand, H. pylori stool antigen tests are reported as an alternative assay because of their reliability and simplicity. However, the comparative performance of H. pylori stool antigen tests for detecting the presence of the bacterium in clinical specimens in the study area is not assessed. Therefore, in this study we evaluated the performance of SD BIOLINE H. pylori Ag rapid test with reference to the commercially available EZ- STEP ELISA and SD BIOLINE H. pylori Ag ELISA tests. Stool samples were collected to analyse the diagnostic performance of SD BIOLINE H. pylori Ag rapid test kit using SD H. pylori Ag ELISA kit and EZ- STEP ELISA tests as a gold standard. Serum samples were also collected from each patient to test for the presence of H. pylori antibodies using dBest H. pylori Test Disk. Sensitivity, specificity, predictive values and kappa value are assessed. P values < 0.05 were taken statistically significant. Stool and serum samples were collected from 201 dyspeptic patients and analysed. The sensitivity, specificity, positive and negative predictive values of the SD BIOLINE H. pylori Ag rapid test were: 95.6% (95% CI, 88.8-98.8), 92.5% (95%CI, 89-94.1%), 86.7% (95% CI, 80.5-89.6), and 97.6% (95% CI, 993.9-99.3) respectively. The performance of SD BIOLINE H. pylori Ag rapid test was better than the currently available antibody test in study area. Therefore, the SD BIOLINE Ag rapid stool test could replace and be used to diagnose active H. pylori infection before the commencement of therapy among dyspeptic patients.

Detecting West Nile Virus in Owls and Raptors by an Antigen-capture Assay

PubMed Central

Campbell, Douglas G.; Barker, Ian K.; Lindsay, Robbin; Hunter, Bruce

2004-01-01

We evaluated a rapid antigen-capture assay (VecTest) for detection of West Nile virus in oropharyngeal and cloacal swabs, collected at necropsy from owls (N = 93) and raptors (N = 27). Sensitivity was 93.5%–95.2% for northern owl species but <42.9% for all other species. Specificity was 100% for owls and 85.7% for raptors. PMID:15663862

Potential for rapid antibody detection to identify tuberculous cattle with non-reactive tuberculin skin test results.

PubMed

Waters, W Ray; Vordermeier, H Martin; Rhodes, Shelley; Khatri, Bhagwati; Palmer, Mitchell V; Maggioli, Mayara F; Thacker, Tyler C; Nelson, Jeffrey T; Thomsen, Bruce V; Robbe-Austerman, Suelee; Bravo Garcia, Doris M; Schoenbaum, Mark A; Camacho, Mark S; Ray, Jean S; Esfandiari, Javan; Lambotte, Paul; Greenwald, Rena; Grandison, Adrian; Sikar-Gang, Alina; Lyashchenko, Konstantin P

2017-06-07

Bovine tuberculosis (TB) control programs generally rely on the tuberculin skin test (TST) for ante-mortem detection of Mycobacterium bovis-infected cattle. Present findings demonstrate that a rapid antibody test based on Dual-Path Platform (DPP ® ) technology, when applied 1-3 weeks after TST, detected 9 of 11 and 34 of 52 TST non-reactive yet M. bovis-infected cattle from the US and GB, respectively. The specificity of the assay ranged from 98.9% (n = 92, US) to 96.0% (n = 50, GB) with samples from TB-free herds. Multi-antigen print immunoassay (MAPIA) revealed the presence of antibodies to multiple antigens of M. bovis in sera from TST non-reactors diagnosed with TB. Thus, use of serologic assays in series with TST can identify a significant number of TST non-reactive tuberculous cattle for more efficient removal from TB-affected herds.

DETECTION OF CANINE PARVOVIRUS ANTIGEN IN DOGS IN KUMASI, GHANA

PubMed Central

Folitse, R. D; Kodie, D.O; Amemor, E.; Dei, D.; Tasiame, W.; Burimuah, V.; Emikpe, B.O

2018-01-01

Background: Canine Parvovirus (CPV) in dogs has been documented in many countries. However, evidence of the infection is scanty in Ghana. This study was conducted to detect canine parvovirus antigen in dogs presented with diarrhoea to the Government Veterinary Clinic in Kumasi, Ghana. Materials and Methods: Faecal samples from 72 dogs presented with diarrhoea were tested for the presence of canine parvovirus antigen using commercially available rapid test kit (BIT® Rapid Colour Canine Parvovirus Ag Test Kit, BIOINDIST Co. Ltd, Korea) based on the principle of immunochromatography. Influence of breed, sex, age, vaccination history and the nature of diarrhoea were assessed. Data obtained was analysed with SPSS and subjected to the chi-square test. Significance was at α0.05 Results: We found 61.11% tested positive (44/72) for CPV. Based on sex, 61.54% of males (20/33) and 60.61% of females tested positive (24/39). A total of 65.67% of samples from puppies below 6 months were positive. 56.25% of CPV vaccinated dogs and 70.83% of unvaccinated dogs were positive respectively. 69.05% of samples from haemorrhagic diarrhoeic dogs and 50.00% from non-haemorrhagic diarrhoeic dogs were positive of CPV. Conclusion: The study is the first documented evidence of the existence of CPV in Ghana. It also revealed that absence of bloody diarrhoea does not necessarily rule out CPV infection. PMID:29302647

DETECTION OF CANINE PARVOVIRUS ANTIGEN IN DOGS IN KUMASI, GHANA.

PubMed

Folitse, R D; Kodie, D O; Amemor, E; Dei, D; Tasiame, W; Burimuah, V; Emikpe, B O

2018-01-01

Canine Parvovirus (CPV) in dogs has been documented in many countries. However, evidence of the infection is scanty in Ghana. This study was conducted to detect canine parvovirus antigen in dogs presented with diarrhoea to the Government Veterinary Clinic in Kumasi, Ghana. Faecal samples from 72 dogs presented with diarrhoea were tested for the presence of canine parvovirus antigen using commercially available rapid test kit (BIT ® Rapid Colour Canine Parvovirus Ag Test Kit, BIOINDIST Co. Ltd, Korea) based on the principle of immunochromatography. Influence of breed, sex, age, vaccination history and the nature of diarrhoea were assessed. Data obtained was analysed with SPSS and subjected to the chi-square test. Significance was at α 0.05 . We found 61.11% tested positive (44/72) for CPV. Based on sex, 61.54% of males (20/33) and 60.61% of females tested positive (24/39). A total of 65.67% of samples from puppies below 6 months were positive. 56.25% of CPV vaccinated dogs and 70.83% of unvaccinated dogs were positive respectively. 69.05% of samples from haemorrhagic diarrhoeic dogs and 50.00% from non-haemorrhagic diarrhoeic dogs were positive of CPV. The study is the first documented evidence of the existence of CPV in Ghana. It also revealed that absence of bloody diarrhoea does not necessarily rule out CPV infection.

The role of rapid antigen testing for influenza in the era of molecular diagnostics.

PubMed

Dale, Suzanne E

2010-08-01

Rapid antigen testing for influenza has been both maligned and revered since its conception. Microbiologists have long lamented the lack of sensitivity of commercial rapid influenza detection tests (RIDTs), whereas many clinicians have eschewed their utility by emphasizing the value of definitely diagnosing influenza at the patient's bedside. RIDTs, although quick and easy to perform, are widely accepted as being less sensitive than traditional culture techniques and newer molecular methods, including reverse-transcription polymerase chain reaction (RT-PCR). Moreover, the performance characteristics of RIDTs vary widely, and their applications as clinical diagnostic tools are not well understood. In contrast, traditional techniques are time consuming and require significant expertise to perform. Often, the delay in diagnosing influenza through these methods has little impact on patient care. The benefits of achieving a diagnosis of influenza at the point of care are numerous and include increased access to appropriate antivirals, appropriate patient cohorting for infection control purposes, and better resource utilization. Therefore, it behooves the microbiology community to communicate these issues to clinicians and to work to improve the sensitivity of RIDTs.

[Evaluation of Prolex for the rapid identification of streptococci isolated in medical microbiology].

PubMed

Loubinoux, J; Mihaila-Amrouche, L; Bouvet, A

2004-10-01

The need to rapidly identify streptococci responsible for acute infectious diseases has led to the development of agglutination techniques that are able to identify streptococcal group antigens (A, B, C, D, F, and G) directly from primoculture colonies on blood agar. The Prolex agglutination tests (Pro-Lab Diagnostics, Richmond Hill, Ontario, Canada), distributed in France by i2a, have been used for the determination of group antigens of 166 isolates of streptococci and enterococci previously identified in the National Reference Center for Streptococci. The results obtained with the Prolex reagents have permitted to correctly identify all pyogenic beta-hemolytic streptococci (23 Streptococcus pyogenes, 21 Streptococcus agalactiae, 33 Streptococcus dysgalactiae subsp. equisimilis including 6 group C and 27 group G, and 5 Streptococcus porcinus including 4 group B). Four differences between unexpected agglutinations (A or F) and species identifications have been obtained. These differences were observed for four non-hemolytic isolates of Streptococcus mutans, Streptococcus gordonii, Streptococcus infantarius, and Streptococcus suis. The anti-D reagent has been of value as a marker for isolates of enterococci. Thus, these results confirm the abilities of these agglutination tests for the grouping of beta-hemolytic streptococci. Moreover, the use of Prolex has the advantage to be rapid because of the non-enzymatic but chemical extraction of streptococcal antigens.

Evaluation of two commercial procedures for rapid identification of Neisseria gonorrhoeae using a reference panel of antigenically diverse gonococci.

PubMed Central

Boehm, D M; Bernhardt, M; Kurzynski, T A; Pennell, D R; Schell, R F

1990-01-01

Two commercial tests for the rapid identification of Neisseria gonorrhoeae were evaluated. Two hundred seventy-nine organisms were tested, including 202 strains of N. gonorrhoeae. The Syva MicroTrak test results were less subjective but required a fluorescence microscope. The Phadebact Monoclonal GC OMNI Test required modification of the manufacturer's interpretive instructions in order to avoid cross-reactions, but it was a practical test. Specificities of both tests were 100%. Sensitivities of the Phadebact Monoclonal GC OMNI and Syva MicroTrak tests were 100% and approximately 100%, respectively. PMID:2121792

Tests for the measurement of factor VII-activating protease (FSAP) activity and antigen levels in citrated plasma, their correlation to PCR testing, and utility for the detection of the Marburg I-polymorphism of FSAP.

PubMed

Stephan, Sina; Schwarz, Herbert; Borchert, Anja; Bussfeld, Delia; Quak, Elfriede; Simshaeuser-Knaub, Beate; Teigelkamp, Stefan; Behrens, Fritz; Vitzthum, Frank

2008-01-01

The single nucleotide Marburg I (MRI) polymorphism of the factor VII-activating protease (FSAP) gene, the prourokinase-activating activity of FSAP, and antigen levels of FSAP in plasma have been associated with incidence and progression of carotid stenosis and venous thromboembolism. However, more information on the extent of these associations, potential further ones, and respective clinical utilities remain to be determined. At present, testing is performed mainly by PCR assays based on probes or SYBR Green I. Some studies include testing for antigen levels of total FSAP and its ability to activate prourokinase. To test large cohorts, it is beneficial to rely on assays that are cost-effective, reliable, easy to use, rapid to perform, and that may eventually be automated. In addition, it appears advantageous to use functional tests or tests that determine antigen levels as they may relate more closely to the phenotype than the genotype does. Tests for the measurements of antigen levels of FSAP and its prourokinase-activating activity were improved and performance characteristics assessed. To determine the FSAP genotypes, an amplification created restriction site (ACRS) PCR test was developed. Key performance characteristics of the FSAP activity and antigen tests were as follows: measuring range: 350-1400 mPEU/mL and 1.8-120 ng/mL, total coefficients of variation (CV): 5%-20% and 5%-14%, within-run CV: 4%-11% and 2.3%-12%, and run-to-run CV: 2%-17% and 4.3%-8.3%, respectively. The ratio of the activity and antigen level of FSAP correctly identified the FSAP genotypes of 126 samples tested. The ACRS PCR test is useful for laboratories that do not have the equipment to perform probe or SYBR Green I based real-time PCR. Furthermore, the tests developed for the determination of FSAP activity and antigen levels are convenient for determining clinical correlations, even for large population studies. The ratio of activity and antigen level of FSAP appears to be a promising and efficient alternative to molecular diagnostic techniques to detect the MRI polymorphism of FSAP.

[Pediatrics. Using the McIsaac score for the indication of rapid diagnostic testing in children with streptococcal pharyngitis].

PubMed

Pauchard, J-Y; Gehri, M; Vaudaux, B

2013-01-16

The McIsaac scoring system is a tool designed to predict the probability of streptococcal pharyngitis in children aged 3 to 17 years with a sore throat. Although it does not allow the physician to make the diagnosis of streptococcal pharyngitis, it enables to identify those children with a sore throat in whom rapid antigen detection tests have a good predictive value.

Evaluation of a monoclonal antibody-based latex agglutination test for diagnosis of cryptococcosis: comparison with two tests using polyclonal antibodies.

PubMed Central

Temstet, A; Roux, P; Poirot, J L; Ronin, O; Dromer, F

1992-01-01

Cryptococcal antigen detection has become a routine biological test performed for patients with AIDS. The poor prognosis of cryptococcosis explains the need for reliable tests. We evaluated the performances of a newly commercialized agglutination test that uses a monoclonal antibody specific for cryptococcal capsular polysaccharide (Pastorex Cryptococcus; Sanofi-Diagnostics Pasteur, Marnes-la-Coquette, France) and compared them with those of tests that use polyclonal immune sera (Cryptococcal Antigen Latex Agglutination System, Meridian Diagnostics, Inc., Cincinnati, Ohio; and Crypto-LA, International Biological Labs Inc., Cranbury, N.J.). The sensitivities and specificities of the tests were compared by using purified polysaccharides and yeast suspensions. Clinical specimens (131 serum samples, 41 cerebrospinal fluid samples, 34 urine samples, and 19 bronchoalveolar lavage samples) from 87 human immunodeficiency virus-positive subjects with (40 patients) and without (47 patients) culture-proven cryptococcosis were retrospectively tested during a blinded study. The effect of pronase treatment of samples was assessed for Pastorex Cryptococcus and the Cryptococcal Antigen Latex Agglutination System, and the antigen titers were compared. Our results show that (i) during the screening, concordance among the three tests was 97%; (ii) the use of pronase enhanced both the sensitivities and specificities of the Pastorex Cryptococcus test; (iii) titers agreed for 67% of the cerebrospinal fluid samples and 60% of the serum samples; and (iv) cryptococcosis was detected equally well with Pastorex Cryptococcus and with the other tests, whatever the infecting serotype (A, B, or D). The meaning of in vitro sensitivity and the relationship between titers and sensitivity are discussed. The results show that Pastorex Cryptococcus is a rapid and reliable test for the detection of cryptococcal antigen in body fluids and suggest that kits cannot be used interchangeably to monitor antigen titers in patients. PMID:1400951

Comparison of in vivo and in vitro survival and fecundity rates of the poultry red mite, Dermanyssus gallinae.

PubMed

Arkle, S; George, D R; Guy, J H; Sparagano, O A E

2010-04-01

To assist in the testing of possible antigens in developing a vaccine against the poultry red mite (Dermanyssus gallinae De Geer), a rapid and reliable in vitro screening method is critical. This short paper describes how D. gallinae survival and fecundity rates in an in vivo feeding device compared to that of mites fed using an in vitro method. Results showed that survival of fed D. gallinae females and mites overall was greater in vitro, although there was no difference between male survival and fecundity between in vivo and in vitro designs. The in vitro feeding device described therefore has the potential to provide reliable results, comparable to those obtained by in vivo testing, to allow for the rapid screening of D. gallinae antigens. Copyright 2009 Elsevier Ltd. All rights reserved.

A case of hyper-reactive malarial splenomegaly. The role of rapid antigen-detecting and PCR-based tests.

PubMed

Mothe, B; Lopez-Contreras, J; Torres, O H; Muñoz, C; Domingo, P; Gurgui, M

2008-03-01

Hyper-reactive malarial splenomegaly (HMS) - originally referred to as tropical splenomegaly syndrome - is characterized by a massive splenomegaly, high titres of anti-malarial antibodies and polyclonal IgM hypergammaglobulinemia. It is believed to be a consequence of an aberrant immunological response to prolonged exposure to malarial parasites. Although it is a frequent disease in the tropics, it is infrequent in western countries and is only seen in long-term residents from endemic areas. We describe the case of a 67-year-old Spanish man, a missionary in Cameroon for 30 years, who presented with a clinical history that fulfilled the diagnosis of HMS. We discuss the role and importance of PCR-based techniques in demonstrating lowgrade malarial parasitemia and the usefulness of new rapid antigen-detecting dipstick tests.

Sensitivity for Diagnosing Group A Streptococcal Pharyngitis from Manufacturers is 10% Higher than Reported in Peer-Reviewed Publications.

PubMed

Vachhani, Raj; Patel, Toral; Centor, Robert M; Estrada, Carlos A

2017-01-01

Meta-analyses based on peer-reviewed publications report a sensitivity of approximately 85% for rapid antigen streptococcus tests to diagnose group A streptococcal (GAS) pharyngitis. Because these meta-analyses excluded package inserts, we examined the test characteristics of rapid antigen streptococcal tests and molecular methods that manufacturers report in their package inserts. We included tests available in the US market (Food and Drug Administration, period searched 1993-2015) and used package insert data to calculate pooled sensitivity and specificity. To examine quality, we used the Quality Assessment of Diagnostic Accuracy Studies-2. We excluded 26 tests having different trade names but identical methods and data. The study design was prospective in 41.7% (10 of 24). The pooled sensitivity of the most commonly used method, lateral flow/immunochromatographic, was 95% (95% confidence interval [CI] 94-96) and the pooled specificity was 98% (96-98); 7108 patients. The pooled sensitivity of the polymerase chain reaction or molecular methods was 98% (95% CI 96-98) and the pooled specificity was 96% (95% CI 95-97); 5685 patients. Package inserts include sponsored studies that overestimate the sensitivity of rapid tests to diagnose GAS pharyngitis by approximately 10%. Physicians should understand that package inserts overestimate diagnostic test utility; a negative test cannot be used to exclude GAS pharyngitis.

Biconically tapered fiber optic probes for rapid label-free immunoassays.

PubMed

Miller, John; Castaneda, Angelica; Lee, Kun Ho; Sanchez, Martin; Ortiz, Adrian; Almaz, Ekrem; Almaz, Zuleyha Turkoglu; Murinda, Shelton; Lin, Wei-Jen; Salik, Ertan

2015-04-01

We report use of U-shaped biconically tapered optical fibers (BTOF) as probes for label-free immunoassays. The tapered regions of the sensors were functionalized by immobilization of immunoglobulin-G (Ig-G) and tested for detection of anti-IgG at concentrations of 50 ng/mL to 50 µg/mL. Antibody-antigen reaction creates a biological nanolayer modifying the waveguide structure leading to a change in the sensor signal, which allows real-time monitoring. The kinetics of the antibody (mouse Ig-G)-antigen (rabbit anti-mouse IgG) reactions was studied. Hydrofluoric acid treatment makes the sensitive region thinner to enhance sensitivity, which we confirmed by experiments and simulations. The limit of detection for the sensor was estimated to be less than 50 ng/mL. Utilization of the rate of the sensor peak shift within the first few minutes of the antibody-antigen reaction is proposed as a rapid protein detection method.

Development of a SARS Coronavirus Vaccine from Recombinant Spike Protein Plus Delta Inulin Adjuvant.

PubMed

McPherson, Clifton; Chubet, Richard; Holtz, Kathy; Honda-Okubo, Yoshikazu; Barnard, Dale; Cox, Manon; Petrovsky, Nikolai

2016-01-01

Given periodic outbreaks of fatal human infections caused by coronaviruses, development of an optimal coronavirus vaccine platform capable of rapid production is an ongoing priority. This chapter describes the use of an insect cell expression system for rapid production of a recombinant vaccine against severe acute respiratory syndrome coronavirus (SARS). Detailed methods are presented for expression, purification, and release testing of SARS recombinant spike protein antigen, followed by adjuvant formulation and animal testing. The methods herein described for rapid development of a highly protective SARS vaccine are equally suited to rapid development of vaccines against other fatal human coronavirus infections, e.g., the MERS coronavirus.

Dot-Blot Immunoassay of Fasciola gigantica Infection using 27 kDa and Adult Worm Regurge Antigens in Egyptian Patients

PubMed Central

Kamel, Hanan H.; Saad, Ghada A.

2013-01-01

The purpose of the present study was to evaluate the potential role of the 27-Kilodalton (KDa) antigen versus Fasciola gigantica adult worm regurge antigens in a DOT-Blot assay and to assess this assay as a practical tool for diagnosis fascioliasis in Egyptian patients. Fasciola gigantica antigen of an approximate molecular mass 27-(KDa) was obtained from adult worms by a simple elution SDS-PAGE. A Dot-Blot was developed comparatively to adult worm regurge antigens for the detection of specific antibodies from patients infected with F. gigantica in Egypt. Control sera were obtained from patients with other parasitic infections and healthy volunteers to assess the test and compare between the antigens. The sensitivity, specificity, positive and negative predictive values of Dot-Blot using the adult worm regurge were 80%, 90%, 94.1%, and 69.2% respectively, while those using 27-KDa were 100% which confirms the diagnostic potential of this antigen. All patients infected with Fasciola were positive, with cross reactivity reported with Schistosoma mansoni serum samples. This 27-KDa Dot-Blot assay showed to be a promising test which can be used for serodiagnosis of fascioliasis in Egyptian patients especially, those presenting with hepatic disease. It is specific, sensitive and easy to perform method for the rapid diagnosis particularly when more complex laboratory tests are unavailable. PMID:23710084

Dot-blot immunoassay of Fasciola gigantica infection using 27 kDa and adult worm regurge antigens in Egyptian patients.

PubMed

Kamel, Hanan H; Saad, Ghada A; Sarhan, Rania M

2013-04-01

The purpose of the present study was to evaluate the potential role of the 27-Kilodalton (KDa) antigen versus Fasciola gigantica adult worm regurge antigens in a DOT-Blot assay and to assess this assay as a practical tool for diagnosis fascioliasis in Egyptian patients. Fasciola gigantica antigen of an approximate molecular mass 27-(KDa) was obtained from adult worms by a simple elution SDS-PAGE. A Dot-Blot was developed comparatively to adult worm regurge antigens for the detection of specific antibodies from patients infected with F. gigantica in Egypt. Control sera were obtained from patients with other parasitic infections and healthy volunteers to assess the test and compare between the antigens. The sensitivity, specificity, positive and negative predictive values of Dot-Blot using the adult worm regurge were 80%, 90%, 94.1%, and 69.2% respectively, while those using 27-KDa were 100% which confirms the diagnostic potential of this antigen. All patients infected with Fasciola were positive, with cross reactivity reported with Schistosoma mansoni serum samples. This 27-KDa Dot-Blot assay showed to be a promising test which can be used for serodiagnosis of fascioliasis in Egyptian patients especially, those presenting with hepatic disease. It is specific, sensitive and easy to perform method for the rapid diagnosis particularly when more complex laboratory tests are unavailable.

Comparative performance of aldolase and lactate dehydrogenase rapid diagnostic tests in Plasmodium vivax detection

PubMed Central

2014-01-01

Background Misdiagnosis of malaria by commercial rapid diagnostic tests (RDTs) is a major cause of concern in the diagnosis of malaria. This retrospective study was aimed at assessing the relative performance of four RDTs with emphasis on the detection of two Plasmodium vivax antigens: aldolase and lactate dehydrogenase (LDH). Methods Three commercially available Plasmodium LDH or aldolase antigen detection kits (One Step Malaria P.f/P.v, ParaHit Total ver. 1.0, SD Bioline Malaria) and an anti-P. vivax aldolase-specific monoclonal antibody (mAb) pair 1C3-12 F10 were evaluated with P. vivax positive as well as non-P. vivax samples and healthy samples using blood smear examination as standard. Each test was read according to the manufacturer’s instructions. Results MAb 1C3-12 F10 pair targeting P. vivax-specific aldolase exhibited very good specificity and sensitivity of 100 and 97.4%, respectively. Positive predictive value (PPV) and negative predictive value (NPV) of 100 and 99.5%, respectively, were also observed. The anti-P. vivax LDH in the One-Step Malaria P.f/P.v test showed sensitivity, specificity, PPV and NPV of 93.5, 98.0, 88.9 and 98.8%, respectively. ParaHit Total ver. 1.0 targeting the pan-aldolase antigen showed sensitivity, specificity of 97.4 and 99.6%, respectively. PPV and NPV were both 99.5%. SD Bioline had sensitivity, specificity, PPV and NPV of 93.5, 100, 100 and 98.8%, respectively. The overall sensitivity and specificity of all four RDTs were acceptable, especially for the aldolase detection tests. Five (6.5%) of the P. vivax-positive samples (n = 77) that were confirmed by microscopic examination as well as the two aldolase detection RDTs (mAb 1C3-12 F10 and ParaHit Total ver.1.0) were undetected by the two LDH detection RDTs (One Step Malaria P.f/P.v and SD Bioline). Similarly, two positive samples (2.6%) that were positively confirmed by the LDH detection RDTs were also undetected by the aldolase detection test kits. Conclusion Aldolase and LDH antigens perform differently in different P. vivax samples; hence there is a high risk of misdiagnosis when monoclonal antibodies are used against only one particular antigen in the test. A combination of both aldolase and LDH in RDTs for the rapid diagnosis of P. vivax will enhance the sensitivity of the assay and reduce misdiagnosis. PMID:25015737

pLDH level of clinically isolated Plasmodium vivax and detection limit of pLDH based malaria rapid diagnostic test.

PubMed

Jang, Jin Woo; Cho, Chi Hyun; Han, Eun Taek; An, Seong Soo A; Lim, Chae Seung

2013-06-03

The malaria rapid diagnostic tests (RDTs) are now widely used in the world. Compared to Plasmodium falciparum, a poor sensitivity of RDTs was reported against Plasmodium vivax based on the adopted antibody against pan-Plasmodium antigen lactate dehydrogenase (pLDH) or aldolase. Levels of pLDH were measured from patient with P. vivax, and the correlations between the levels of pLDH and the sensitivities of RDTs were analysed among Republic of Korea (ROK) isolates. Three RDTs, OptiMAL test, SD BIOLINE Malaria Ag P.f/Pan test, Humasis Malaria Pf/Pan antigen test, and the Genedia pLDH antigen ELISA were performed with blood samples from 152 febrile patients and 100 healthy controls. Three malaria RDTs revealed sensitivities between 85.5 (131/152) and 86.8% (132/152) with highest sensitivity for the detection of P.vivax by pLDH antigen ELISA test (145/152, 95.4%) in comparison to traditional microscopy using Giemsa-stained slides. None of the healthy control tested positive by three RDTs or ELISA, indicating 100% specificity in their respective test. Levels of pLDH among Korean P. vivax isolates ranged between 0 ng/mL and 22,387.2 ng/mL (mean ± standard deviation 3,917.5 ± 6,120.9 ng/mL). The lower detection limits of three RDTs were between 25 and 50 ng/mL with artificially diluted samples. The moderate degree of correlation was observed between parasitaemia and concentrations of pLDH (r = 0.4, p < 0.05). The pLDH levels of P. vivax are the main explanation for the variations in the performance of pLDH-based RDTs. Therefore, comparing sensitivities of RDT may need to include targeted biomarker value of patients.

Evaluation of Nucleic Acid Preservation Cards for West Nile Virus Testing in Dead Birds

PubMed Central

Foss, Leslie; Reisen, William K.; Fang, Ying; Kramer, Vicki; Padgett, Kerry

2016-01-01

The California West Nile virus (WNV) Dead Bird Surveillance Program (DBSP) is an important component of WNV surveillance in the state. We evaluated FTA™ and RNASound™ cards as an alternative method for sampling dead birds for WNV molecular testing as these cards allow for more cost effective, rapid, and safer diagnostic sampling than the shipment of bird carcasses. To evaluate accuracy of results among avian sampling regimes, Reverse-Transcription Polymerase Chain Reaction (RT-PCR) results from FTA™ and RNASound™ cards were compared with results from kidney tissue, brain tissue, or oral swabs in lysis buffer in 2012–2013. In addition, RT-PCR results were compared with results from oral swabs tested by rapid antigen tests (RAMP™ and VecTOR™). While test results from the cards were not as sensitive as kidney tissue testing, they were more likely to provide accurate results than rapid antigen tests, and detected WNV in corvids as well as in other passerines, raptors, and waterfowl. Overall, WNV RT-PCR cycle threshold (Ct) scores from the cards were higher than those from tissue testing, but both card products displayed high sensitivity and specificity. American Crow samples provided the highest sensitivity. The cards also proved to be easier and more convenient vehicles for collecting and shipping samples, and in 2014 our program launched use of RNASound™ cards in the DBSP. Both FTA™ and RNASound™ products displayed 96% agreement with tissue results and are an adequate alternative sampling method for WNV dead bird testing. PMID:27341492

Validation of the Puumala virus rapid field test for bank voles in Germany.

PubMed

Reil, D; Imholt, C; Rosenfeld, U M; Drewes, S; Fischer, S; Heuser, E; Petraityte-Burneikiene, R; Ulrich, R G; Jacob, J

2017-02-01

Puumala virus (PUUV) causes many human infections in large parts of Europe and can lead to mild to moderate disease. The bank vole (Myodes glareolus) is the only reservoir of PUUV in Central Europe. A commercial PUUV rapid field test for rodents was validated for bank-vole blood samples collected in two PUUV-endemic regions in Germany (North Rhine-Westphalia and Baden-Württemberg). A comparison of the results of the rapid field test and standard ELISAs indicated a test efficacy of 93-95%, largely independent of the origin of the antigens used in the ELISA. In ELISAs, reactivity for the German PUUV strain was higher compared to the Swedish strain but not compared to the Finnish strain, which was used for the rapid field test. In conclusion, the use of the rapid field test can facilitate short-term estimation of PUUV seroprevalence in bank-vole populations in Germany and can aid in assessing human PUUV infection risk.

The dual function of the splenic marginal zone: essential for initiation of anti-TI-2 responses but also vital in the general first-line defense against blood-borne antigens

PubMed Central

ZANDVOORT, A; TIMENS, W

2002-01-01

The splenic marginal zone (S-MZ) is especially well equipped for rapid humoral responses and is unique in its ability to initiate an immune response to encapsulated bacteria (T-cell independent type 2 (TI-2) antigens). Because of the rapid spreading through the blood, infections with blood-borne bacteria form a major health risk. To cope with blood-borne antigens, a system is needed that can respond rapidly to a great diversity of organisms. Because of a number of unique features, S-MZ B cells can respond rapid and efficient to all sorts of blood-borne antigens. These unique features include a low blood flow microenvironment, low threshold for activation, high expression of complement receptor 2 (CR2, CD21) and multireactivity. Because of the unique high expression of CD21 in a low flow compartment, S-MZ B cells can bind and respond to TI-2 antigens even with relatively low-avid B cell receptors. Although TI-2 antigens are in general poorly opsonized by classic opsonins, a particular characteristic of these antigens is their ability to bind very rapidly to complement fragment C3d without the necessity of previous immunoglobulin binding. TI-2 primed S-MZ B cells, already by first passage through the germinal centre, will meet antigen-C3d complexes bound to follicular dendritic cells, allowing unique immediate isotype switching. This explains that the primary humoral response to TI-2 antigens is unique in its characterization by a rapid increase in IgM concurrent with IgG antibody levels. PMID:12296846

Evaluating the Use of Commercial West Nile Virus Antigens as Positive Controls in the Rapid Analyte Measurement Platform West Nile Virus Assay.

PubMed

Burkhalter, Kristen L; Savage, Harry M

2015-12-01

We evaluated the utility of 2 types of commercially available antigens as positive controls in the Rapid Analyte Measurement Platform (RAMP®) West Nile virus (WNV) assay. Purified recombinant WNV envelope antigens and whole killed virus antigens produced positive RAMP results and either type would be useful as a positive control. Killed virus antigens provide operational and economic advantages and we recommend their use over purified recombinant antigens. We also offer practical applications for RAMP positive controls and recommendations for preparing them.

Paper-based assay for red blood cell antigen typing by the indirect antiglobulin test.

PubMed

Yeow, Natasha; McLiesh, Heather; Guan, Liyun; Shen, Wei; Garnier, Gil

2016-07-01

A rapid and simple paper-based elution assay for red blood cell antigen typing by the indirect antiglobulin test (IAT) was established. This allows to type blood using IgG antibodies for the important blood groups in which IgM antibodies do not exist. Red blood cells incubated with IgG anti-D were washed with saline and spotted onto the paper assay pre-treated with anti-IgG. The blood spot was eluted with an elution buffer solution in a chromatography tank. Positive samples were identified by the agglutinated and fixed red blood cells on the original spotting area, while red blood cells from negative samples completely eluted away from the spot of origin. Optimum concentrations for both anti-IgG and anti-D were identified to eliminate the washing step after the incubation phase. Based on the no-washing procedure, the critical variables were investigated to establish the optimal conditions for the paper-based assay. Two hundred ten donor blood samples were tested in optimal conditions for the paper test with anti-D and anti-Kell. Positive and negative samples were clearly distinguished. This assay opens up new applications of the IAT on paper including antibody detection and blood donor-recipient crossmatching and extends its uses into non-blood typing applications with IgG antibody-based diagnostics. Graphical abstract A rapid and simple paper-based assay for red blood cell antigen typing by the indirect antiglobulin test.

Country-wide assessment of the genetic polymorphism in Plasmodium falciparum and Plasmodium vivax antigens detected with rapid diagnostic tests for malaria

PubMed Central

Mariette, Natacha; Barnadas, Céline; Bouchier, Christiane; Tichit, Magali; Ménard, Didier

2008-01-01

Background Rapid diagnostic tests (RDTs) are becoming increasingly indispensable in malaria management, as a means of increasing the accuracy of diagnosis. The WHO has issued recommendations, but the selection of the most suitable RDT remains difficult for users in endemic countries. The genetic variability of the antigens detected with RDTs has been little studied, but may affect the sensitivity of RDTs. This factor has been studied by comparisons between countries at continental level, but little information is available concerning antigen variability within a given country. Methods A country-wide assessment of polymorphism of the PfHRP2, PfHRP3, pLDH and aldolase antigens was carried out in 260 Plasmodium falciparum and 127 Plasmodium vivax isolates, by sequencing the genes encoding these antigens in parasites originating from the various epidemiological strata for malaria in Madagascar. Results Higher levels of polymorphism were observed for the pfhrp2 and pfhrp3 genes than for the P. falciparum and P. vivax aldolase and pldh genes. Pfhrp2 sequence analysis predicted that 9% of Malagasy isolates would not be detected at parasite densities ≤ 250 parasites/μl (ranging from 6% in the north to 14% in the south), although RDTs based on PfHRP2 detection are now recommended in Madagascar. Conclusion These findings highlight the importance of training of health workers and the end users of RDTs in the provision of information about the possibility of false-negative results for patients with clinical symptoms of malaria, particularly in the south of Madagascar. PMID:18957099

Prevalence and levels of Renibacterium salmoninarum in spring-summer Chinook salmon (Oncorhynchus tshawytscha) smolts at dams on the Columbia and Snake Rivers.

USGS Publications Warehouse

Elliott, D.G.; Pascho, R.J.; Jackson, L.M.; Mathews, G.M.; Harmon, J.R.

1997-01-01

We evaluated Renibaeterium salmoninarum infection in smolts of hatchery and wild spring-summer Chinook salmon Oncorhynchus tshawytscha sampled during most of the outmigration at Little Goose (1988) and Lower Granite dams (1988–1991) on the Snake River and at Priest Rapids and McNary dams on the Columbia River (1988–1990). We sampled 860–2,178 fish per dam each year. Homogenates of kidney–spleen tissue from all fish were tested for the presence of R. salmoninarum antigens by the enzyme-linked immunosorbent assay (ELISA), and homogenates from 10% of the fish were examined by the fluorescent antibody technique (FAT). Although only 1–11% of fish sampled at a given dam during any l year exhibited lesions characteristic of bacterial kidney disease, 86–100% of the fish tested positive for R. salmoninarum antigen by ELISA, whereas 4–17% of the fish tested positive by the FAT. During most years, a majority (68–87%) of fish testing positive by the ELISA had low R. salmoninarum antigen levels, but in 1989, 53% of positive fish from Lower Granite Dam and 52% from McNary Dam showed medium-to-high antigen levels. For most years, the highest mean antigen levels were measured in fish sampled after 75% of the total out-migrants had passed a given dam. When the largest numbers of fish were being collected for bypass or downriver transportation, mean antigen levels were relatively low.

Renibacterium salmoninarum in spring-summer chinook salmon smolts at dams on the Columbia and Snake Rivers

USGS Publications Warehouse

Elliott, D.G.; Pascho, R.J.; Jackson, L.M.; Matthews, G.M.; Harmon, J.R.

1997-01-01

We evaluated Renibacterium salmoninarum infection in smolts of hatchery and wild spring-summer chinook salmon Oncorhynchus tshawytscha sampled during most of the out-migration at Little Goose (1988) and Lower Granite dams (1988-1991) on the Snake River and at Priest Rapids and McNary dams on the Columbia River (1988-1990). We sampled 860-2,178 fish per dam each year. Homogenates of kidney-spleen tissue from all fish were tested for the presence of R. salmoninarum antigens by the enzyme-linked immunosorbent assay (ELISA), and homogenates from 10% of the fish were examined by the fluorescent antibody technique (FAT). Although only 1-11% of fish sampled at a given dam during any 1 year exhibited lesions characteristic of bacterial kidney disease, 86-100% of the fish tested positive for R. salmoninarum antigen by ELISA, whereas 4-17% of the fish tested positive by the FAT. During most years, a majority (68-87%) of fish testing positive by the ELISA had low R. salmoninarum antigen levels, but in 1989, 53% of positive fish from Lower Granite Dam and 52% from McNary Dam showed medium-to-high antigen levels. For most years, the highest mean antigen levels were measured in fish sampled after 75% of the total out-migrants had passed a given dam. When the largest numbers of fish were being collected for bypass or downriver transportation, mean antigen levels were relatively low.

Acute Legionella pneumophila infection masquerading as acute alcoholic hepatitis.

PubMed

Hunter, Jonathan Michael; Chan, Julian; Reid, Angeline Louise; Tan, Chistopher

2013-01-25

A middle-aged man had deteriorated rapidly in hospital after being misdiagnosed with acute alcoholic hepatitis. Acute Legionnaires disease (Legionellosis) was subsequently diagnosed on rapid antigen urinary testing and further confirmed serologically. This led to appropriate antibiotic treatment and complete clinical resolution. Physicians caring for patients with alcohol-related liver disease should consider Legionella pneumophila in their differential diagnosis even with a paucity of respiratory symptoms.

Comparison of BioStar Strep A OIA optical immune assay, Abbott TestPack Plus Strep A, and culture with selective media for diagnosis of group A streptococcal pharyngitis.

PubMed Central

Roe, M; Kishiyama, C; Davidson, K; Schaefer, L; Todd, J

1995-01-01

We directly compared three techniques for the diagnosis of group A streptococcal pharyngitis in 500 symptomatic children seen in the Emergency Department or Child Care Clinic of The Children's Hospital of Denver. Throats were vigorously swabbed with two rayon swabs, which were transported immediately to the Microbiology Laboratory. Each swab was cultured aerobically on Strep A Isolation Agar (Remel) and then tested for antigen-one swab by the Strep A OIA optical immune assay (BioStar) and the other by the TestPack Plus Strep A (Abbott) technique. Each test was performed blind to the others. The refrigerated pledget was cultured in Todd-Hewitt broth if an antigen test was positive and both direct plate cultures were negative (the "gold standard" was any culture positive). All isolates were serologically grouped. Of 500 complete patient cultures, 151 (30%) were positive for group A streptococcal growth. The two antigen tests gave comparable results with an average sensitivity of 83%. Each was significantly (P < 0.02) less sensitive than its corresponding culture. The BioStar Strep A OIA optical immune assay produced significantly (P < 0.003) more false-positive results than did the Abbott test. Rapid antigen testing is not sensitive enough to eliminate the need for backup cultures. PMID:7650184

Detection of K1 antigen of Escherichia coli rods isolated from pregnant women and neonates.

PubMed

Kaczmarek, Agnieszka; Budzyńska, Anna; Gospodarek, Eugenia

2014-09-01

The K1 antigen is an important virulence determinant of Escherichia coli strains and has been shown to be associated particularly with neonatal meningitis, bacteraemia and septicaemia. Thus, its detection seems to be useful, especially in the case of E. coli strains isolated from pregnant women and newborns. In this study, the sensitivity and specificity of the latex agglutination test (Pastorex Meningitis) for identification of E. coli serogroup K1 were assessed, using PCR as the gold standard. Our results showed that consistency of results between latex agglutination test and PCR amounted to 98.5%. Therefore, Pastorex Meningitis is a good alternative to PCR and could be used for rapid K1 antigen detection, especially in local non-specialized laboratories with limited resources where PCR assay is not applied.

A Rapid and Sensitive Method for the Quantitation of Legionella Pneumophila Antigen from Human Urine.

DTIC Science & Technology

1980-11-12

320-325. 4. Hedlund, K. W., V. G. McGann, D. S . Copeland, S . F. Little, and R. G. Allen. 1979. Immunologic protection against the Legionnaires ’ disease ...reverse% passive hemagglutination test was developed to assay concentrations of solubl4 antigen of Legionnaires ’ Disease (Legionella pneumophila) in...URINE U. S . Arm" Medical Research Institute of Infectious Diseases Fort Detrick, Frederick, Maryland 21701 The views of the authors do not purport to

STUDIES ON SCRUB TYPHUS

PubMed Central

Smadel, Joseph E.; Rights, Fred L.; Jackson, Elizabeth B.

1946-01-01

A complement-fixing antigen specific for scrub typhus occurs in the body fluids and tissues of infected mice, white rats, and cotton rats. The specific serological substance is demonstrable only in those animals which develop a rapidly fatal disease after an incubation period of a few days. Such an experimental infection is induced in mice and rats by the intravenous injection of suspensions of yolk sac rich in R. orientalis. Ether extraction is an important step in the preparation of a complement-fixing antigen from tissues of mice dying with scrub typhus. The Imphal No. 8 and Calcutta strains of R. orientalis are indistinguishable on the basis of complement fixation and cross-immunity tests. The complement-fixing antigen in body fluids of infected mice and rats and in our preparations of tissues from such animals occurs as a soluble antigen. Under the proper conditions the soluble antigen can be stored or dehydrated without loss of serological activity. PMID:19871518

Rapid Assay of Cellular Immunity in Q Fever.

DTIC Science & Technology

1995-10-01

Integrated Diagnostics for activity by re-incubation with L929 cells and no infectious material was detected. This antigen was tested for the ability to...UNCLASSIFIED •%E L E• M1 lt*’E••l DEC 1 119954 F A CONTRACT NUMBER: DAND17-95-C-5057 TITLE: Rapid Assay of Cellular Immunity in Q Fever PRINCIPAL INVESTIGATOR...SUBTITLE 5. FUNDING NUMBERS Rapid Assay of Cellular Immunity in Q Fever DAMDI7-95-C-5057 6. AUTHOR(S) Marjorie Wier, Ph.D. 7. PERFORMING ORGANIZATION

A new rapid immunohistochemical staining technique using the EnVision antibody complex.

PubMed

Kämmerer, U; Kapp, M; Gassel, A M; Richter, T; Tank, C; Dietl, J; Ruck, P

2001-05-01

Rapid immunohistochemical investigation, in addition to staining with hematoxylin and eosin, would be useful during intraoperative frozen section diagnosis in some cases. This study was undertaken to investigate whether the recently described EnVision system, a highly sensitive two-step immunohistochemical technique, could be modified for rapid immunostaining of frozen sections. Forty-five primary antibodies were tested on frozen sections from various different tissues. After fixation in acetone for 1 min and air-drying, the sections were incubated for 3 min each with the primary antibody, the EnVision complex (a large number of secondary antibodies and horseradish peroxidase coupled to a dextran backbone), and the chromogen (3,3'diaminobenzidine or 3-amino-9-ethylcarbazole). All reactions were carried out at 37C. Specific staining was seen with 38 antibodies (including HMB-45 and antibodies against keratin, vimentin, leukocyte common antigen, smooth muscle actin, synaptophysin, CD34, CD3, CD20, and prostate-specific antigen). A modification of the EnVision method allows the detection of a broad spectrum of antigens in frozen sections in less than 13 min. This method could be a useful new tool in frozen section diagnosis and research. (J Histochem Cytochem 49:623-630, 2001)

Detection of rabies antigen in the saliva and brains of apparently healthy dogs slaughtered for human consumption and its public health implications in abia state, Nigeria.

PubMed

Mshelbwala, P P; Ogunkoya, A B; Maikai, B V

2013-01-01

The study was carried out in eight dogs slaughtering outlets within four Local Government Areas of the State for the determination of rabies antigen in the saliva and brain of apparently healthy dogs slaughtered for human consumption. A total of one hundred (100) samples each of saliva and brain were collected before and after slaughter, respectively, between April to June, 2013, in the selected areas. The saliva was subjected to rapid immune-chromatographic test (RICT) while direct fluorescent antibody test (DFAT) was carried out on the brain samples. Structured questionnaire was administered to nineteen (19) dog meat processors comprising 18 males and 1 female in the selected areas. Sixty four percent of the samples tested were from female dogs while 36% were from males, 5% tested positive for rabies antigen with the use of both tests; there was no statistical association between sex and rabies status of the dogs sampled (P > 0.05). Butchers bitten during the course of slaughtering were 94.7% out of which 72.8% utilized traditional method of treatment and only 27.8% reported to the hospital for proper medical attention. This study has established the presence of rabies antigen in apparently healthy dogs in the study area.

[New Radiopharmaceuticals Based on Prostate-Specific Inhibitors of Membrane Antigen for Diagnostics and Therapy of Metastatic Prostate Cancer].

PubMed

Vlasova, O P; German, K E; Krilov, V V; Petriev, V M; Epstein, N B

2015-01-01

About 10.7% cases of prostate cancer were registered in Russia in 2011 (40,000 patients). More than half of cancer cases were revealed in advanced (III-IV) stages when metastases inevitably developed quickly. Clinical problem of early diagnostics and treatment of metastatic prostate cancer is still not solved. Anatomical imaging techniques have low sensitivity and specificity for the detection of this disease. Metabolic visualization methods which use prostate specific antigen (PSA) as a marker are also ineffective. This article describes prostate-specific membrane antigens (PSMA) that are proposed as a marker for diagnostics and therapy of prostate cancer. The most promising PSMA-based radiopharmaceutical agent for diagnostics has been developed and clinically tested in the European countries. These pharmaceuticals are based on small peptide molecules modified with urea, and have the highest affinity to PSMA. Favorable phannacokinetics, rapid accumulation in the tumor and rapid excretion from the body are beneficial features of these pharmaceuticals.

Comparative Study of rK39 Leishmania Antigen for Serodiagnosis of Visceral Leishmaniasis: Systematic Review with Meta-Analysis

PubMed Central

Maia, Zuinara; Lírio, Monique; Mistro, Sóstenes; Mendes, Carlos Maurício Cardeal; Mehta, Sanjay R.; Badaro, Roberto

2012-01-01

Background The rK39 recombinant protein is derived from a specific antigen produced by the Leishmania donovani complex, and has been used in the last two decades for the serodiagnosis of visceral leishmaniasis. We present here a systematic review and meta-analysis of studies evaluating serologic assays to diagnose visceral leishmaniasis to determine the accuracy of rK39 antigen in comparison to the use of other antigen preparations. Methodology/Principal Findings A systematic review with meta-analysis of the literature was performed to compare the rK39 strip-test and ELISA formats against serological tests using promastigote antigens derived from whole or soluble parasites for Direct Aglutination Test (DAT), Indirect Immunofluorescence test (IFAT) and ELISA with a promastigote antigen preparation (p-ELISA). Gold standard diagnosis was defined by the demonstration of amastigotes on hematological specimens. A database search was performed on Medline, Lilacs, Scopus, Isi Web of Science, and Cochrane Library. Quality of data was assessed using the QUADAS questionnaire. A search of the electronic databases found 352 papers of which only 14 fulfilled the selection criteria. Three evaluated the rK39 ELISA, while 13 evaluated the rK39 immunochromatographic strip test. The summarized sensitivity for the rK39-ELISA was 92% followed by IFAT 88% and p-ELISA 87%. The summarized specificity for the three diagnostic tests was 81%, 90%, and 77%. Studies comparing the rK39 strip test with DAT found a similar sensitivity of 94%, although the DAT had a slightly higher specificity. The rK39 strip test was more sensitive and specific than the IFAT and p-ELISA. We did not detect any difference in the sensitivity and specificity between strips produced by different manufacturers. Conclusions The rK39 protein used either in a strip test or in an ELISA, and the DAT are the best choices for implementation of rapid, easy and efficient test for serodiagnosis of VL. PMID:22303488

Comparative study of rK39 Leishmania antigen for serodiagnosis of visceral leishmaniasis: systematic review with meta-analysis.

PubMed

Maia, Zuinara; Lírio, Monique; Mistro, Sóstenes; Mendes, Carlos Maurício Cardeal; Mehta, Sanjay R; Badaro, Roberto

2012-01-01

The rK39 recombinant protein is derived from a specific antigen produced by the Leishmania donovani complex, and has been used in the last two decades for the serodiagnosis of visceral leishmaniasis. We present here a systematic review and meta-analysis of studies evaluating serologic assays to diagnose visceral leishmaniasis to determine the accuracy of rK39 antigen in comparison to the use of other antigen preparations. A systematic review with meta-analysis of the literature was performed to compare the rK39 strip-test and ELISA formats against serological tests using promastigote antigens derived from whole or soluble parasites for Direct Aglutination Test (DAT), Indirect Immunofluorescence test (IFAT) and ELISA with a promastigote antigen preparation (p-ELISA). Gold standard diagnosis was defined by the demonstration of amastigotes on hematological specimens. A database search was performed on Medline, Lilacs, Scopus, Isi Web of Science, and Cochrane Library. Quality of data was assessed using the QUADAS questionnaire. A search of the electronic databases found 352 papers of which only 14 fulfilled the selection criteria. Three evaluated the rK39 ELISA, while 13 evaluated the rK39 immunochromatographic strip test. The summarized sensitivity for the rK39-ELISA was 92% followed by IFAT 88% and p-ELISA 87%. The summarized specificity for the three diagnostic tests was 81%, 90%, and 77%. Studies comparing the rK39 strip test with DAT found a similar sensitivity of 94%, although the DAT had a slightly higher specificity. The rK39 strip test was more sensitive and specific than the IFAT and p-ELISA. We did not detect any difference in the sensitivity and specificity between strips produced by different manufacturers. The rK39 protein used either in a strip test or in an ELISA, and the DAT are the best choices for implementation of rapid, easy and efficient test for serodiagnosis of VL.

Impact of the Mycobaterium africanum West Africa 2 Lineage on TB Diagnostics in West Africa: Decreased Sensitivity of Rapid Identification Tests in The Gambia.

PubMed

Ofori-Anyinam, Boatema; Kanuteh, Fatoumatta; Agbla, Schadrac C; Adetifa, Ifedayo; Okoi, Catherine; Dolganov, Gregory; Schoolnik, Gary; Secka, Ousman; Antonio, Martin; de Jong, Bouke C; Gehre, Florian

2016-07-01

MPT64 rapid speciation tests are increasingly being used in diagnosis of tuberculosis (TB). Mycobacterium africanum West Africa 2 (Maf 2) remains an important cause of TB in West Africa and causes one third of disease in The Gambia. Since the introduction of MPT64 antigen tests, a higher than expected rate of suspected non-tuberculous mycobacteria (NTM) was seen among AFB smear positive TB suspects, which led us to prospectively assess sensitivity of the MPT64 antigen test in our setting. We compared the abundance of mRNA encoded by the mpt64 gene in sputa of patients with untreated pulmonary TB caused by Maf 2 and Mycobacterium tuberculosis (Mtb). Subsequently, prospectively collected sputum samples from presumptive TB patients were inoculated in the BACTEC MGIT 960 System. One hundred and seventy-three acid fast bacilli (AFB)-positive and blood agar negative MGIT cultures were included in the study. Cultures were tested on the day of MGIT positivity with the BD MGIT TBc Identification Test. A random set of positives and all negatives were additionally tested with the SD Bioline Ag MPT64 Rapid. MPT64 negative cultures were further incubated at 37°C and retested until positive. Bacteria were spoligotyped and assigned to different lineages. Maf 2 isolates were 2.52-fold less likely to produce a positive test result and sensitivity ranged from 78.4% to 84.3% at the beginning and end of the recommended 10 day testing window, respectively. There was no significant difference between the tests. We further showed that the decreased rapid test sensitivity was attributable to variations in mycobacterial growth behavior and the smear grades of the patient. In areas where Maf 2 is endemic MPT64 tests should be cautiously used and MPT64 negative results confirmed by a second technique, such as nucleic acid amplification tests, to avoid their misclassification as NTMs.

Acute Legionella pneumophila infection masquerading as acute alcoholic hepatitis

PubMed Central

Hunter, Jonathan Michael; Chan, Julian; Reid, Angeline Louise; Tan, Chistopher

2013-01-01

A middle-aged man had deteriorated rapidly in hospital after being misdiagnosed with acute alcoholic hepatitis. Acute Legionnaires disease (Legionellosis) was subsequently diagnosed on rapid antigen urinary testing and further confirmed serologically. This led to appropriate antibiotic treatment and complete clinical resolution. Physicians caring for patients with alcohol-related liver disease should consider Legionella pneumophila in their differential diagnosis even with a paucity of respiratory symptoms. PMID:23355576

Quantifying serum antibody in bird fanciers' hypersensitivity pneumonitis.

PubMed

McSharry, Charles; Dye, George M; Ismail, Tengku; Anderson, Kenneth; Spiers, Elizabeth M; Boyd, Gavin

2006-06-26

Detecting serum antibody against inhaled antigens is an important diagnostic adjunct for hypersensitivity pneumonitis (HP). We sought to validate a quantitative fluorimetric assay testing serum from bird fanciers. Antibody activity was assessed in bird fanciers and control subjects using various avian antigens and serological methods, and the titer was compared with symptoms of HP. IgG antibody against pigeon serum antigens, quantified by fluorimetry, provided a good discriminator of disease. Levels below 10 mg/L were insignificant, and increasing titers were associated with disease. The assay was unaffected by total IgG, autoantibodies and antibody to dietary hen's egg antigens. Antigens from pigeon serum seem sufficient to recognize immune sensitivity to most common pet avian species. Decreasing antibody titers confirmed antigen avoidance. Increasing antibody titer reflected the likelihood of HP, and decreasing titers confirmed antigen avoidance. Quantifying antibody was rapid and the increased sensitivity will improve the rate of false-negative reporting and obviate the need for invasive diagnostic procedures. Automated fluorimetry provides a method for the international standardization of HP serology thereby improving quality control and improving its suitability as a diagnostic adjunct.

Rapid diagnostic tests duo as alternative to conventional serological assays for conclusive Chagas disease diagnosis.

PubMed

Egüez, Karina E; Alonso-Padilla, Julio; Terán, Carolina; Chipana, Zenobia; García, Wilson; Torrico, Faustino; Gascon, Joaquim; Lozano-Beltran, Daniel-Franz; Pinazo, María-Jesús

2017-04-01

Chagas disease is caused by the parasite Trypanosoma cruzi. It affects several million people, mainly in Latin America, and severe cardiac and/or digestive complications occur in ~30% of the chronically infected patients. Disease acute stage is mostly asymptomatic and infection goes undiagnosed. In the chronic phase direct parasite detection is hampered due to its concealed presence and diagnosis is achieved by serological methods, like ELISA or indirect hemagglutination assays. Agreement in at least two tests must be obtained due to parasite wide antigenic variability. These techniques require equipped labs and trained personnel and are not available in distant regions. As a result, many infected people often remain undiagnosed until it is too late, as the two available chemotherapies show diminished efficacy in the advanced chronic stage. Easy-to-use rapid diagnostic tests have been developed to be implemented in remote areas as an alternative to conventional tests. They do not need electricity, nor cold chain, they can return results within an hour and some even work with whole blood as sample, like Chagas Stat-Pak (ChemBio Inc.) and Chagas Detect Plus (InBIOS Inc.). Nonetheless, in order to qualify a rapidly diagnosed positive patient for treatment, conventional serological confirmation is obligatory, which might risk its start. In this study two rapid tests based on distinct antigen sets were used in parallel as a way to obtain a fast and conclusive Chagas disease diagnosis using whole blood samples. Chagas Stat-Pak and Chagas Detect Plus were validated by comparison with three conventional tests yielding 100% sensitivity and 99.3% specificity over 342 patients seeking Chagas disease diagnosis in a reference centre in Sucre (Bolivia). Combined used of RDTs in distant regions could substitute laborious conventional serology, allowing immediate treatment and favouring better adhesion to it.

Evaluation of surface antigen TF1.17 in feline Tritrichomonas foetus isolates.

PubMed

Gould, E N; Corbeil, L B; Kania, S A; Tolbert, M K

2017-09-15

Tritrichomonas foetus (T. foetus) is a flagellated protozoa that infects the distal ileum and proximal colon of domestic cats, as well as the urogenital tract of cattle. Feline trichomonosis is recognized as a prevalent cause of chronic diarrhea in cats worldwide. The suspected route of transmission is fecal-oral, with cats in densely crowded environments at highest risk for infection. Thus, the recommended strategy for minimizing spread of infection is to identify and isolate T. foetus-positive cats from the general population. Rapid identification of infected cats can be challenging due to the inability to accurately and quickly detect the organism in samples at point of care facilities. Thus, identification of targets for use in development of a novel diagnostic test, as well as a vaccine or therapy for T. foetus infection is a significant area of research. Despite a difference in organ tropism between T. foetus genotypes, evidence exists for conserved virulence factors between feline and bovine T. foetus. The bovine T. foetus surface antigen, TF1.17, is an adhesin that is conserved across isolates. Vaccination with the purified antigen results in amelioration of cytopathogenicity and more rapid clearance of infection in cattle. We previously showed that three feline isolates of T. foetus were positive for TF1.17 antigen so we further hypothesized that TF1.17 is conserved across feline T. foetus isolates and that this antigen would represent an attractive target for development of a novel diagnostic test or therapy for feline trichomonosis. In these studies, we used monoclonal antibodies previously generated against 1.15 and 1.17 epitopes of the bovine T. foetus TF1.17 antigen, to evaluate for the presence and role of TF1.17 in the cytopathogenicity of feline T. foetus. A previously validated in vitro co-culture approach was used to model feline T. foetus infection. Immunoblotting, immunofluorescence assays, and flow cytometric analysis confirmed the presence and surface localization of antigen TF1.17 across all feline T. foetus isolates tested. Antigen TF1.17 was notably absent in the presumably nonpathogenic intestinal trichomonad, Pentatrichomonas hominis, a parasite that can be confused microscopically with T. foetus. Similar to bovine trichomoniasis, TF1.17 was found to promote T. foetus adhesion to the intestinal epithelium. These results support further characterization and development of the TF1.17 antigen as a possible target for the diagnosis and prevention of feline T. foetus infection. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of twenty-two rapid antigen detection tests in the diagnosis of Equine Influenza caused by viruses of H3N8 subtype.

PubMed

Yamanaka, Takashi; Nemoto, Manabu; Bannai, Hiroshi; Tsujimura, Koji; Kondo, Takashi; Matsumura, Tomio; Gildea, Sarah; Cullinane, Ann

2016-03-01

Equine influenza (EI) is a highly contagious disease caused by viruses of the H3N8 subtype. The rapid diagnosis of EI is essential to reduce the disease spread. Many rapid antigen detection (RAD) tests for diagnosing human influenza are available, but their ability to diagnose EI has not been systematically evaluated. The aim of this study was to compare the performance of 22 RAD tests in the diagnosis of EI. The 22 RAD tests were performed on fivefold serial dilutions of EI virus to determine their detection limits. The four most sensitive RAD tests (ImmunoAce Flu, BD Flu examan, Quick chaser Flu A, B and ESPLINE Influenza A&B-N) were further evaluated using nasopharyngeal samples collected from experimentally infected and naturally infected horses. The results were compared to those obtained using molecular tests. The detection limits of the 22 RAD tests varied hugely. Even the four RAD tests showing the best sensitivity were 125-fold less sensitive than the molecular techniques. The duration of virus detection in the experimentally infected horses was shorter using the RAD tests than using the molecular techniques. The RAD tests detected between 27% and 73% of real-time RT-PCR-positive samples from naturally infected horses. The study demonstrated the importance of choosing the right RAD tests as only three of 22 were fit for diagnosing EI. It was also indicated that even RAD tests with the highest sensitivity serve only as an adjunct to molecular tests because of the potential for false-negative results. © 2015 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

Six-week follow-up after HIV-1 exposure: a position statement from the Public Health Agency of Sweden and the Swedish Reference Group for Antiviral Therapy.

PubMed

Gaines, Hans; Albert, Jan; Axelsson, Maria; Berglund, Torsten; Gisslén, Magnus; Sönnerborg, Anders; Blaxhult, Anders; Bogdanovic, Gordana; Brytting, Maria; Carlander, Christina; Flamholc, Leo; Follin, Per; Haggar, Axana; Hagstam, Per; Johansson, Marcus; Navér, Lars; Persson Blom, Jenny; Samuelson, Agneta; Ström, Helena; Sundqvist, Martin; Svedhem Johansson, Veronica; Tegmark Wisell, Karin; Tegnell, Anders; Thorstensson, Rigmor

2016-02-01

In 2014 the Public Health Agency of Sweden and the Swedish Reference Group for Antiviral Therapy (RAV) conducted a review and analysis of the state of knowledge on the duration of follow-up after exposure to human immunodeficiency virus (HIV). Up until then a follow-up of 12 weeks after exposure had been recommended, but improved tests and new information on early diagnosis motivated a re-evaluation of the national recommendations by experts representing infectious diseases and microbiology, county medical officers, the RAV, the Public Health Agency, and other national authorities. Based on the current state of knowledge the Public Health Agency of Sweden and the RAV recommend, starting in April 2015, a follow-up period of 6 weeks after possible HIV-1 exposure, if HIV testing is performed using laboratory-based combination tests detecting both HIV antibody and antigen. If point-of-care rapid HIV tests are used, a follow-up period of 8 weeks is recommended, because currently available rapid tests have insufficient sensitivity for detection of HIV-1 antigen. A follow-up period of 12 weeks is recommended after a possible exposure for HIV-2, since presently used assays do not include HIV-2 antigens and only limited information is available on the development of HIV antibodies during early HIV-2 infection. If pre- or post-exposure prophylaxis is administered, the follow-up period is recommended to begin after completion of prophylaxis. Even if infection cannot be reliably excluded before the end of the recommended follow-up period, HIV testing should be performed at first contact for persons who seek such testing.

[Laboratory diagnosis of bacterial meningitis: usefulness of various tests for the determination of the etiological agent].

PubMed

Carbonnelle, E

2009-01-01

Despite breakthroughs in the diagnosis and treatment of infectious diseases, meningitis still remains an important cause of mortality and morbidity. An accurate and rapid diagnosis of acute bacterial meningitis is essential for a good outcome. The gold-standard test for diagnosis is CSF analysis. Gram staining of CSF reveals bacteria in about 50 to 80 % of cases and cultures are positive in at best 80 % of cases. However, the sensitivity of both tests is less than 50 % in patients who are already on antibiotic treatment. CSF leukocyte count and concentration of protein and glucose lack specificity and sensitivity for the diagnosis of meningitis. Other biological tests are available for the diagnosis. Latex agglutination test were adapted for rapid and direct detection of soluble bacterial antigens in CSF of patients suspected with bacterial meningitis. This test is efficient in detecting antigens of most common central nervous system bateria but lacks sensibility. Furthermore, in the early phases of acute bacterial and viral meningitis, signs and symptoms are often non specific and it is not always possible to make a differential diagnosis. Markers like CRP, procalcitonin, or sTREM-1 may be very useful for the diagnosis and to differentiate between viral and bacterial meningitis. Bacterial meningitis diagnosis and management require various biological tests and a multidisciplinary approach.

Cloning and Expression of Genes for Dengue Virus Type-2 Encoded-Antigens for Rapid Diagnosis and Vaccine Development

DTIC Science & Technology

1988-10-31

00 0 Cloning and Expression of Genes for Dengue Virus (Type-2 Encoded-Antigens for Rapid ODiagnosis and Vaccine DevelopmentN| ANNUAL PROGRESS REPORT...11. TITLE (include Security Classification) Cloning and Expression of Genes f or Dengue Virus Type 2 Fncoded Antigens for Rapid Diagnosis and Vaccine ...epidemics in Central and South Americas and the Caribbean is a cause of major concern. An effective vaccine is not available to protect individuals

Cell mediated immunity cross-reactions of mycobacteria: polymorphism of target bacterial antigens.

PubMed Central

Kulkarni, S; Hattikudur, S; Kamat, R S

1986-01-01

Swiss white mice were immunized with different mycobacteria and delayed type hypersensitivity (DTH) responses were studied by the foot-pad swelling technique of Gray and Jennings (1955). Extensive cross-reactions in DTH, outside the limits of Runyon's groups were observed. As a general trend slow growing mycobacteria showed greater cross-reactivity with slow growers than with rapid growers and vice versa. The implied cross-protective significance of DTH cross-reactions was further confirmed by demonstration of the ability of DTH cross-reacting sonicates to generate activated macrophages in M. avium immunized mice. An antiserum was raised against the earlier reported DTH eliciting antigen of M. tuberculosis H37Rv (DTH-H37Rv). The sero-reactivity of anti-DTH-H37Rv against the sonicates of different mycobacteria was studied with the objective of investigating the molecular basis of DTH cross-reactivity. Immunoprecipitation reactions of different mycobacterial sonicates with anti-DTH-H37Rv showed that the antigen was shared by all the mycobacteria tested irrespective of their cross-reactivity in a DTH response. All of the slow growers showed reactions of total identity with DTH-H37Rv. However with rapid growers DTH-H37Rv showed only a partial identity. From these data it was concluded that an antigen participating in DTH response is shared by all mycobacteria and that it is polymorphous, having genus specific and group specific (as slow and rapid grower groups) determinants. Images Fig. 2 PMID:3082545

Rapid non-invasive tests for diagnostics of infectious diseases

NASA Astrophysics Data System (ADS)

Malamud, Daniel

2014-06-01

A rapid test for an infectious disease that can be used at point-of-care at a physician's office, a pharmacy, or in the field is critical for the prompt and appropriate therapeutic intervention. Ultimately by treating infections early on will decrease transmission of the pathogen. In contrast to metabolic diseases or cancer where multiple biomarkers are required, infectious disease targets (e.g. antigen, antibody, nucleic acid) are simple and specific for the pathogen causing the disease. Our laboratory has focused on three major infectious disease; HIV, Tuberculosis, and Malaria. These diseases are pandemic in much of the world thus putting natives, tourists and military personnel at risk for becoming infected, and upon returning to the U.S., transmitting these diseases to their contacts. Our devices are designed to detect antigens, antibodies or nucleic acids in blood or saliva samples in less than 30 minutes. An overview describing the current status of each of the three diagnostic platforms is presented. These microfluidic point-of-care devices will be relatively inexpensive, disposable, and user friendly.

Evaluation of ICT filariasis card test using whole capillary blood: comparison with Knott's concentration and counting chamber methods.

PubMed

Njenga, S M; Wamae, C N

2001-10-01

An immunochromatographic card test (ICT) that uses fingerprick whole blood instead of serum for diagnosis of bancroftian filariasis has recently been developed. The card test was validated in the field in Kenya by comparing its sensitivity to the combined sensitivity of Knott's concentration and counting chamber methods. A total of 102 (14.6%) and 117 (16.7%) persons was found to be microfilaremic by Knott's concentration and counting chamber methods, respectively. The geometric mean intensities (GMI) were 74.6 microfilariae (mf)/ml and 256.5 mf/ml by Knott's concentration and counting chamber methods, respectively. All infected individuals detected by both Knott's concentration and counting chamber methods were also antigen positive by the ICT filariasis card test (100% sensitivity). Further, of 97 parasitologically amicrofilaremic persons, 24 (24.7%) were antigen positive by the ICT. The overall prevalence of antigenemia was 37.3%. Of 100 nonendemic area control persons, none was found to be filarial antigen positive (100% specificity). The results show that the new version of the ICT filariasis card test is a simple, sensitive, specific, and rapid test that is convenient in field settings.

A multiplex immunochromatographic test using gold nanoparticles for the rapid and simultaneous detection of four nitrofuran metabolites in fish samples.

PubMed

Wang, Quan; Liu, Yingchun; Wang, Mingyan; Chen, Yongjun; Jiang, Wei

2018-01-01

There is an urgent need for the rapid and simultaneous detection of multiple analytes present in a sample matrix. Here, a multiplex immunochromatographic test (multi-ICT) was developed that successfully allowed for the rapid and simultaneous detection of four major nitrofuran metabolites, i.e., 3-amino-2-oxazolidinone (AOZ), semicarbazide (SEM), 3-amino-5-methylmorpholino-2-oxazolidinone (AMOZ), and 1-aminohydantoin (AHD), in fish samples. Four different antigens were separately immobilized in four test lines on a nitrocellulose membrane. Goat anti-mouse immunoglobulin (IgG) was used as a control. Sensitive and specific monoclonal antibodies (mAbs) that recognize the corresponding antigens were selected for the assay, and no cross-reactivity between the antibodies in the detection assay was observed. The free analytes in samples or standards were pre-incubated with freeze-dried mAb-gold conjugates to improve the sensitivity of the detection assay. The multi-ICT detection was accomplished in less than 15 min by the naked eye. The cutoff values for the strip test were 0.5 ng/mL for AOZ and 0.75 ng/mL for AHD, SEM, and AMOZ, which were all below the maximum residue levels set by the European Union and China. A high degree of consistency was observed between the multi-ICT method and commercially available enzyme-linked immunosorbent assay (ELISA) kits using spiked, incurred, and "blind" fish samples, indicating the accuracy, reproducibility, and reliability of the novel test strip. This newly developed multi-ICT strip assay is suitable for the rapid and high-throughput screening of four nitrofuran metabolites in fish samples on-site, with no treatment or devices required. Graphical abstract A multiplex immunochromatographic test (multi-ICT) was developed that successfully allowed for the rapid and simultaneous detection of four major nitrofuran metabolites (AOZ, SEM, AMOZ, and AHD) in fish samples.

Two cases of false-positive dengue non-structural protein 1 (NS1) antigen in patients with hematological malignancies and a review of the literature on the use of NS1 for the detection of Dengue infection.

PubMed

Chung, Shimin J; Krishnan, Prabha U; Leo, Yee Sin

2015-02-01

Early diagnosis of dengue has been made easier in recent years owing to the advancement in diagnostic technologies. The rapid non-structural protein 1 (NS1) test strip is widely used in many developed and developing regions at risk of dengue. Despite the relatively high specificity of this test, we recently encountered two cases of false-positive dengue NS1 antigen in patients with underlying hematological malignancies. We reviewed the literature for causes of false-positive dengue NS1. © The American Society of Tropical Medicine and Hygiene.

Etiologic predictive value of a rapid immunoassay for the detection of group A Streptococcus antigen from throat swabs in patients presenting with a sore throat.

PubMed

Orda, Ulrich; Gunnarsson, Ronny; Orda, Sabine; Fitzgerald, Mark; Rofe, Geoff; Dargan, Anna

2016-04-01

Clinical reasoning utilizing certain symptoms and scores has not proven to be a reliable decision-making tool to determine whether or not to suspect a group A Streptococcus (GAS) infection in the patient presenting with a sore throat. Culture as the so-called 'gold standard' is impracticable because it takes 1 to 2 days (and even longer in remote locations) for a result, and thus treatment decisions will be made without the result available. Rapid diagnostic antigen tests have demonstrated sufficient sensitivities and specificities in detecting GAS antigens to identify GAS throat infections. Throat swab samples were collected from patients attending the Mount Isa Hospital emergency department for a sore throat; these samples were compared to swab samples collected from healthy controls who did not have a sore throat. Both groups were aged 3-15 years. All swab samples were analyzed with a point-of-care test (Alere Test Pack +Plus with OBC Strep A). The etiologic predictive value (EPV) of the throat swab was calculated. The 95% confidence interval for positive EPV was 88-100% and for negative EPV was 97-99%, depending on assumptions made. This study demonstrates that the point-of-care test Alere Test Pack +Plus Strep A has a high positive predictive value and is able to rule in GAS infection as long as the proportion of carriers is low. Also the negative predictive value for ruling out GAS as the etiologic agent is very high irrespective of the carrier rate. Hence, this test is always useful to rule out GAS infection. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Determination of neuronal antibodies in suspected and definite Creutzfeldt-Jakob disease.

PubMed

Grau-Rivera, Oriol; Sánchez-Valle, Raquel; Saiz, Albert; Molinuevo, José Luis; Bernabé, Reyes; Munteis, Elvira; Pujadas, Francesc; Salvador, Antoni; Saura, Júlia; Ugarte, Antonio; Titulaer, Maarten; Dalmau, Josep; Graus, Francesc

2014-01-01

Creutzfeldt-Jakob disease (CJD) and autoimmune encephalitis with antibodies against neuronal surface antigens (NSA-abs) may present with similar clinical features. Establishing the correct diagnosis has practical implications in the management of care for these patients. To determine the frequency of NSA-abs in the cerebrospinal fluid of patients with suspected CJD and in patients with pathologically confirmed (ie, definite) CJD. A mixed prospective (suspected) and retrospective (definite) CJD cohort study was conducted in a reference center for detection of NSA-abs. The population included 346 patients with suspected CJD and 49 patients with definite CJD. Analysis of NSA-abs in cerebrospinal fluid with brain immunohistochemistry optimized for cell-surface antigens was performed. Positive cases in the suspected CJD group were further studied for antigen specificity using cell-based assays. All definite CJD cases were comprehensively tested for NSA-abs, with cell-based assays used for leucine-rich glioma-inactivated 1 (LGI1), contactin-associated protein-like 2 (CASPR2), N-methyl-d-aspartate (NMDA), and glycine (GlY) receptors. Neuronal surface antigens were detected in 6 of 346 patients (1.7%) with rapid neurologic deterioration suggestive of CJD. None of these 6 patients fulfilled the diagnostic criteria for probable or possible CJD. The target antigens included CASPR2, LGI1, NMDAR, aquaporin 4, Tr (DNER [δ/notch-like epidermal growth factor-related receptor]), and an unknown protein. Four of the patients developed rapidly progressive dementia, and the other 2 patients had cerebellar ataxia or seizures that were initially considered to be myoclonus without cognitive decline. The patient with Tr-abs had a positive 14-3-3 test result. Small cell lung carcinoma was diagnosed in the patient with antibodies against an unknown antigen. All patients improved or stabilized after appropriate treatment. None of the 49 patients with definite CJD had NSA-abs. A low, but clinically relevant, number of patients with suspected CJD had potentially treatable disorders associated with NSA-abs. In contrast, none of 49 patients with definite CJD had NSA-abs, including NMDAR-abs, GlyR-abs, LGI1-abs, or CASPR2-abs. These findings suggest that cerebrospinal fluid NSA-abs analysis should be included in the diagnostic workup of patients with rapidly progressive central nervous system syndromes, particularly when they do not fulfill the diagnostic criteria of probable or possible CJD.

Countrywide Reassessment of Schistosoma mansoni Infection in Burundi Using a Urine-Circulating Cathodic Antigen Rapid Test: Informing the National Control Program

PubMed Central

Ortu, Giuseppina; Ndayishimiye, Onésime; Clements, Michelle; Kayugi, Donatien; Campbell, Carl H.; Lamine, Mariama Sani; Zivieri, Antonio; Magalhaes, Ricardo Soares; Binder, Sue; King, Charles H.; Fenwick, Alan; Colley, Daniel G.; Jourdan, Peter Mark

2017-01-01

Following implementation of the national control program, a reassessment of Schistosoma mansoni prevalence was conducted in Burundi to determine the feasibility of moving toward elimination. A countrywide cluster-randomized cross-sectional study was performed in May 2014. At least 25 schools were sampled from each of five eco-epidemiological risk zones for schistosomiasis. Fifty randomly selected children 13–14 years of age per school were included for a single urine-circulating cathodic antigen (CCA) rapid test and, in a subset of schools, for duplicate Kato-Katz slide preparation from a single stool sample. A total of 17,331 children from 347 schools were tested using CCA. The overall prevalence of S. mansoni infection, when CCA trace results were considered negative, was 13.5% (zone range [zr] = 4.6–17.8%), and when CCA trace results were considered positive, it was 42.8% (zr = 34.3–49.9%). In 170 schools, prevalence of this infection determined using Kato-Katz method was 1.5% (zr ==0–2.7%). The overall mean intensity of S. mansoni infection determined using Kato-Katz was 0.85 eggs per gram (standard deviation = 10.86). A majority of schools (84%) were classified as non-endemic (prevalence = 0) using Kato-Katz; however, a similar proportion of schools were classified as endemic when CCA trace results were considered negative (85%) and nearly all (98%) were endemic when CCA trace results were considered positive. The findings of this nationwide reassessment using a CCA rapid test indicate that Schistosoma infection is still widespread in Burundi, although its average intensity is probably low. Further evidence is now needed to determine the association between CCA rapid test positivity and low-intensity disease transmission. PMID:28115675

Development of rapid immunochromatographic test for hemagglutinin antigen of H7 subtype in patients infected with novel avian influenza A (H7N9) virus.

PubMed

Kang, Keren; Chen, Li; Zhao, Xiang; Qin, Chengfeng; Zhan, Zanwu; Wang, Jihua; Li, Wenmei; Dzakah, Emmanuel E; Huang, Weijuang; Shu, Yuelong; Jiang, Tao; Cao, Wuchun; Xie, Mingquan; Luo, Xiaochun; Tang, Shixing

2014-01-01

Since human infection with the novel H7N9 avian influenza virus was identified in China in March 2013, the relatively high mortality rate and possibility of human-to-human transmission have highlighted the urgent need for sensitive and specific assays for diagnosis of H7N9 infection. We developed a rapid diagnostic test for the novel avian influenza A (H7N9) virus using anti-hemagglutinin (HA) monoclonal antibodies specifically targeting H7 in an immunochromatographic assay system. The assay limit of detection was 103.5 pfu/ml or 103TCID50 of H7N9 virus. The assay specifically detected H7N9 viral isolates and recombinant HA proteins of H7 subtypes including H7N7 and H7N9, but did not react with non-H7 subtypes including H1N1, H3N2, H5N1, H5N9, and H9N2. The detection sensitivity was 59.4% (19/32) for H7N9 patients confirmed by RT-PCR. Moreover, the highest sensitivity of 61.5% (16/26) was obtained when testing H7N9 positive sputum samples while 35.7% (5/14) of nasopharyngeal swabs and 20% (2/10) of fecal samples tested positive. No false positive detection was found when testing 180 H7N9 negative samples. Our novel rapid assay can specifically detect H7 HA antigen, facilitating rapid diagnosis for prevention and control of the on-going H7N9 epidemic.

Development of Rapid Immunochromatographic Test for Hemagglutinin Antigen of H7 Subtype in Patients Infected with Novel Avian Influenza A (H7N9) Virus

PubMed Central

Kang, Keren; Chen, Li; Zhao, Xiang; Qin, Chengfeng; Zhan, Zanwu; Wang, Jihua; Li, Wenmei; Dzakah, Emmanuel E.; Huang, Weijuang; Shu, Yuelong; Jiang, Tao; Cao, Wuchun; Xie, Mingquan; Luo, Xiaochun; Tang, Shixing

2014-01-01

Background Since human infection with the novel H7N9 avian influenza virus was identified in China in March 2013, the relatively high mortality rate and possibility of human-to-human transmission have highlighted the urgent need for sensitive and specific assays for diagnosis of H7N9 infection. Methodology/Principal Findings We developed a rapid diagnostic test for the novel avian influenza A (H7N9) virus using anti-hemagglutinin (HA) monoclonal antibodies specifically targeting H7 in an immunochromatographic assay system. The assay limit of detection was 103.5 pfu/ml or 103TCID50 of H7N9 virus. The assay specifically detected H7N9 viral isolates and recombinant HA proteins of H7 subtypes including H7N7 and H7N9, but did not react with non-H7 subtypes including H1N1, H3N2, H5N1, H5N9, and H9N2. The detection sensitivity was 59.4% (19/32) for H7N9 patients confirmed by RT-PCR. Moreover, the highest sensitivity of 61.5% (16/26) was obtained when testing H7N9 positive sputum samples while 35.7% (5/14) of nasopharyngeal swabs and 20% (2/10) of fecal samples tested positive. No false positive detection was found when testing 180 H7N9 negative samples. Conclusions/Significance Our novel rapid assay can specifically detect H7 HA antigen, facilitating rapid diagnosis for prevention and control of the on-going H7N9 epidemic. PMID:24647358

Evaluation of a rapid agglutination method for detection of equine red cell surface antigens (Ca and Aa) as part of pretransfusion testing.

PubMed

Owens, Sean D; Snipes, Joy; Magdesian, K Gary; Christopher, Mary M

2008-03-01

Blood typing before transfusion minimizes the risk of transfusion reactions and prevents immunization of the recipient against incompatible RBC antigens. The major RBC antigens that warrant identification before packed RBC or whole blood transfusions in horses are Ca and Aa. Standard blood-typing protocols are time-consuming (2.5-3.0 hours) and impractical in emergency settings. The purpose of this study was to determine whether equine RBCs could be typed for Ca and Aa antigens using sera from horses with RBC antibodies in a modified rapid (15 minute) blood-typing protocol. Serum was obtained from a horse with anti-Ca antibodies and from another horse with anti-Aa antibodies. The presence of agglutinating antibodies was confirmed with antibody screening. Venous blood samples, collected in citrate-phosphate-dextrose, were obtained from 21 horses of various breeds. Samples were blood typed in the Veterinary Medical Teaching Hospital Hematology Laboratory using standard methodology. Washed RBCs from each of the 21 horses were incubated individually with anti-Ca and anti-Aa sera at dilutions of 1:4, 1:8, and 1:16 for 15 and 30 minutes at room temperature and 37 degrees C. Of the 21 horses, 13 were identified as Aa+/Ca+, four were Aa+/Ca-, two were Aa-/Ca+, and two were Aa-/Ca-. All 17 Aa-positive horses had a positive agglutination reaction at all dilutions of anti-Aa serum, incubation times, and temperatures, while all Aa-negative horses were negative. Each Ca-positive horse had a positive agglutination reaction at all incubation time points and temperatures up to the 1:16 dilution of the anti-Ca serum. All Ca-negative horses were negative at all times, temperatures, and dilutions of anti-Ca serum. Use of the modified protocol on 26 hospitalized horses resulted in accurate typing, based on complete antibody screens. These results support the hypothesis that equine RBCs can be blood typed using a rapid (15 minute) protocol, at room temperature, for the presence of Ca and Aa antigens using equine-derived antisera. This technique may be beneficial for pretransfusion testing of equine patients in an emergency setting.

Comparative evaluation of the diagnostic potential of recombinant envelope proteins and native cell culture purified viral antigens of Chikungunya virus.

PubMed

Khan, Mohsin; Dhanwani, Rekha; Kumar, Jyoti S; Rao, P V Lakshmana; Parida, Manmohan

2014-07-01

Despite the fact that Chikungunya resurgence is associated with epidemic of unprecedented magnitude, there are challenges in the field of its clinical diagnosis. However, serological tests in an ELISA format provide a rapid tool for the diagnosis of Chikungunya infection. Indeed, ELISAs based on recombinant proteins hold a great promise as these methods are cost effective and are free from the risk of handling biohazardous material. In this study, the performance of recombinant CHIKV antigens was compared in various ELISA formats for the diagnosis of Chikungunya. Two recombinant antigens derived from the envelope proteins of Chikungunya virus were prepared and evaluated by comparing their competence for detecting circulating antibodies in serum samples of patients infected with CHIKV using MAC-ELISA and indirect IgM-ELISA. The efficacy of the recombinant antigens was also compared with the native antigen. The indirect antibody capture IgM microplate ELISA revealed ≥90% concordance with the native antigen in detecting the CHIKV specific IgM antibodies whereas the recombinant antigen based MAC-ELISA showed 100% specificity. The recombinant antigens used in this study were effective and reliable targets for the diagnosis of CHIKV infection and also provide an alternative for native antigen use which is potentially biohazardous. © 2013 Wiley Periodicals, Inc.

Development and testing of a rapid diagnostic test for bubonic and pneumonic plague.

PubMed

Chanteau, Suzanne; Rahalison, Lila; Ralafiarisoa, Lalao; Foulon, Jeanine; Ratsitorahina, Mahery; Ratsifasoamanana, Lala; Carniel, Elisabeth; Nato, Farida

2003-01-18

Plague is often fatal without prompt and appropriate treatment. It affects mainly poor and remote populations. Late diagnosis is one of the major causes of human death and spread of the disease, since it limits the effectiveness of control measures. We aimed to develop and assess a rapid diagnostic test (RDT) for plague. We developed a test that used monoclonal antibodies to the F1 antigen of Yersinia pestis. Sensitivity and specificity were assessed with a range of bacterial cultures and clinical samples, and compared with findings from available ELISA and bacteriological tests for plague. Samples from patients thought to have plague were tested with the RDT in the laboratory and by health workers in 26 pilot sites in Madagascar. The RDT detected concentrations of F1 antigen as low as 0.5 ng/mL in up to 15 min, and had a shelf life of 21 days at 60 degrees C. Its sensitivity and specificity were both 100%. RDT detected 41.6% and 31% more positive clinical specimens than did bacteriological methods and ELISA, respectively. The agreement rate between tests done at remote centres and in the laboratory was 89.8%. With the combination of bacteriological methods and F1 ELISA as reference standard, the positive and negative predictive values of the RDT were 90.6% and 86.7%, respectively. Our RDT is a specific, sensitive, and reliable test that can easily be done by health workers at the patient's bedside, for the rapid diagnosis of pneumonic and bubonic plague. This test will be of key importance for the control of plague in endemic countries.

A comparative study of blood smear, QBC and antigen detection for diagnosis of malaria.

PubMed

Parija, S C; Dhodapkar, Rahul; Elangovan, Subashini; Chaya, D R

2009-01-01

Rapid diagnosis is prerequisite for effective treatment and reducing mortality and morbidity of malaria. This study was taken up to compare the efficacy of various methods available, i.e., thick and thin smear, quantitative buffy coat (QBC), plasmodium lactate dehydrogenase and aldolase in blood of patient. A total of 411 samples were collected from patients presenting with classic symptoms of malaria. For traditional microscopy; thick and thin smears were prepared and stained with Leishman's stain, taking thick smear as gold standard, thin smear had a sensitivity and specificity of 54.8% and 100%, respectively. QBC and antigen detection was done using commercially available kits; out of 411 samples, QBC and Malariagen were positive in 66 and 62 cases, with a sensitivity of 78% and 75%, respectively. Leishman's thick smear, although cost effective, is difficult to interpret for inexperienced microscopists; so if facilities are available, QBC should be used for routine diagnosis. In places where facilities are not available, rapid, simple and easy to interpret antigen detection test can be used despite low sensitivity.

Use of a Dual-Antigen Rapid Diagnostic Test to Screen Children for Severe Plasmodium falciparum Malaria in a High-Transmission, Resource-Limited Setting.

PubMed

Boyce, Ross; Reyes, Raquel; Matte, Michael; Ntaro, Moses; Mulogo, Edgar; Siedner, Mark J

2017-10-16

In rural areas, many patients with malaria seek care at peripheral health facilities or community case management programs. While this strategy is effective for the management of uncomplicated malaria, severe malaria necessitates prompt detection and referral to facilities with adequate resources. In this prospective, observational cohort study, we assessed the accuracy of a dual-band (histidine-rich protein-2/pan-lactate dehydrogenase [HRP2/pLDH]) rapid diagnostic test (RDT) to differentiate uncomplicated from severe malaria. We included children aged <12 years who presented to a rural clinic in western Uganda with a positive HRP2 or HRP2/pLDH RDT. We estimated the test characteristics of a dual-antigen (HRP2+/pLDH+) band positive RDT compared to World Health Organization-defined clinical and laboratory criteria to detect severe malaria. A total of 2678 children underwent testing for malaria with an RDT, and 83 (9.0%) satisfied criteria for severe malaria. The sensitivity and specificity of a HRP2+/pLDH+ result for severe malaria was 97.6% (95% confidence interval [CI], 90.8%-99.6%) and 75.6% (95% CI, 73.8%-77.4%), respectively. An HRP2+/pLDH+ result was significantly more sensitive (97.6% vs 68.7%, P < .001) for the detection of severe malaria compared to algorithms that incorporate screening for danger signs. A positive dual-antigen (HRP2/pLDH) RDT has higher sensitivity than the use of clinical manifestations to detect severe malaria, making it a promising tool in the triage of children with malaria in low-resource settings. Additional work is needed to operationalize diagnostic and treatment algorithms that include dual-antigen RDTs to avoid over referral. © The Author 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Modification of the Rappaport rapid test in large-scale testing for syphilis. Evaluation of the rapid plate and rapid card tests.

PubMed

Ghinsberg, R; Meir, E; Blumstein, G; Kafeman, R

1975-11-01

The Rappaport rapid (RR) plate and card tests were developed as modifications of the RR tube test to permit rapid and inexpensive screening of large numbers of subjects for the diagnosis of syphilis. More than 2,000 sera were examined in parallel by the Venereal Disease Research Laboratory (VDRL) slide test, the rapid plasma reagin (RPR) card test and the RR plate and card tests. There was complete agreement between the RR plate and card tests and the VDRL slide and RPR card tests in 96.6% of sera. In a selected group of 1,530 sera examined, in addition, by the fluorescent treponemal antibody absorption (FTA-ABS) test, there was agreement between the RR plate and card tests and the FTA-ABS test in 74.3% of sera and between the VDRL and RPR tests and the FTA-ABS test in 73.7% of sera. The RR plate test was found to be sufficiently sensitive and specific for the diagnosis of syphilis, although the VDRL slide test is perhaps more sensitive in primary and late latent syphilis. Since the antigen used in the RR tests is colored and stable and the sera do not require inactivation before the test, the tests are easier to perform than the VDRL slide test: the RR plate and card tests could therefore replace the VDRL test as a screening test, with hardly any loss of accuracy.

A Protein-Conjugate Approach to Develop a Monoclonal Antibody-Based Antigen Detection Test for the Diagnosis of Human Brucellosis

PubMed Central

Patra, Kailash P.; Saito, Mayuko; Atluri, Vidya L.; Rolán, Hortensia G.; Young, Briana; Kerrinnes, Tobias; Smits, Henk; Ricaldi, Jessica N.; Gotuzzo, Eduardo; Gilman, Robert H.; Tsolis, Renee M.; Vinetz, Joseph M.

2014-01-01

Human brucellosis is most commonly diagnosed by serology based on agglutination of fixed Brucella abortus as antigen. Nucleic acid amplification techniques have not proven capable of reproducibly and sensitively demonstrating the presence of Brucella DNA in clinical specimens. We sought to optimize a monoclonal antibody-based assay to detect Brucella melitensis lipopolysaccharide in blood by conjugating B. melitensis LPS to keyhole limpet hemocyanin, an immunogenic protein carrier to maximize IgG affinity of monoclonal antibodies. A panel of specific of monoclonal antibodies was obtained that recognized both B. melitensis and B. abortus lipopolysaccharide epitopes. An antigen capture assay was developed that detected B. melitensis in the blood of experimentally infected mice and, in a pilot study, in naturally infected Peruvian subjects. As a proof of principle, a majority (7/10) of the patients with positive blood cultures had B. melitensis lipopolysaccharide detected in the initial blood specimen obtained. One of 10 patients with relapsed brucellosis and negative blood culture had a positive serum antigen test. No seronegative/blood culture negative patients had a positive serum antigen test. Analysis of the pair of monoclonal antibodies (2D1, 2E8) used in the capture ELISA for potential cross-reactivity in the detection of lipopolysaccharides of E. coli O157:H7 and Yersinia enterocolitica O9 showed specificity for Brucella lipopolysaccharide. This new approach to develop antigen-detection monoclonal antibodies against a T cell-independent polysaccharide antigen based on immunogenic protein conjugation may lead to the production of improved rapid point-of-care-deployable assays for the diagnosis of brucellosis and other infectious diseases. PMID:24901521

Genome-wide screening and identification of antigens for rickettsial vaccine development

USDA-ARS?s Scientific Manuscript database

The capacity to identify immunogens for vaccine development by genome-wide screening has been markedly enhanced by the availability of complete microbial genome sequences coupled to rapid proteomic and bioinformatic analysis. Critical to this genome-wide screening is in vivo testing in the context o...

Evaluation of a Cryptococcal antigen Lateral Flow Assay in serum and cerebrospinal fluid for rapid diagnosis of cryptococcosis in Colombia.

PubMed

Cáceres, Diego H; Zuluaga, Alejandra; Tabares, Ángela M; Chiller, Tom; González, Ángel; Gómez, Beatriz L

2017-12-21

A Lateral Flow Assay to detect cryptococcal antigen (CrAg® LFA) in serum and cerebrospinal fluid for the rapid diagnosis of cryptococcosis was evaluated. A retrospective validation was performed. Sensitivity and specificity of the CrAg® LFA was 100%. High concordance (kappa index=1.0) between Cryptococcal Antigen Latex Agglutination System (CALAS®) and CrAg® LFA was observed. CrAg® LFA showed higher analytical sensitivity for detecting low concentrations of cryptococcal antigen.

Evaluation of a Cryptococcal antigen Lateral Flow Assay in serum and cerebrospinal fluid for rapid diagnosis of cryptococcosis in Colombia

PubMed Central

Cáceres, Diego H.; Zuluaga, Alejandra; Tabares, Ángela M.; Chiller, Tom; González, Ángel; Gómez, Beatriz L.

2017-01-01

ABSTRACT A Lateral Flow Assay to detect cryptococcal antigen (CrAg® LFA) in serum and cerebrospinal fluid for the rapid diagnosis of cryptococcosis was evaluated. A retrospective validation was performed. Sensitivity and specificity of the CrAg® LFA was 100%. High concordance (kappa index=1.0) between Cryptococcal Antigen Latex Agglutination System (CALAS®) and CrAg® LFA was observed. CrAg® LFA showed higher analytical sensitivity for detecting low concentrations of cryptococcal antigen. PMID:29267584

Natural selection promotes antigenic evolvability.

PubMed

Graves, Christopher J; Ros, Vera I D; Stevenson, Brian; Sniegowski, Paul D; Brisson, Dustin

2013-01-01

The hypothesis that evolvability - the capacity to evolve by natural selection - is itself the object of natural selection is highly intriguing but remains controversial due in large part to a paucity of direct experimental evidence. The antigenic variation mechanisms of microbial pathogens provide an experimentally tractable system to test whether natural selection has favored mechanisms that increase evolvability. Many antigenic variation systems consist of paralogous unexpressed 'cassettes' that recombine into an expression site to rapidly alter the expressed protein. Importantly, the magnitude of antigenic change is a function of the genetic diversity among the unexpressed cassettes. Thus, evidence that selection favors among-cassette diversity is direct evidence that natural selection promotes antigenic evolvability. We used the Lyme disease bacterium, Borrelia burgdorferi, as a model to test the prediction that natural selection favors amino acid diversity among unexpressed vls cassettes and thereby promotes evolvability in a primary surface antigen, VlsE. The hypothesis that diversity among vls cassettes is favored by natural selection was supported in each B. burgdorferi strain analyzed using both classical (dN/dS ratios) and Bayesian population genetic analyses of genetic sequence data. This hypothesis was also supported by the conservation of highly mutable tandem-repeat structures across B. burgdorferi strains despite a near complete absence of sequence conservation. Diversification among vls cassettes due to natural selection and mutable repeat structures promotes long-term antigenic evolvability of VlsE. These findings provide a direct demonstration that molecular mechanisms that enhance evolvability of surface antigens are an evolutionary adaptation. The molecular evolutionary processes identified here can serve as a model for the evolution of antigenic evolvability in many pathogens which utilize similar strategies to establish chronic infections.

Natural Selection Promotes Antigenic Evolvability

PubMed Central

Graves, Christopher J.; Ros, Vera I. D.; Stevenson, Brian; Sniegowski, Paul D.; Brisson, Dustin

2013-01-01

The hypothesis that evolvability - the capacity to evolve by natural selection - is itself the object of natural selection is highly intriguing but remains controversial due in large part to a paucity of direct experimental evidence. The antigenic variation mechanisms of microbial pathogens provide an experimentally tractable system to test whether natural selection has favored mechanisms that increase evolvability. Many antigenic variation systems consist of paralogous unexpressed ‘cassettes’ that recombine into an expression site to rapidly alter the expressed protein. Importantly, the magnitude of antigenic change is a function of the genetic diversity among the unexpressed cassettes. Thus, evidence that selection favors among-cassette diversity is direct evidence that natural selection promotes antigenic evolvability. We used the Lyme disease bacterium, Borrelia burgdorferi, as a model to test the prediction that natural selection favors amino acid diversity among unexpressed vls cassettes and thereby promotes evolvability in a primary surface antigen, VlsE. The hypothesis that diversity among vls cassettes is favored by natural selection was supported in each B. burgdorferi strain analyzed using both classical (dN/dS ratios) and Bayesian population genetic analyses of genetic sequence data. This hypothesis was also supported by the conservation of highly mutable tandem-repeat structures across B. burgdorferi strains despite a near complete absence of sequence conservation. Diversification among vls cassettes due to natural selection and mutable repeat structures promotes long-term antigenic evolvability of VlsE. These findings provide a direct demonstration that molecular mechanisms that enhance evolvability of surface antigens are an evolutionary adaptation. The molecular evolutionary processes identified here can serve as a model for the evolution of antigenic evolvability in many pathogens which utilize similar strategies to establish chronic infections. PMID:24244173

Diagnostic, therapeutic and economic consequences of a positive urinary antigen test for Legionella spp. in patients admitted with community-acquired pneumonia: a 7-year retrospective evaluation.

PubMed

Engel, M F; van Manen, L; Hoepelman, A I M; Thijsen, S; Oosterheert, J J

2013-09-01

A positive urinary antigen test for Legionella spp. (Legionella urinary antigen test; LUAT) allows an early switch from empiric to targeted treatment (TT) in hospitalised, community-acquired pneumonia (CAP) patients. We aimed to evaluate the diagnostic, therapeutic and economic consequences of this frequently used test 7 years after its implementation. We retrospectively evaluated LUATs performed between 2005 and 2011 in two teaching hospitals. All tests performed in hospitalised CAP patients were used in the economic evaluation and positive tests were included in the treatment evaluation. Data on patient characteristics, admission and outcome were retrieved from the patients' files. The number of days gained by making a rapid aetiological diagnosis, the number of days TT could be provided and their costs were calculated. Of 4485 LUATs, 2504 (56%) were performed for CAP including 55 (1%) positive tests (€1041/positive test). In 26 (60%) of the 43 included positive tests, LUAT was the only test showing Legionella spp. Subsequently, earlier TT was possible in the remaining cases during 209 cumulative admission days (€274/TT day). LUAT led to detection of Legionella spp. 13 days earlier per case (€203/day) as compared with culture/serology alone. Timely LUAT use in accordance with current guidelines allows early detection and treatment of CAP caused by Legionella spp. at considerable expense.

Ultrasensitive immunochromatographic assay for the simultaneous detection of five chemicals in drinking water.

PubMed

Xing, Changrui; Liu, Liqiang; Song, Shanshan; Feng, Min; Kuang, Hua; Xu, Chuanlai

2015-04-15

In this paper, we describe the development of a multicomponent lateral-flow assay based on an antibody-antigen reaction for the rapid and simultaneous detection of trace contaminants in water, including a heavy metal, algal toxin, antibiotic, hormone, and pesticide. The representative analytes chosen for the study were lead (Pb(II), microcystin-leucine-arginine (MC-LR), chloramphenicol (CAP), testosterone (T), and chlorothalonil (CTN). Five different antigens were immobilized separately in five test lines on a nitrocellulose membrane. The monoclonal antibodies specifically recognized the corresponding antigens, and there was no cross-reactivity between the antibodies in the detection assay. Samples or standards containing the five analytes were preincubated with the freeze-dried colloidal-gold-labeled monoclonal antibody conjugates to improve the sensitivity of the assay. The results were obtained within 20min with a paper-based sensor. The cut-off values for the strip test were 4ng/mL for Pb(II), 1ng/mL for MC-LR, 0.1ng/mL for CAP, 5ng/mL for T, and 5ng/mL for CTN. The assay was evaluated using spiked water samples, and the accuracy and reproducibility of the results were good. In summary, this lateral-flow device provides an effective and rapid method for the onsite detection of multiple contaminants in water samples, with no treatment or devices required. Copyright © 2014 Elsevier B.V. All rights reserved.

Coproantigens in taeniasis and echinococcosis.

PubMed

Allan, James C; Craig, Philip S

2006-01-01

The application of modern immunodiagnostic or molecular diagnostic techniques has improved the diagnosis of the taeniid cestode infections, echinococcosis and taeniasis. One particularly promising approach is the detection of parasite-specific antigens in faeces (coproantigens). This approach has been applied to both Echinoccocus and Taenia species and it has gained increasingly widespread use. Taeniid coproantigen tests are based on either monoclonal or polyclonal antibodies raised against adult tapeworm antigens. These tests have the following common characteristics; they are largely genus-specific, specificity is high (>95%), parasite antigen can be detected in faeces weeks prior to patency, levels of coproantigen are independent of egg output, coproantigen is stable for days at a range of temperatures (-80 degrees C to 35 degrees C), for several months in formalin-fixed faecal samples, and coproantigen levels drop rapidly (1-5 days) following successful treatment. In the genus Taenia, most work has been done on Taenia solium and coproantigen tests have reliably detected many more tapeworm carriers than microscopy. For Echinococcus species, there is a broad positive correlation between test sensitivity and worm burden with a reliable threshold level for the test of >50 worms. Characterisation of taeniid coproantigens in order to further improve the tests is ongoing. Studies indicate taeniid coproantigens to include high molecular weight (>150 kDa), heavily glycosylated molecules with carbohydrate moieties contributing substantially to the levels of antigen detected in faeces. Application of the existing coproantigen tests in epidemiological and control programmes for Echinococcus and Taenia species infection has begun to contribute to an improved understanding of transmission and of surveillance of these important zoonotic cestodes.

Utility of a stool antigen test to detect the incidence of helicobacter pylori infection and familial and community enviromental risk factors for this infection in pediatric age.

PubMed

Sabbi, T; Dall'Oglio, L; De Angelis, P; Torroni, E; Colistro, F; Azzolina, M; Santoni, A; Di Ciommo, V; Benedetto, M

2012-01-01

Helicobacter pylori (Hp) infection is mainly acquired during childhood; it is recognised as a cause of gastritis and peptic ulcer and it has been classified as a group A carcinogen by World Health Organization. The exact mode of transmission is as yet, not known. Aim of our study has been to identify risk factors associated with Helicobacter pylori infection in a preschool and school population and to confirm if Hp antigen in faeces is useful as screening in epidemiological studies. We interviewed, with questionnaire, 400 children (203 male; age range 3-10 years; mean age 6 years) of 3 different schools and stool samples were collected of all children too. 35 of 400 (8%) children underwent to upper gastrointestinal endoscopy because of a suspect of upper gastrointestinal disease. stool were collected from 400 school children and 35 of them shown positivity of Hp antigen test. A questionnaire about presence of nausea, vomit, recurrent abdominal pain, family size, parent's occupations and education, use of antibiotics, country of birth of child and parents, personal hygiene, breast feeding, presence of the animals was completed. 35 children with positive Hp stool antigen test and a suspicious of upper gastrointestinal disease (recurrent abdominal pain, diurnal or nocturnal abdominal pain, nausea, vomiting, iron deficiency) underwent to esophagogastroduodenoscopy (EGDS) that demonstrated antral gastritis and positive histology and urease rapid test. the results of this study suggest that risk factors for Hp infection are low socioeconomics factors, hygiene and living conditions and that Hp antigen in faeces is useful as screening test.

Rapid classification of biological components

DOEpatents

Thompson, Vicki S.; Barrett, Karen B.; Key, Diane E.

2013-10-15

A method is disclosed for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an illustrative embodiment of the invention, the analyte is a drug, such as marijuana, cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method involves attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein the locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to antigens in the array, thereby forming immune complexes; washing away antibodies that do not form immune complexes; and detecting the immune complexes, thereby forming an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to a subject's identity.

Rapid classification of biological components

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thompson, Vicki S.; Barrett, Karen B.; Key, Diane E.

A method is disclosed for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an illustrative embodiment of the invention, the analyte is a drug, such as marijuana, cocaine, methamphetamine, methyltestosterone, or mesterolone. The method involves attaching antigens to the surface of a solid support in a preselected pattern to form an array wherein the locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to antigens in the array, thereby forming immunemore » complexes; washing away antibodies that do form immune complexes; and detecting the immune complexes, thereby forming an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to the subject's identity.« less

[Membrane-filtration immunoassay: reagents, methods and the diagnostic and technical means for detection].

PubMed

Khramov, E N; Osin, N S; Pomelova, V G; Vikha, I V; Bychenkova, T A; Smirnova, V G; Grakina, G I; Kas'ianova, T A

1999-01-01

The comprehensive development of dot-EIA made at the State Research Institute of Biological Instrument-Making Industry has provided devices KIMF-02 and KIMF-03), a base of chemical reagents, immunoassays, test systems for detection of a wide range of causative agents of viral and bacterial infections, that of serodiagnosis of their related diseases. The KIMF-02 kit has undergone engineering and medical tests and recommended for the Ministry of Health of the Russian Federation to produce them in stock. The kit includes all required for analysis even in an ill-equipped laboratory, a set of attached agents ensures a valid visual recording of results. The developed procedures and test systems allow the immunoassay to be as sensitive as TIFA; however, they are laborious and much simpler in design. The simple and rapid procedures of dot-EIA are recommended for incorporation into the a package of laboratory methods for verification of the accumulation of virus-specific antigens in various biological substrata, environmental samples, for control of the activity of antigens and antibodies used in serological tests, for detection of specific antigens in the clinical samples, and for serodiagnosis of infections.

Rapid and selective detection of botulinum neurotoxin serotypes-A and –B with a single immunochromatographic test strip

USDA-ARS?s Scientific Manuscript database

Seven, antigenically distinct botulinum neurotoxins (BoNT) are produced by the bacterium Clostridium botulinum and classified into serotypes designated A-G. In animals these potent toxin acts to inhibit acetylcholine release, resulting in paralysis and death. BoNT/A and /B, together represent >80% o...

ANTIGENIC MODULATION

PubMed Central

Old, Lloyd J.; Stockert, Elisabeth; Boyse, Edward A.; Kim, Jae Ho

1968-01-01

Antigenic modulation (the loss of TL antigens from TL+ cells exposed to TL antibody in the absence of lytic complement) has been demonstrated in vitro. An ascites leukemia, phenotype TL.1,2,3, which modulates rapidly and completely when incubated with TL antiserum in vitro, was selected for further study of the phenomenon. Over a wide range of TL antibody concentrations modulation at 37°C was detectable within 10 min and was complete within approximately 1 hr. The cells were initially sensitized to C' by their contact with antibody, thereafter losing this sensitivity to C' lysis together with their sensitivity to TL antibody and C' in the cytotoxic test. The capacity of the cells to undergo modulation was abolished by actinomycin D and by iodoacetamide, and by reducing the temperature of incubation to 0°C. Thus modulation apparently is an active cellular process. Antigens TL. 1,2, and 3 are all modulated by anti-TL.1,3 serum and by anti-TL.3 serum. This modulation affects all three TL components together, even when antibody to one or two of them is lacking. aAnti-TL.2 serum does not induce modulation and in fact impairs modulation by the other TL antibodies. The influence of the TL phenotype of cells upon the demonstrable content of H-2 (D region) isoantigen, first shown in cells modulated in vivo, has been observed with cells modulated in vitro. Cells undergoing modulation show a progressive increase in H-2 (D region) antigen over a period of 4 hr, with no change in H-2 antigens of the K region. Restoration of the TL+ phenotype of modulated cells after removal of antibody is less rapid than TL+ → TL- modulation and may require several cell divisions. PMID:5636556

Prevalence of Plasmodium falciparum transmission reducing immunity among primary school children in a malaria moderate transmission region in Zimbabwe.

PubMed

Paul, Noah H; Vengesai, Arthur; Mduluza, Takafira; Chipeta, James; Midzi, Nicholas; Bansal, Geetha P; Kumar, Nirbhay

2016-11-01

Malaria continues to cause alarming morbidity and mortality in more than 100 countries worldwide. Antigens in the various life cycle stages of malaria parasites are presented to the immune system during natural infection and it is widely recognized that after repeated malaria exposure, adults develop partially protective immunity. Specific antigens of natural immunity represent among the most important targets for the development of malaria vaccines. Immunity against the transmission stages of the malaria parasite represents an important approach to reduce malaria transmission and is believed to become an important tool for gradual elimination of malaria. Development of immunity against Plasmodium falciparum sexual stages was evaluated in primary school children aged 6-16 years in Makoni district of Zimbabwe, an area of low to modest malaria transmission. Malaria infection was screened by microscopy, rapid diagnostic tests and finally using nested PCR. Plasma samples were tested for antibodies against recombinant Pfs48/45 and Pfs47 by ELISA. Corresponding serum samples were used to test for P. falciparum transmission reducing activity in Anopheles stephensi and An. gambiae mosquitoes using the membrane feeding assay. The prevalence of malaria diagnosed by rapid diagnostic test kit (Paracheck)™ was 1.7%. However, of the randomly tested blood samples, 66% were positive by nested PCR. ELISA revealed prevalence (64% positivity at 1:500 dilution, in randomly selected 66 plasma samples) of antibodies against recombinant Pfs48/45 (mean A 405nm=0.53, CI=0.46-0.60) and Pfs47 (mean A405nm=0.91, CI=0.80-1.02); antigens specific to the sexual stages. The mosquito membrane feeding assay demonstrated measurable transmission reducing ability of the samples that were positive for Pfs48/45 antibodies by ELISA. Interestingly, 3 plasma samples revealed enhancement of infectivity of P. falciparum in An. stephensi mosquitoes. These studies revealed the presence of antibodies with transmission reducing immunity in school age children from a moderate transmission area of malaria, and provide further support to exploit target antigens such as Pfs48/45 for further development of a malaria transmission blocking vaccine. Copyright © 2016 Elsevier B.V. All rights reserved.

[Efficiency and specificity of the KAT-test for rapid diagnosis of falciparum malaria].

PubMed

Cong, Le Dinh; Sergiev, V P; Rabinovich, S A; Nhah, Doan Hanh; Huong, Nguyen Van; Morozov, E N; Kukina, I V; Thinh, Ta Thi; Maksakovskaia, E V; Dao, Le Minh; Chalyĭ, V F; To, Dang Thi; Fandeev, V A; Hoa, Ngo Viet; Due, Nguyen Thi

2002-01-01

A new rapid KAT Quick Malaria test for the diagnosis of falciparum malaria, which is based on the detection of a monoclonal antibody-antigen complex of malaria parasites, has been worked out by the KAT Medical CC in South Africa. The efficiency and specificity of the KAT test were compared with those of the microscopic method and with the ICT test for rapid diagnosis of P. falciparum and P. vivax. The polymerase chain reaction was used as a control test. Testing for malaria was performed on 98 blood samples from feverish patients in Vietnam and Tadjikistan and among the persons who had returned to Moscow from endemic regions. The efficiency of the KAT test for falciparum-malaria was found to be 100% versus 90.5% with ICT. The absence of cross-reactions with P. vivax and the presence of pseudopositive results of the KAT test for fever cases of non-malaria origin indicate its high specificity. There was no correlation between the rate of test line colouring and the level of parasitemia. The KAT test yielded positive results only when gametocytes were found in blood specimens.

Fluorescence polarization immunoassays for rapid, accurate and sensitive determination of mycotoxins

USDA-ARS?s Scientific Manuscript database

Fluorescence polarization immunoassay (FPIA) is a type of homogeneous assay. For low molecular weight antigens, such as mycotoxins, it is based on the competition between an unlabeled antigen and its fluorescent-labeled derivative (tracer) for an antigen-specific antibody. The antigen content is det...

AntigenMap 3D: an online antigenic cartography resource.

PubMed

Barnett, J Lamar; Yang, Jialiang; Cai, Zhipeng; Zhang, Tong; Wan, Xiu-Feng

2012-05-01

Antigenic cartography is a useful technique to visualize and minimize errors in immunological data by projecting antigens to 2D or 3D cartography. However, a 2D cartography may not be sufficient to capture the antigenic relationship from high-dimensional immunological data. AntigenMap 3D presents an online, interactive, and robust 3D antigenic cartography construction and visualization resource. AntigenMap 3D can be applied to identify antigenic variants and vaccine strain candidates for pathogens with rapid antigenic variations, such as influenza A virus. http://sysbio.cvm.msstate.edu/AntigenMap3D

Evaluation of two new commercial tests for the diagnosis of acute dengue virus infection using NS1 antigen detection in human serum.

PubMed

Dussart, Philippe; Petit, Laure; Labeau, Bhety; Bremand, Laetitia; Leduc, Alexandre; Moua, David; Matheus, Séverine; Baril, Laurence

2008-08-20

We compared the performance of two new commercial tests for the detection of dengue NS1 protein during the clinical phase of dengue virus (DENV) infection-an immunochromatographic test allowing rapid detection of the NS1 antigen, Dengue NS1 Ag STRIP (Bio-Rad Laboratories - Marnes La Coquette, France), and a two-step sandwich-format microplate enzyme-linked immunosorbent assay (ELISA), pan-E Dengue Early ELISA (Panbio - Brisbane, Australia)-with a one-step sandwich-format microplate ELISA, the Platelia Dengue NS1 Ag test (Bio-Rad). We tested 272 serum samples from patients with dengue disease. Of these, 222 were from patients with acute infection of one of the four dengue serotypes, detected by RT-PCR and/or virus isolation. Forty-eight acute-phase serum samples from patients not infected with dengue virus were also included. The sensitivity of the Platelia Dengue NS1 Ag test on acute serum samples (n = 222) was 87.4% (95% confidence interval: 82.3% to 91.5%); that of Dengue NS1 Ag STRIP was 81.5% (95% CI: 75.8% to 86.4%) after 15 minutes and 82.4% (95% CI: 76.8% to 87.2%) after 30 minutes. Both tests had a specificity of 100% (97.5% CI, one-sided test: 92.6% to 100.0%). The pan-E Dengue Early ELISA had a sensitivity of 60.4% (95% CI: 53.4% to 66.8%) and a specificity of 97.9% (95% CI: 88.9% to 99.9%). Our findings support the use of diagnostic tools based on the NS1 antigen detection for the diagnosis of acute DENV infection. The immunochromatographic test, Dengue NS1 Ag STRIP-the first rapid diagnostic test for DENV infection-was highly sensitive and specific, and would therefore be a suitable first-line test in the field. The pan-E Dengue Early ELISA was less sensitive than the Platelia test; this two-step ELISA should be combined with DENV IgM antibody detection for the diagnosis of DENV infection.

Rapid detection of hepatitis B virus surface antigen by an agglutination assay mediated by a bispecific diabody against both human erythrocytes and hepatitis B virus surface antigen.

PubMed

Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi

2007-06-01

Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens.

[Effect of immune modulation on immunogenic and protective activity of a live plague vaccine].

PubMed

Karal'nik, B V; Ponomareva, T S; Deriabin, P N; Denisova, T G; Mel'nikova, N N; Tugambaev, T I; Atshabar, B B; Zakarian, S B

2014-01-01

Comparative evaluation of the effect of polyoxidonium and betaleukin on immunogenic and protective activity of a live plague vaccine in model animal experiments. Plague vaccine EV, polyoxidonium, betaleukin, erythrocytic antigenic diagnosticum for determination of F1 antibodies and immune reagents for detection of lymphocytes with F1 receptors (LFR) in adhesive test developed by the authors were used. The experiments were carried out in 12 rabbits and 169 guinea pigs. Immune modulation accelerated the appearance and disappearance of LFR (early phase) and ensured a more rapid and intensive antibody formation (effector phase). Activation by betaleukin is more pronounced than by polyoxidonium. The more rapid and intensive was the development of early phase, the more effective was antibody response to the vaccine. Immune modulation in the experiment with guinea pigs significantly increased protective activity of the vaccine. The use of immune modulators increased immunogenic (in both early and effector phases of antigen-specific response) and protective activity of the EV vaccine. A connection between the acceleration of the first phase of antigen-specific response and general intensity of effector phase of immune response to the EV vaccine was detected. ,

EBOLA Ag K-SeT rapid test: field evaluation in Sierra Leone.

PubMed

Colavita, F; Biava, M; Mertens, P; Gilleman, Q; Borlon, C; Delli Guanti, M; Petrocelli, A; Cataldi, G; Kamara, A T; Kamara, S A; Konneh, K; Vincenti, D; Castilletti, C; Abdurahman, S; Mirazimi, A; Capobianchi, M R; Ippolito, G; Miccio, R; Di Caro, A

2018-06-01

Efficient interruption of Ebola virus disease (EVD) transmission chains critically depends on reliable and fast laboratory diagnosis. We evaluated the performance of the EBOLA Virus Antigen Detection K-SeT (EBOLA Ag K-SeT), a new rapid diagnostic antigen test in field settings. The study was conducted in a field laboratory located in Freetown (Sierra Leone) by the Italian National Institute for Infectious Diseases 'L. Spallanzani' and the EMERGENCY Onlus NGO. The EBOLA Ag K-SeT was tested on 210 residual plasma samples (EVD prevalence 50%) from patients hospitalized at the EMERGENCY Ebola treatment center in Goderich (Freetown), comparing the results with quantitative real-time PCR. Overall, the sensitivity of EBOLA Ag K-SeT was 88.6% (95% confidence interval (CI), 82.5-94.7), and the corresponding specificity was 98.1% (95% CI, 95.5-100.7). The positive and negative predictive values were 97.9% (95% CI, 95.0-100.8) and 89.6% (95% CI, 84-95.2), respectively. The sensitivity strongly increased up to 98.7% (95% CI, 96.1-101.2) for those samples with high virus load (≥6.2 log RNA copies/mL). Our results suggest that EBOLA Ag K-SeT could represent a new effective diagnostic tool for EVD, meeting a need for resource-poor settings and rapid diagnosis for individuals with suspected EVD. Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Rapid Diagnosis of Arbovirus and Arenavirus Infections by Immunofluorescence.

DTIC Science & Technology

1984-12-31

rivers have been tested against Ebola, Lassa and Marburg viruses . Only positives with Ebola virus were found with the monovalent slides. One serum gave...recognition as a disease entity. DIVELOPMK OF THE ELISA TEST FOR CCHF VIRUSES . We have used detected CCHF virus infected cells by ELISA. This system offers...CCHF) virus was developed using infected, formalin-fixed CER cells as antigen. A retrospective serologic survey of equatorial Africa for antibodies

Evaluation of recombinant LigB antigen-based indirect ELISA and latex agglutination test for the serodiagnosis of bovine leptospirosis in India.

PubMed

Deneke, Yosef; Sabarinath, T; Gogia, Neha; Lalsiamthara, Jonathan; Viswas, K N; Chaudhuri, Pallab

2014-08-01

Leptospirosis is a zoonotic disease caused by pathogenic spirochetes of the genus Leptospira, causing febrile infection characterized by multi-organ failure in humans and animals. Leptospiral Ig-like protein B (LigB) is a surface-expressed antigen that mediates host cell invasion or attachment. In this study, N-terminal conserved region of LigB protein (46 kDa) was evaluated for its diagnostic potential to detect anti-leptospiral antibodies in the sera of various animal species. Dot blot analysis revealed immunoreactivity of Leptospira-positive sera of cattle, buffalo, dog, sheep and goat to purified LigB protein. We have analyzed 1126 bovine serum samples, collected from Northern and Eastern part of India, by microscopic agglutination test (MAT) and recombinant LigB (rLigB) based ELISA and latex agglutination test (LAT). The sensitivity of rLigB based ELISA for 554 MAT positive sera was 96.9% and the specificity with 572 MAT negative sera was 91.08% whereas LAT showed sensitivity and specificity of 93.68% and 92.31%, respectively. Kappa values of 0.879 and 0.860 for recombinant antigen based ELISA and LAT indicate excellent agreement with the gold standard serological test, MAT, for the detection of anti-leptospiral antibodies in sera. Further, LAT based on rLigB antigen is a simple and rapid test, suitable for serodiagnosis of leptospirosis under field conditions, owing to its portability and longer shelf life. Copyright © 2014 Elsevier Ltd. All rights reserved.

Evaluation of a surface plasmon resonance imaging-based multiplex O-antigen serogrouping for Escherichia coli using eleven major serotypes of Shiga -toxin-producing E. coli.

PubMed

Nakano, Satoshi; Nagao, Miki; Yamasaki, Tomomi; Morimura, Hiroyuki; Hama, Natsuki; Iijima, Yoshio; Shinomiya, Hiroto; Tanaka, Michio; Yamamoto, Masaki; Matsumura, Yasufumi; Miyake, Shiro; Ichiyama, Satoshi

2018-06-01

The early detection of Shiga toxin-producing Escherichia coli (STEC) is important for early diagnosis and preventing the spread of STEC. Although the confirmatory test for STEC should be based on the detection of Shiga toxin using molecular analysis, isolation permits additional characterization of STEC using a variety of methods, including O:H serotyping. The conventional slide agglutination O-antigen serogrouping used in many clinical laboratories is laborious and time-consuming. Surface plasmon resonance (SPR)-based immunosensors are commonly used to investigate a large variety of bio-interactions such as antibody/antigen, peptide/antibody, DNA/DNA, and antibody/bacteria interactions. SPR imaging (SPRi) is characterized by multiplexing capabilities for rapidly screening (approximately 100 to several hundred sensorgrams in parallel) molecules. SPRi-based O-antigen serogrouping method for STEC was recently developed by detecting the interactions between O-antigen-specific antibodies and bacterial cells themselves. The aim of this study was to evaluate its performance for E. coli serogrouping using clinical STEC isolates by comparing the results of slide agglutination tests. We tested a total of 188 isolates, including O26, O45, O91, O103, O111, O115, O121, O128, O145, O157, and O159. The overall sensitivity of SPRi-based O-antigen serogrouping was 98.9%. Only two O157 isolates were misidentified as nontypeable and O121. The detection limits of all serotypes were distributed between 1.1 × 10 6 and 17.6 × 10 6  CFU/ml. Pulsed-field gel electrophoresis (PFGE) revealed the heterogeneity of the examined isolates. In conclusion, SPRi is a useful method for the O-antigen serogrouping of STEC isolates, but the further evaluation of non-O157 minor serogroups is needed. Copyright © 2018 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

γδ T cells recognize a microbial encoded B cell antigen to initiate a rapid antigen specific Interleukin 17 response

PubMed Central

Zeng, Xun; Wei, Yu-ling; Huang, Jun; Newell, Evan W.; Yu, Hongxiang; Kidd, Brian A.; Kuhns, Michael S.; Waters, Ray W.; Davis, Mark M.; Weaver, Casey T.; Chien, Yueh-hsiu

2012-01-01

Summary γδ T cells contribute uniquely to host immune defense. However, how they function remains an enigma. Although it is unclear what most γδ T cells recognize, common dogma asserts that they recognize self-antigens. While they are the major initial Interleukin-17 (IL-17) producers in infections, it is unclear what is required to trigger these cells to act. Here, we report that a noted B cell antigen, the algae protein-phycoerythrin (PE) is an antigen for murine and human γδ T cells. PE also stained specific bovine γδ T cells. Employing this specificity, we demonstrated that antigen recognition, but not extensive clonal expansion, was required to activate naïve γδ T cells to make IL-17. In this activated state, γδ T cells gained the ability to respond to cytokine signals that perpetuated the IL-17 production. These results underscore the adaptability of lymphocyte antigen receptors and suggest a previously unrecognized antigen-driven rapid response in protective immunity prior to the maturation of classical adaptive immunity. PMID:22960222

Testing Eurasian wild boar piglets for serum antibodies against Mycobacterium bovis.

PubMed

Che' Amat, A; González-Barrio, D; Ortiz, J A; Díez-Delgado, I; Boadella, M; Barasona, J A; Bezos, J; Romero, B; Armenteros, J A; Lyashchenko, K P; Venteo, A; Rueda, P; Gortázar, C

2015-09-01

Animal tuberculosis (TB) caused by infection with Mycobacterium bovis and closely related members of the M. tuberculosis complex (MTC), is often reported in the Eurasian wild boar (Sus scrofa). Tests detecting antibodies against MTC antigens are valuable tools for TB monitoring and control in suids. However, only limited knowledge exists on serology test performance in 2-6 month-old piglets. In this age-class, recent infections might cause lower antibody levels and lower test sensitivity. We examined 126 wild boar piglets from a TB-endemic site using 6 antibody detection tests in order to assess test performance. Bacterial culture (n=53) yielded a M. bovis infection prevalence of 33.9%, while serum antibody prevalence estimated by different tests ranged from 19% to 38%, reaching sensitivities between 15.4% and 46.2% for plate ELISAs and between 61.5% and 69.2% for rapid immunochromatographic tests based on dual path platform (DPP) technology. The Cohen kappa coefficient of agreement between DPP WTB (Wildlife TB) assay and culture results was moderate (0.45) and all other serological tests used had poor to fair agreements. This survey revealed the ability of several tests for detecting serum antibodies against the MTC antigens in 2-6 month-old naturally infected wild boar piglets. The best performance was demonstrated for DPP tests. The results confirmed our initial hypothesis of a lower sensitivity of serology for detecting M. bovis-infected piglets, as compared to older wild boar. Certain tests, notably the rapid animal-side tests, can contribute to TB control strategies by enabling the setup of test and cull schemes or improving pre-movement testing. However, sub-optimal test performance in piglets as compared to that in older wild boar should be taken into account. Copyright © 2015 Elsevier B.V. All rights reserved.

CD137+CD154− Expression As a Regulatory T Cell (Treg)-Specific Activation Signature for Identification and Sorting of Stable Human Tregs from In Vitro Expansion Cultures

PubMed Central

Nowak, Anna; Lock, Dominik; Bacher, Petra; Hohnstein, Thordis; Vogt, Katrin; Gottfreund, Judith; Giehr, Pascal; Polansky, Julia K.; Sawitzki, Birgit; Kaiser, Andrew; Walter, Jörn; Scheffold, Alexander

2018-01-01

Regulatory T cells (Tregs) are an attractive therapeutic tool for several different immune pathologies. Therapeutic Treg application often requires prolonged in vitro culture to generate sufficient Treg numbers or to optimize their functionality, e.g., via genetic engineering of their antigen receptors. However, purity of clinical Treg expansion cultures is highly variable, and currently, it is impossible to identify and separate stable Tregs from contaminating effector T cells, either ex vivo or after prior expansion. This represents a major obstacle for quality assurance of expanded Tregs and raises significant safety concerns. Here, we describe a Treg activation signature that allows identification and sorting of epigenetically imprinted Tregs even after prolonged in vitro culture. We show that short-term reactivation resulted in expression of CD137 but not CD154 on stable FoxP3+ Tregs that displayed a demethylated Treg-specific demethylated region, high suppressive potential, and lack of inflammatory cytokine expression. We also applied this Treg activation signature for rapid testing of chimeric antigen receptor functionality in human Tregs and identified major differences in the signaling requirements regarding CD137 versus CD28 costimulation. Taken together, CD137+CD154− expression emerges as a universal Treg activation signature ex vivo and upon in vitro expansion allowing the identification and isolation of epigenetically stable antigen-activated Tregs and providing a means for their rapid functional testing in vitro. PMID:29467769

CD137+CD154- Expression As a Regulatory T Cell (Treg)-Specific Activation Signature for Identification and Sorting of Stable Human Tregs from In Vitro Expansion Cultures.

PubMed

Nowak, Anna; Lock, Dominik; Bacher, Petra; Hohnstein, Thordis; Vogt, Katrin; Gottfreund, Judith; Giehr, Pascal; Polansky, Julia K; Sawitzki, Birgit; Kaiser, Andrew; Walter, Jörn; Scheffold, Alexander

2018-01-01

Regulatory T cells (Tregs) are an attractive therapeutic tool for several different immune pathologies. Therapeutic Treg application often requires prolonged in vitro culture to generate sufficient Treg numbers or to optimize their functionality, e.g., via genetic engineering of their antigen receptors. However, purity of clinical Treg expansion cultures is highly variable, and currently, it is impossible to identify and separate stable Tregs from contaminating effector T cells, either ex vivo or after prior expansion. This represents a major obstacle for quality assurance of expanded Tregs and raises significant safety concerns. Here, we describe a Treg activation signature that allows identification and sorting of epigenetically imprinted Tregs even after prolonged in vitro culture. We show that short-term reactivation resulted in expression of CD137 but not CD154 on stable FoxP3+ Tregs that displayed a demethylated Treg-specific demethylated region, high suppressive potential, and lack of inflammatory cytokine expression. We also applied this Treg activation signature for rapid testing of chimeric antigen receptor functionality in human Tregs and identified major differences in the signaling requirements regarding CD137 versus CD28 costimulation. Taken together, CD137+CD154- expression emerges as a universal Treg activation signature ex vivo and upon in vitro expansion allowing the identification and isolation of epigenetically stable antigen-activated Tregs and providing a means for their rapid functional testing in vitro .

Early Diagnosis of Scrub Typhus with a Rapid Flow Assay Using Recombinant Major Outer Membrane Protein Antigen (r56) of Orientia tsutsugamushi

PubMed Central

Ching, W.-M.; Rowland, D.; Zhang, Z.; Bourgeois, A. L.; Kelly, D.; Dasch, G. A.; Devine, P. L.

2001-01-01

The variable 56-kDa major outer membrane protein of Orientia tsutsugamushi is the immunodominant antigen in human scrub typhus infections. We developed a rapid immunochromatographic flow assay (RFA) for the detection of immunoglobulin M (IgM) and IgG antibodies to O. tsutsugamushi. The RFA employs a truncated recombinant 56-kDa protein from the Karp strain as the antigen. The performance of the RFA was evaluated with a panel of 321 sera (serial bleedings of 85 individuals suspected of scrub typhus) which were collected in the Pescadore Islands, Taiwan, from 1976 to 1977. Among these 85 individuals, IgM tests were negative for 7 cases by both RFA and indirect fluorescence assay (IFA) using Karp whole-cell antigen. In 29 cases specific responses were detected by the RFA earlier than by IFA, 44 cases had the same detection time, and 5 cases were detected earlier by IFA than by RFA. For IgG responses, 4 individuals were negative with both methods, 37 cases exhibited earlier detection by RFA than IFA, 42 cases were detected at the same time, and 2 cases were detected earlier by IFA than by RFA. The sensitivities of RFA detection of antibody in sera from confirmed cases were 74 and 86% for IgM and IgG, respectively. When IgM and IgG results were combined, the sensitivity was 89%. A panel of 78 individual sera collected from patients with no evidence of scrub typhus was used to evaluate the specificity of the RFA. The specificities of the RFA were 99% for IgM and 97% for IgG. The sensitivities of IFA were 53 and 73% for IgM and IgG, respectively, and were 78% when the results of IgM and IgG were combined. The RFA test was significantly better than the IFA test for the early detection of antibody to scrub typhus in primary infections, while both tests were equally sensitive with reinfected individuals. PMID:11238230

Evaluation of a new rapid kit, BD MGIT TBc identification test for confirmation of Mycobacterium tuberculosis complex.

PubMed

Kandhakumari, Gandhi; Stephen, Selvaraj

2017-01-01

At present, three rapid kits are available globally for the confirmation of Mycobacterium tuberculosis complex (MTBC) in cultures by MPT64 antigen (MPT64 Ag) detection. These include Capilia TB, SD Bioline, and BD MGIT TBc Identification (TBcID). The third kit is yet to be validated in India. We have tested this kit and compared with SD Bioline using conventional tests as gold standard. Seventy-one MTBC (70 M. tuberculosis and one Mycobacterium bovis) and four nontuberculous mycobacteria (NTM) were isolated from 649 clinical specimens in MGIT 960 and/or Lowenstein-Jensen slants (LJ). MPT64 Ag was detected by both TBcID and SD Bioline kits in all the 71 clinical isolates and the reference strain M. tuberculosis H37Rv. All NTM species tested were negative by the two different kits. Thus, TBcID kit showed 100% concordance in terms of sensitivity and specificity. Rapid kits confirm MTBC cultures within 15 min in contrast to several weeks' time required by conventional techniques.

Comparison of Simplexa universal direct PCR with cytotoxicity assay for diagnosis of Clostridium difficile infection: performance, cost, and correlation with disease.

PubMed

Landry, Marie L; Ferguson, David; Topal, Jeffrey

2014-01-01

Simplexa Clostridium difficile universal direct PCR, a real-time PCR assay for the detection of the C. difficile toxin B (tcdB) gene using the 3M integrated cycler, was compared with a two-step algorithm which includes the C. Diff Chek-60 glutamate dehydrogenase (GDH) antigen assay followed by cytotoxin neutralization. Three hundred forty-two liquid or semisolid stools submitted for diagnostic C. difficile testing, 171 GDH antigen positive and 171 GDH antigen negative, were selected for the study. All samples were tested by the C. Diff Chek-60 GDH antigen assay, cytotoxin neutralization, and Simplexa direct PCR. Of 171 GDH-positive samples, 4 were excluded (from patients on therapy or from whom duplicate samples were obtained) and 88 were determined to be true positives for toxigenic C. difficile. Of the 88, 67 (76.1%) were positive by the two-step method and 86 (97.7%) were positive by PCR. Seventy-nine were positive by the GDH antigen assay only. Of 171 GDH antigen-negative samples, none were positive by PCR. One antigen-negative sample positive by the cytotoxin assay only was deemed a false positive based on chart review. Simplexa C. difficile universal direct PCR was significantly more sensitive for detecting toxigenic C. difficile bacteria than cytotoxin neutralization (P = 0.0002). However, most PCR-positive/cytotoxin-negative patients did not have clear C. difficile disease. The estimated cost avoidance provided by a more rapid molecular diagnosis was outweighed by the cost of isolating and treating PCR-positive/cytotoxin-negative patients. The costs, clinical consequences, and impact on nosocomial transmission of treating and/or isolating patients positive for toxigenic C. difficile by PCR but negative for in vivo toxin production merit further study.

Multiantigen Print Immunoassay for Comparison of Diagnostic Antigens for Taenia solium Cysticercosis and Taeniasis▿

PubMed Central

Handali, Sukwan; Klarman, Molly; Gaspard, Amanda N.; Noh, John; Lee, Yeuk-Mui; Rodriguez, Silvia; Gonzalez, Armando E.; Garcia, Hector H.; Gilman, Robert H.; Tsang, Victor C. W.; Wilkins, Patricia P.

2010-01-01

One of the best-characterized tests for the diagnosis of neurocysticercosis is the enzyme-linked immunoelectrotransfer blot assay, developed at the CDC, which uses lentil lectin-purified glycoproteins (LLGPs) extracted from Taenia solium cysticerci. The purification of the LLGP antigens has been difficult to standardize, and the polyacrylamide gel system used for the immunoblot assay is not easily transferable to other laboratories. In this study, we developed a multiantigen printing immunoassay (MAPIA) to compare the performance of multiple recombinant Taenia solium proteins with the potential for the detection of cysticercosis and taeniasis. We prepared MAPIA strips using six cysticercosis and two taeniasis diagnostic proteins and compared the performance of the proteins with sera collected from defined cysticercosis and taeniasis cases. Of the six cysticercosis antigens, rT24H performed well in detecting cases with two or more viable cysts in the brain (sensitivity and specificity, 97% and 99.4%, respectively); the use of a combination of cysticercosis antigens did not improve the sensitivity of the test and decreased the specificity. None of the antigens could differentiate the different clinical presentations of cysticercosis. Both of the taeniasis antigens (rES33 and rES38) had the same sensitivity of 99.4% and specificities of 93.9% and 94.5%, respectively. Some cross-reactivity against rES33 and rES38 was found, especially with sera from cases infected with Schistosoma mansoni. We conclude that MAPIA is a simple and effective tool that may be used to compare antibody responses to different cysticercosis and taeniasis antigens and, in this case, may be useful for the rapid detection of T. solium cases. PMID:19906893

Multiantigen print immunoassay for comparison of diagnostic antigens for Taenia solium cysticercosis and taeniasis.

PubMed

Handali, Sukwan; Klarman, Molly; Gaspard, Amanda N; Noh, John; Lee, Yeuk-Mui; Rodriguez, Silvia; Gonzalez, Armando E; Garcia, Hector H; Gilman, Robert H; Tsang, Victor C W; Wilkins, Patricia P

2010-01-01

One of the best-characterized tests for the diagnosis of neurocysticercosis is the enzyme-linked immunoelectrotransfer blot assay, developed at the CDC, which uses lentil lectin-purified glycoproteins (LLGPs) extracted from Taenia solium cysticerci. The purification of the LLGP antigens has been difficult to standardize, and the polyacrylamide gel system used for the immunoblot assay is not easily transferable to other laboratories. In this study, we developed a multiantigen printing immunoassay (MAPIA) to compare the performance of multiple recombinant Taenia solium proteins with the potential for the detection of cysticercosis and taeniasis. We prepared MAPIA strips using six cysticercosis and two taeniasis diagnostic proteins and compared the performance of the proteins with sera collected from defined cysticercosis and taeniasis cases. Of the six cysticercosis antigens, rT24H performed well in detecting cases with two or more viable cysts in the brain (sensitivity and specificity, 97% and 99.4%, respectively); the use of a combination of cysticercosis antigens did not improve the sensitivity of the test and decreased the specificity. None of the antigens could differentiate the different clinical presentations of cysticercosis. Both of the taeniasis antigens (rES33 and rES38) had the same sensitivity of 99.4% and specificities of 93.9% and 94.5%, respectively. Some cross-reactivity against rES33 and rES38 was found, especially with sera from cases infected with Schistosoma mansoni. We conclude that MAPIA is a simple and effective tool that may be used to compare antibody responses to different cysticercosis and taeniasis antigens and, in this case, may be useful for the rapid detection of T. solium cases.

Preliminary multiplex microarray IgG immunoassay for the diagnosis of toxoplasmosis and rubella.

PubMed

Baschirotto, Priscila T; Krieger, Marco A; Foti, Leonardo

2017-06-01

During pregnancy, toxoplasmosis and rubella can cause serious damage to the mother and the foetus through vertical transmission. Early diagnosis enables implementation of health measures aimed at preventing vertical transmission and minimising damage caused by these diseases. Here, we report the development of a multiplex assay for simultaneous detection of IgG antibodies produced during toxoplasmosis and rubella infection. This assay is based on xMap technology. Initially, by singleplex assays, we evaluated the following antigens: one Toxoplasma gondii lysate; two antigenic extracts of T. gondii (TOX8131 and TOX8122); fragments of T. gondii antigens [SAG-1 (amino acids 45-198), GRA-7 (24-100), GRA-1 (57-149), ROP-4, and MIC-3 (234-306)]; two chimeric antigens composed of fragments of SAG-1, GRA-7, and P35 (CTOX and CTOXH); and fragments of Rubella virus antigens [E-1 (157-176, 213-239, 374-390), E-2 (31-105), and C (1-123)]. A multiplex assay to simultaneously diagnose toxoplasmosis and rubella was designed with the best-performing antigens in singleplex and multiplex assays, which included CTOXH, T. gondii lysate, TOX8131, E-1, and E-2. The multiplex assay showed 100% sensitivity and specificity for anti-T. gondii IgG detection and 95.6% sensitivity and 100% specificity for anti-R. virus IgG detection. We found that, despite the difficulties related to developing multiplex systems, different types of antigens (extracts and recombinant proteins) can be used to develop high-performance diagnostic tests. The assay developed is suitable to screen for prior T. gondii and R. virus infections, because it is a rapid, high-throughput, low-cost alternative to the current standard diagnostic tools, which require multiple individual tests.

Miniature silicon electronic biological assay chip and applications for rapid battlefield diagnostics

NASA Astrophysics Data System (ADS)

Cunningham, Brian T.; Regan, Robert A.; Clapp, Christopher; Hildebrant, Eric; Weinberg, Marc S.; Williams, John

1999-07-01

Assessing the medical condition of battlefield personnel requires the development of rapid, portable biological diagnostic assays for a wide variety of antigens and enzymes. Ideally, such an assay would be inexpensive, small, and require no added reagents while maintaining the sensitivity and accuracy of laboratory-based assays. In this work, a microelectromechanical (MEMS) based biological assay sensor is presented which is expected to meet the above requirements. The sensor is a thin silicon membrane resonator (SMR) which registers a decrease in resonant frequency when mass is adsorbed onto its surface. By coating the sensor surface with a monolayer of antibody, for example, we have detected the corresponding antigen with a detection resolution of 0.25 ng/ml in phosphate buffer solution. Micromachining techniques are being used to integrate many (64 elements on the first test chip) identical SMR sensors into a single silicon chip which would be capable of simultaneously performing a wide variety of biomedical assays. The sensors require only a small printed circuit board and 8V power supply to operate and provide a readout. The presentation will describe the operation of the SMR sensor, the fabrication of the sensor array, and initial test results using commercially available animal immunoglobulins in laboratory-prepared test solutions.

Rapid polyclonal desensitization with antibodies to IgE and FcεRIα.

PubMed

Khodoun, Marat V; Kucuk, Zeynep Yesim; Strait, Richard T; Krishnamurthy, Durga; Janek, Kevin; Lewkowich, Ian; Morris, Suzanne C; Finkelman, Fred D

2013-06-01

Rapid desensitization, a procedure in which persons allergic to an antigen are treated at short intervals with increasing doses of that antigen until they tolerate a large dose, is an effective, but risky, way to induce temporary tolerance. We wanted to determine whether this approach can be adapted to suppress all IgE-mediated allergies in mice by injecting serially increasing doses of monoclonal antibodies (mAbs) to IgE or FcεRIα. Active and passive models of antigen- and anti-IgE mAb-induced IgE-mediated anaphylaxis were used. Mice were desensitized with serially increasing doses of anti-IgE mAb, anti-FcεRIα mAb, or antigen. Development of shock (hypothermia), histamine and mast cell protease release, cytokine secretion, calcium flux, and changes in cell number and FcεRI and IgE expression were evaluated. Rapid desensitization with anti-IgE mAb suppressed IgE-mediated immediate hypersensitivity; however, some mice developed mild anaphylaxis during desensitization. Rapid desensitization with anti-FcεRIα mAb that only binds FcεRI that is not occupied by IgE suppressed both active and passive IgE-mediated anaphylaxis without inducing disease. It quickly, but temporarily, suppressed IgE-mediated anaphylaxis by decreasing mast cell signaling through FcεRI, then slowly induced longer lasting mast cell unresponsiveness by removing membrane FcεRI. Rapid desensitization with anti-FcεRIα mAb was safer and longer lasting than rapid desensitization with antigen. A rapid desensitization approach with anti-FcεRIα mAb safely desensitizes mice to IgE-mediated anaphylaxis by inducing mast cell anergy and later removing all mast cell IgE. Rapid desensitization with an anti-human FcεRIα mAb may be able to prevent human IgE-mediated anaphylaxis. Copyright © 2013 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.

Rapid polyclonal desensitization with antibodies to IgE and FcεRIα

PubMed Central

Khodoun, Marat V.; Kucuk, Zeynep Yesim; Strait, Richard T.; Krishnamurthy, Durga; Janek, Kevin; Lewkowich, Ian; Morris, Suzanne C.; Finkelman, Fred D.

2013-01-01

Background Rapid desensitization,a procedure in which individuals allergic to an antigen are treated at short intervals with increasing doses of that antigen until they tolerate a large dose, is an effective, but risky way to induce temporary tolerance. Objective To determine whether this approach can be adapted to suppress all IgE-mediated in mice by injecting serially increasing doses of monoclonal antibodies (mAbs) to IgE or FcεRIα. Methods Active and passive models of antigen- and anti-IgE mAb-induced IgE-mediated anaphylaxis were used. Mice were desensitized with serially increasing doses of anti-IgE mAb, anti-FcεRIα mAb or antigen. Development of shock (hypothermia), histamine and mast cell protease release, cytokine secretion, calcium flux and changes in cell number and FcεRI and IgE expression were evaluated. Results Rapid desensitization with anti-IgE mAb suppressed IgE-mediated immediate hypersensitivity; however, some mice developed mild anaphylaxis during desensitization. Rapid desensitization with anti-FcεRIα mAb that only binds FcεRI that is not occupied by IgE suppressed both active and passive IgE-mediated anaphylaxis without inducing disease. It quickly, but temporarily, suppressed IgE-mediated anaphylaxis by decreasing mast cell signaling through FcεRI, then slowly slowlyinduced longer lasting mast cell unresponsiveness by removing membrane FcεRI. Rapid desensitization with anti-FcεRIα mAb was safer and longer-lasting than rapid desensitization with antigen. Conclusion A rapid desensitization approach with anti-FcεRIα mAb safely desensitizes mice to IgE-mediated anaphylaxis by inducing mast cell anergy and later, removing all mast cell IgE. Rapid desensitization with an anti-human FcεRIα mAb may be able to prevent human IgE-mediated anaphylaxis. PMID:23632296

Evaluation of two rapid antigen assays, BioStar Strep A OIA and Pacific Biotech CARDS O.S., and culture for detection of group A streptococci in throat swabs.

PubMed Central

Dale, J C; Vetter, E A; Contezac, J M; Iverson, L K; Wollan, P C; Cockerill, F R

1994-01-01

Two rapid methods, BioStar Strep A OIA (OIA; BioStar, Inc., Boulder, Colo.), an optical immunoassay, and CARDS O.S. (O.S.; Pacific Biotech, Inc., San Diego, Calif.), a color immunochromographic assay, and two culture methods, one with 5% sheep blood agar (SBA) and one with Todd-Hewitt broth (TH; Remel, Lenexa, Kans.), were evaluated for use in the detection of Streptococcus pyogenes from pharnygeal swabs. Seven hundred forty-six double swabs (Culturette II) were processed, with OIA and SBA culture performed on one swab and O.S. and SBA culture performed on the other swab. The pledget from the Culturette II was incubated overnight in TH and was subcultured onto SBA for an additional 48 h in ambient air. All beta-hemolytic streptococci from culture were tested by a direct fluorescent-antibody test (Difco Laboratories, Detroit, Mich.). Specimens with discordant fluorescent-antibody test and rapid test results were also tested by using the Streptex latex agglutination reagent (Murex Diagnostics Limited, Dartford, England). The results obtained by all testing methods were compared with a combined test result ("gold standard"), which was defined as any positive culture detected by the SBA or TH culture methods and confirmed by Streptex latex agglutination or, in the case of negative results by both culture methods, a concomitant positive result by OIA and O.S. antigen testing. Sensitivity and specificity results for each of the methods were as follows, respectively: OIA, 81.0 and 97.5%; O.S., 74.4 and 99.0%; SBA culture, 92.3 and 98.3%; and TH culture, 86.4 and 100%. Both OIA and O.S. are suitable screening methods for detecting S. pyogenes directly from throat swabs but are of insufficient sensitivity to eliminate the need for backup cultures for specimens with negative OIA and O.S. results. PMID:7852559

Rapid Detection of the Varicella Zoster Virus

NASA Technical Reports Server (NTRS)

Lewis, Michelle P.; Harding, Robert

2011-01-01

1.Technology Description-Researchers discovered that when the Varicella Zoster Virus (VZV) reactivates from latency in the body, the virus is consistently present in saliva before the appearance of skin lesions. A small saliva sample is mixed with a specialized reagent in a test kit. If the virus is present in the saliva sample, the mixture turns a red color. The sensitivity and specificity emanates from an antibody-antigen reaction. This technology is a rapid, non-invasive, point of-of-care testing kit for detecting the virus from a saliva sample. The device is easy to use and can be used in clinics and in remote locations to quickly detect VZV and begin treatment with antiviral drugs. 2.Market Opportunity- RST Bioscience will be the first and only company to market a rapid, same day test kit for the detection of VZV in saliva. The RST detection test kit will have several advantages over existing, competitive technology. The test kit is self contained and laboratory equipment is not required for analysis of the sample. Only a single saliva sample is required to be taken instead of blood or cerebral spinal fluid. The test kit is portable, sterile and disposable after use. RST detection test kits require no electrical power or expensive storage equipment and can be used in remote locations. 3.Market Analysis- According to the CDC, it is estimated that 1 million cases of shingles occur each year in the U.S. with more than half over the age of sixty. There is a high demand for rapid diagnostics by the public. The point-of-care testing (POCT) market is growing faster than other segments of in vitro diagnostics. According to a July 2007 InteLab Corporation industry report the overall market for POCT was forecast to increase from $10.3 billion in 2005 to $18.7 billion by 2011. The market value of this test kit has not been determined. 4.Competition- The VZV vaccine prevents 50% of cases and reduces neuralgia by 66%. The most popular test detects VZV-specific IgM antibody in blood. Other tests include running a sample in a polymerase chain reaction analyzer, enzyme immunoassay, latex agglutination, indirect fluorescent antibody and fluorescent antibody to membrane antigen assay. These existing tests require laboratory analysis by trained personnel, expensive equipment, invasive procedures and a longer period of time to obtain test results.

Performance evaluation of the FDA-approved Determine™ HIV-1/2 Ag/Ab Combo assay using plasma and whole blood specimens.

PubMed

Masciotra, Silvina; Luo, Wei; Westheimer, Emily; Cohen, Stephanie E; Gay, Cynthia L; Hall, Laura; Pan, Yi; Peters, Philip J; Owen, S Michele

2017-06-01

The Determine™ HIV-1/2 Ag/Ab Combo (DC) rapid test can identify HIV-1 infection earlier than rapid antibody-only tests in plasma specimens. We compared the performance of DC with a laboratory-based antigen/antibody (Ag/Ab) combo assay in plasma and evaluated antigen reactivity in whole blood specimens. We tested by DC 508 plasma specimens collected in a prospective study and 107 sequential plasma and simulated whole blood specimens from 20 seroconversion panels. Previous results using the ARCHITECT (ARC) Ag/Ab combo assay were compared to DC results. In seroconversion panels, the days from the first HIV1 RNA-positive test to first DC-reactive in plasma and whole blood was compared. McNemar's and Wilcoxon signed rank tests were used for statistical analysis. Of 415 HIV-positive samples, ARC detected 396 (95.4%) and DC 337 (81.2%) (p<0.0001). DC was reactive in 50.0% of ARC-reactive/MS-negative, 78.6% of ARC-reactive/MS-indeterminate, and 99.6% of ARC-reactive/MS-HIV-1-positive or -undifferentiated specimens. DC antigen reactivity was higher among ARC-reactive/MS-negative than MS-indeterminate samples. In 20 HIV-1 seroconversion panels, there was a significant difference between DC reactivity in plasma (91.1%) and whole blood (56.4%) (p<0.0001). DC with whole blood showed a significant delay in reactivity compared to plasma (p=0.008). In plasma, DC was significantly less sensitive than an instrumented laboratory-based Ag/Ab combo assay. DC in plasma was significantly more sensitive compared to whole blood in early HIV-1 infections. With the U.S. laboratory-based diagnostic algorithm, DC as the first step would likely miss a high proportion of HIV-1 infections in early stages of seroconversion. Published by Elsevier B.V.

Rapid classification of biological components

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thompson, Vicki S.; Barrett, Karen B.; Key, Diane E.

A method is disclosed for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an illustrative embodiment of the invention, the analyte is a drug, such as marijuana, Cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method involves attaching antigens of the surface of a solid support in a preselected pattern to form an array wherein the locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to antigens in the array,more » thereby forming immune complexes; washing away antibodies that do not form immune complexes; and detecting the immune complexes, thereby forming an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to a subject's identity.« less

Rapid classification of biological components

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thompson, Vicki S.; Barrett, Karen B.; Key, Diane E.

A method is disclosed for analyzing a biological sample by antibody profiling for identifying forensic samples or for detecting the presence of an analyte. In an illustrative embodiment of the invention, the analyte is a drug, such as marijuana, cocaine (crystalline tropane alkaloid), methamphetamine, methyltestosterone, or mesterolone. The method involves attaching antigens to a surface of a solid support in a preselected pattern to form an array wherein the locations of the antigens are known; contacting the array with the biological sample such that a portion of antibodies in the sample reacts with and binds to antigens in the array,more » thereby forming immune complexes; washing away antibodies that do not form immune complexes; and detecting the immune complexes, thereby forming an antibody profile. Forensic samples are identified by comparing a sample from an unknown source with a sample from a known source. Further, an assay, such as a test for illegal drug use, can be coupled to a test for identity such that the results of the assay can be positively correlated to a subject's identity.« less

Indirect enzyme-linked immunosorbent assay method based on Streptococcus agalactiae rSip-Pgk-FbsA fusion protein for detection of bovine mastitis.

PubMed

Bu, Ri-E; Wang, Jin-Liang; Wu, Jin-Hua; Xilin, Gao-Wa; Chen, Jin-Long; Wang, Hua

2017-03-01

The aim of this study was to establish a rapid and accurate method for the detection of the Streptococcus agalactiae antibody (SA-Ab) to determine the presence of the bovine mastitis (BM)-causative pathogen. The multi-subunit fusion protein rSip-Pgk-FbsA was prokaryotically expressed and purified. The triple activities of the membrane surface-associated proteins Sip, phosphoglycerate kinase (Pgk), and fibronectin (FbsA) were used as the diagnostic antigens to establish an indirect enzyme-linked immunosorbent assay (ELISA) method for the detection of SA-Ab in BM. The optimal antigen coating concentration was 2 μg/mL, the optimal serum dilution was 1:160, and the optimal dilution of the enzyme-labeled secondary antibody was 1:6000. The sensitivity, specificity, and repeatability tests showed that the method established in this study had no cross-reaction with antibodies to Streptococcus pyogenes, Escherichia coli, Staphylococcus aureus, and Staphylococcus epidermidis in the sera. The results of the sensitivity test showed that a positive result could be obtained even if the serum dilution reached 1:12,800, indicating the high sensitivity and good repeatability of the method. The positive coincidence rate of this method was 98.6%, which is higher than that of previous tests established with the Sip or Pgk mono-antigen fusion protein, respectively, demonstrating the relatively higher sensitivity of this newly established method. The detection rate for 389 clinical samples was 46.53%. The indirect ELISA method established in this study could provide a more accurate and reliable serological method for the rapid detection of S. agalactiae in cases of BM.

The accuracy of the first response histidine-rich protein2 rapid diagnostic test compared with malaria microscopy for guiding field treatment in an outbreak of falciparum malaria.

PubMed

Ghouth, Abdulla Salim Bin; Nasseb, Faraj Mubarak; Al-Kaldy, Khaled Hussin

2012-01-01

Recent WHO guidelines recommended a universal "test and treat" strategy for malaria mainly by use of the rapid diagnostic test (RDT) in all areas. There are concerns about RDT that use the antigen histidine-rich protein2 (HRP2) to detect Plasmodium falciparum, because infection can persist after effective treatment. The aim of this paper is to describe the accuracy of the first response (HRP2)-RDT compared with malaria microscopy used for guiding the field treatment of patients in an outbreak situation in the Al-Rahabah area in Al-Rydah district in Hadramout/Yemen. An ad hoc cross sectional survey of all febrile patients in the affected area was conducted in May 2011. The field team was developed including the case management group and the entomology group. The group of case management prepared their plan based on "test and treat" strategy by using First Response Malaria Antigen HRP2 rapid diagnostic test for falciparum malaria, artemsinin-based combination therapy (ACT) according to the national policy of anti-malaria drugs in Yemen were supplied to treat those who were found to be RDT positive in the field; also blood smear films were taken from every patient with fever in order to validate the use of the RDT in the field. Blood film slides prepared and read by skilled lab technicians, the fourth reading was done by one lab expert in the malaria referral lab. The accuracy parameters of HRP2 compared with microscopy are: Sensitivity (74%), specificity (94%). The positive predictive value is 68% and the negative predictive value is 96%. Total agreement is 148/162 (93%) and the overall prevalence is 14%. All the positive malaria cases were of P. falciparum either coming from RDT or microscopy. HRP2-rapid test is an acceptable test as a guide for field treatment in an outbreak situation where prompt response is indicated. Good prepared blood film slides should be used as it is feasible to evaluate the accuracy of RDTs as a quality control tool.

A novel assay for detecting canine parvovirus using a quartz crystal microbalance biosensor.

PubMed

Kim, Yong Kwan; Lim, Seong-In; Choi, Sarah; Cho, In-Soo; Park, Eun-Hye; An, Dong-Jun

2015-07-01

Rapid and accurate diagnosis is crucial to reduce both the shedding and clinical signs of canine parvovirus (CPV). The quartz crystal microbalance (QCM) is a new tool for measuring frequency changes associated with antigen-antibody interactions. In this study, the QCM biosensor and ProLinker™ B were used to rapidly diagnosis CPV infection. ProLinker™ B enables antibodies to be attached to a gold-coated quartz surface in a regular pattern and in the correct orientation for antigen binding. Receiver operating characteristics (ROC) curves were used to set a cut-off value using reference CPVs (two groups: one CPV-positive and one CPV-negative). The ROC curves overlapped and the point of intersection was used as the cut-off value. A QCM biosensor with a cut-off value of -205 Hz showed 95.4% (104/109) sensitivity and 98.0% (149/152) specificity when used to test 261 field fecal samples compared to PCR. In conclusion, the QCM biosensor described herein is eminently suitable for the rapid diagnosis of CPV infection with high sensitivity and specificity. Therefore, it is a promising analytical tool that will be useful for clinical diagnosis, which requires rapid and reliable analyses. Copyright © 2015 Elsevier B.V. All rights reserved.

Expression of the VP40 antigen from the Zaire ebolavirus in tobacco plants.

PubMed

Monreal-Escalante, Elizabeth; Ramos-Vega, Abel A; Salazar-González, Jorge A; Bañuelos-Hernández, Bernardo; Angulo, Carlos; Rosales-Mendoza, Sergio

2017-07-01

The plant cell is able to produce the VP40 antigen from the Zaire ebolavirus , retaining the antigenicity and the ability to induce immune responses in BALB/c mice. The recent Ebola outbreak evidenced the need for having vaccines approved for human use. Herein we report the expression of the VP40 antigen from the Ebola virus as an initial effort in the development of a plant-made vaccine that could offer the advantages of being cheap and scalable, which is proposed to overcome the rapid need for having vaccines to deal with future outbreaks. Tobacco plants were transformed by stable DNA integration into the nuclear genome using the CaMV35S promoter and a signal peptide to access the endoplasmic reticulum, reaching accumulation levels up to 2.6 µg g -1 FW leaf tissues. The antigenicity of the plant-made VP40 antigen was evidenced by Western blot and an initial immunogenicity assessment in test animals that revealed the induction of immune responses in BALB/c mice following three weekly oral or subcutaneous immunizations at very low doses (125 and 25 ng, respectively) without accessory adjuvants. Therefore, this plant-based vaccination prototype is proposed as an attractive platform for the production of vaccines in the fight against Ebola virus disease outbreaks.

Enhanced Direct Major Histocompatibility Complex Class I Self-Antigen Presentation Induced by Chlamydia Infection

PubMed Central

Cram, Erik D.; Simmons, Ryan S.; Palmer, Amy L.; Hildebrand, William H.; Rockey, Daniel D.

2015-01-01

The direct major histocompatibility complex (MHC) class I antigen presentation pathway ensures intracellular peptides are displayed at the cellular surface for recognition of infected or transformed cells by CD8+ cytotoxic T lymphocytes. Chlamydia spp. are obligate intracellular bacteria and, as such, should be targeted by CD8+ T cells. It is likely that Chlamydia spp. have evolved mechanisms to avoid the CD8+ killer T cell responses by interfering with MHC class I antigen presentation. Using a model system of self-peptide presentation which allows for posttranslational control of the model protein's stability, we tested the ability of various Chlamydia species to alter direct MHC class I antigen presentation. Infection of the JY lymphoblastoid cell line limited the accumulation of a model host protein and increased presentation of the model-protein-derived peptides. Enhanced self-peptide presentation was detected only when presentation was restricted to defective ribosomal products, or DRiPs, and total MHC class I levels remained unaltered. Skewed antigen presentation was dependent on a bacterial synthesized component, as evidenced by reversal of the observed phenotype upon preventing bacterial transcription, translation, and the inhibition of bacterial lipooligosaccharide synthesis. These data suggest that Chlamydia spp. have evolved to alter the host antigen presentation machinery to favor presentation of defective and rapidly degraded forms of self-antigen, possibly as a mechanism to diminish the presentation of peptides derived from bacterial proteins. PMID:26597986

Changing selective pressure during antigenic changes in human influenza H3.

PubMed

Blackburne, Benjamin P; Hay, Alan J; Goldstein, Richard A

2008-05-02

The rapid evolution of influenza viruses presents difficulties in maintaining the optimal efficiency of vaccines. Amino acid substitutions result in antigenic drift, a process whereby antisera raised in response to one virus have reduced effectiveness against future viruses. Interestingly, while amino acid substitutions occur at a relatively constant rate, the antigenic properties of H3 move in a discontinuous, step-wise manner. It is not clear why this punctuated evolution occurs, whether this represents simply the fact that some substitutions affect these properties more than others, or if this is indicative of a changing relationship between the virus and the host. In addition, the role of changing glycosylation of the haemagglutinin in these shifts in antigenic properties is unknown. We analysed the antigenic drift of HA1 from human influenza H3 using a model of sequence change that allows for variation in selective pressure at different locations in the sequence, as well as at different parts of the phylogenetic tree. We detect significant changes in selective pressure that occur preferentially during major changes in antigenic properties. Despite the large increase in glycosylation during the past 40 years, changes in glycosylation did not correlate either with changes in antigenic properties or with significantly more rapid changes in selective pressure. The locations that undergo changes in selective pressure are largely in places undergoing adaptive evolution, in antigenic locations, and in locations or near locations undergoing substitutions that characterise the change in antigenicity of the virus. Our results suggest that the relationship of the virus to the host changes with time, with the shifts in antigenic properties representing changes in this relationship. This suggests that the virus and host immune system are evolving different methods to counter each other. While we are able to characterise the rapid increase in glycosylation of the haemagglutinin during time in human influenza H3, an increase not present in influenza in birds, this increase seems unrelated to the observed changes in antigenic properties.

A TeGM6-4r antigen-based immunochromatographic test (ICT) for animal trypanosomosis.

PubMed

Nguyen, Thu-Thuy; Ruttayaporn, Ngasaman; Goto, Yasuyuki; Kawazu, Shin-ichiro; Sakurai, Tatsuya; Inoue, Noboru

2015-11-01

Animal trypanosomosis is a disease that is distributed worldwide which results in huge economic losses due to reduced animal productivity. Endemic regions are often located in the countryside where laboratory diagnosis is costly or inaccessible. The establishment of simple, effective, and accurate field tests is therefore of great interest to the farming and veterinary sectors. Our study aimed to develop a simple, rapid, and sensitive immunochromatographic test (ICT) for animal trypanosomosis utilizing the recombinant tandem repeat antigen TeGM6-4r, which is conserved amongst salivarian trypanosome species. In the specificity analysis, TeGM6-4r/ICT detected all of Trypanosoma evansi-positive controls from experimentally infected water buffaloes. As expected, uninfected controls tested negative. All sera samples collected from Tanzanian and Ugandan cattle that were Trypanosoma congolense- and/or Trypanosoma vivax-positive by microscopic examination of the buffy coat were found to be positive by the newly developed TeGM6-4r/ICT, which was comparable to results from TeGM6-4r/ELISA (kappa coefficient [κ] = 0.78). TeGM6/ICT also showed substantial agreement with ELISA using Trypanosoma brucei brucei (κ = 0.64) and T. congolense (κ = 0.72) crude antigen, suggesting the high potential of TeGM6-4r/ICT as a field diagnostic test, both for research purposes and on-site diagnosis of animal trypanosomosis.

Development of Lateral Flow Immunoassay for Antigen Detection in Human Angiostrongylus cantonensis Infection

PubMed Central

Chen, Mu-Xin; Chen, Jia-Xu; Chen, Shao-Hong; Huang, Da-Na; Ai, Lin; Zhang, Ren-Li

2016-01-01

Angiostrongyliasis is difficult to be diagnosed for the reason that no ideal method can be used. Serologic tests require specific equipment and are not always available in poverty-stricken zone and are time-consuming. A lateral flow immunoassay (LFIA) may be useful for angiostrongyliasis control. We established a LFIA for the diagnosis of angiostrongyliasis based on 2 monoclonal antibodies (mAbs) against antigens of Angiostrongylus cantonensis adults. The sensitivity and specificity were 91.1% and 100% in LFIA, while those of commercial ELISA kit was 97.8% and 86.3%, respectively. Youden index was 0.91 in LFIA and 0.84 in commercial ELISA kit. LFIA showed detection limit of 1 ng/ml of A. cantonensis ES antigens. This LFIA was simple, rapid, highly sensitive and specific, which opened an alternative approach for the diagnosis of human angiostrongyliasis. PMID:27417097

Detection of Antigenic Variants of Subtype H3 Swine Influenza A Viruses from Clinical Samples

PubMed Central

Martin, Brigitte E.; Li, Lei; Nolting, Jacqueline M.; Smith, David R.; Hanson, Larry A.

2017-01-01

ABSTRACT A large population of genetically and antigenically diverse influenza A viruses (IAVs) are circulating among the swine population, playing an important role in influenza ecology. Swine IAVs not only cause outbreaks among swine but also can be transmitted to humans, causing sporadic infections and even pandemic outbreaks. Antigenic characterizations of swine IAVs are key to understanding the natural history of these viruses in swine and to selecting strains for effective vaccines. However, influenza outbreaks generally spread rapidly among swine, and the conventional methods for antigenic characterization require virus propagation, a time-consuming process that can significantly reduce the effectiveness of vaccination programs. We developed and validated a rapid, sensitive, and robust method, the polyclonal serum-based proximity ligation assay (polyPLA), to identify antigenic variants of subtype H3N2 swine IAVs. This method utilizes oligonucleotide-conjugated polyclonal antibodies and quantifies antibody-antigen binding affinities by quantitative reverse transcription-PCR (RT-PCR). Results showed the assay can rapidly detect H3N2 IAVs directly from nasal wash or nasal swab samples collected from laboratory-challenged animals or during influenza surveillance at county fairs. In addition, polyPLA can accurately separate the viruses at two contemporary swine IAV antigenic clusters (H3N2 swine IAV-α and H3N2 swine IAV-ß) with a sensitivity of 84.9% and a specificity of 100.0%. The polyPLA can be routinely used in surveillance programs to detect antigenic variants of influenza viruses and to select vaccine strains for use in controlling and preventing disease in swine. PMID:28077698

Detection of genome, antigen, and antibodies in oral fluids from pigs infected with foot-and-mouth disease virus.

PubMed

Senthilkumaran, Chandrika; Yang, Ming; Bittner, Hilary; Ambagala, Aruna; Lung, Oliver; Zimmerman, Jeffrey; Giménez-Lirola, Luis G; Nfon, Charles

2017-04-01

Virus nucleic acids and antibody response to pathogens can be measured using swine oral fluids (OFs). Detection of foot-and-mouth disease virus (FMDV) genome in swine OFs has previously been demonstrated. Virus isolation and viral antigen detection are additional confirmatory assays for diagnosing FMDV, but these methods have not been evaluated using swine OF. The objectives of this study were to further validate the molecular detection of FMDV in oral fluids, evaluate antigen detection and FMDV isolation from swine OFs, and develop an assay for isotypic anti-FMDV antibody detection in OFs. Ribonucleic acid (RNA) from FMDV was detected in OFs from experimentally infected pigs by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) from 1 day post-infection (dpi) to 21 dpi. Foot-and-mouth disease virus (FMDV) was isolated from OFs at 1 to 5 dpi. Additionally, FMDV antigens were detected in OFs from 1 to 6 dpi using a lateral flow immunochromatographic strip test (LFIST), which is a rapid pen-side test, and from 2 to 3 dpi using a double-antibody sandwich enzyme-linked immunosorbent assay (DAS ELISA). Furthermore, FMDV-specific immunoglobulin A (IgA) was detected in OFs using an isotype-specific indirect ELISA starting at dpi 14. These results further demonstrated the potential use of oral fluids for detecting FMDV genome, live virus, and viral antigens, as well as for quantifying mucosal IgA antibody response.

A rapid field test for sylvatic plague exposure in wild animals.

PubMed

Abbott, Rachel C; Hudak, Robert; Mondesire, Roy; Baeten, Laurie A; Russell, Robin E; Rocke, Tonie E

2014-04-01

Plague surveillance is routinely conducted to predict future epizootics in wildlife and exposure risk for humans. The most common surveillance method for sylvatic plague is detection of antibodies to Yersinia pestis F1 capsular antigen in sentinel animals, such as coyotes (Canis latrans). Current serologic tests for Y. pestis, hemagglutination (HA) test and enzyme-linked immunosorbent assay (ELISA), are expensive and labor intensive. To address this need, we developed a complete lateral flow device for the detection of specific antibodies to Y. pestis F1 and V antigens. Our test detected anti-F1 and anti-V antibodies in serum and Nobuto filter paper samples from coyotes, and in serum samples from prairie dogs (Cynomys ludovicianus), lynx (Lynx canadensis), and black-footed ferrets (Mustela nigripes). Comparison of cassette results for anti-F1 and anti-V antibodies with results of ELISA or HA tests showed correlations ranging from 0.68 to 0.98. This device provides an affordable, user-friendly tool that may be useful in plague surveillance programs and as a research tool.

A rapid field test for sylvatic plague exposure in wild animals

USGS Publications Warehouse

Abbott, Rachel C.; Hudak, Robert; Mondesire, Roy; Baeten, Laurie A.; Russell, Robin E.; Rocke, Tonie E.

2014-01-01

Plague surveillance is routinely conducted to predict future epizootics in wildlife and exposure risk for humans. The most common surveillance method for sylvatic plague is detection of antibodies to Yersinia pestis F1 capsular antigen in sentinel animals, such as coyotes (Canis latrans). Current serologic tests for Y. pestis, hemagglutination (HA) test and enzyme-linked immunosorbent assay (ELISA), are expensive and labor intensive. To address this need, we developed a complete lateral flow device for the detection of specific antibodies to Y. pestis F1 and V antigens. Our test detected anti-F1 and anti-V antibodies in serum and Nobuto filter paper samples from coyotes, and in serum samples from prairie dogs (Cynomys ludovicianus), lynx (Lynx canadensis), and black-footed ferrets (Mustela nigripes). Comparison of cassette results for anti-F1 and anti-V antibodies with results of ELISA or HA tests showed correlations ranging from 0.68 to 0.98. This device provides an affordable, user-friendly tool that may be useful in plague surveillance programs and as a research tool.

Methods of rapid diagnosis for the etiology of meningitis in adults

PubMed Central

Bahr, Nathan C; Boulware, David R

2014-01-01

Infectious meningitis may be due to bacterial, mycobacterial, fungal or viral agents. Diagnosis of meningitis must take into account numerous items of patient history and symptomatology along with regional epidemiology and basic cerebrospinal fluid testing (protein, etc.) to allow the clinician to stratify the likelihood of etiology possibilities and rationally select additional diagnostic tests. Culture is the mainstay for diagnosis in many cases, but technology is evolving to provide more rapid, reliable diagnosis. The cryptococcal antigen lateral flow assay (Immuno-Mycologics) has revolutionized diagnosis of cryptococcosis and automated nucleic acid amplification assays hold promise for improving diagnosis of bacterial and mycobacterial meningitis. This review will focus on a holistic approach to diagnosis of meningitis as well as recent technological advances. PMID:25402579

Duration of detection of anti-BmR1 IgG4 antibodies after mass-drug administration (MDA) in Sarawak, Malaysia.

PubMed

Noordin, R; Muhi, J; Md Idris, Z; Arifin, N; Kiyu, A

2012-03-01

The detection rates of brugian filariasis in three regions of Sarawak namely Central, North and South after three courses of mass drug administration (MDA) from year 2004 to 2006 was investigated. A recombinant BmR1 antigen-based IgG4 detection test, named Brugia Rapid and night blood smear for microfilaria (mf) detection were used. All three regions recorded a sharp fall in mf positive rates after a year post-MDA. Meanwhile Brugia Rapid positive rates declined more gradually to 3.8% and 5.6% of the pre-MDA levels in the Central and North regions, respectively. This study showed that in filariasis endemic areas in Sarawak, anti-filarial IgG4 antibodies to BmR1, as detected by the Brugia Rapid test, were positive for one to two years after mf disappearance.

Two pseudo-outbreaks of infectious mononucleosis.

PubMed

Armstrong, C W; Hackler, R L; Miller, G B

1986-01-01

Two outbreaks of suspected infectious mononucleosis (IM) were investigated. In the first outbreak IM was diagnosed in nine children attending a day care center. They had been tested in physicians' offices for heterophile antibody using rapid differential slide tests; all tests had been reported positive. On retesting, none of the suspected cases had detectable serum heterophile antibody. The initial test results had been falsely positive as a result of poor laboratory technique. In the second outbreak IM had been diagnosed in 285 college students. Suspected cases had been found to have serum IgG antibody to the viral capsid antigen of Epstein-Barr virus, but most had not been tested for the presence of heterophile antibody. Retesting of 64 students within 1 month of initial testing yielded only one with heterophile antibody. With the exception of young children (less than 4 years of age), differential slide tests for heterophile antibody are sensitive and specific for recent Epstein-Barr virus infection if properly performed. Viral capsid antigen to Epstein-Barr virus (IgG) titers are of limited usefulness in diagnosing acute IM. The misdiagnosis of IM can be prevented by the appropriate selection, performance and interpretation of diagnostic laboratory tests.

In vitro evaluation of five rapid antigen detection tests for group A beta-haemolytic streptococcal sore throat infections.

PubMed

Lasseter, Gemma M; McNulty, Cliodna A M; Richard Hobbs, F D; Mant, David; Little, Paul

2009-12-01

Using accurate and easy to use rapid antigen detection tests (RADTs) to identify group A beta-haemolytic Streptococci (GABHS) sore throat infections could reduce unnecessary antibiotic prescribing and antimicrobial resistance. Although there is no international consensus on the use of RADTs, these kits have been widely adopted in Finland, France and the USA. Yet in the UK, the Clinical Knowledge Summaries, that provide the main online guidance for GPs, discourage RADTs use, citing their poor sensitivity and inability to impact on prescribing decisions in acute sore throat infections. The purpose of this study was to evaluate the ease of use and in vitro accuracy (sensitivity and specificity) of the five most commonly used RADTs in Europe (OSOM Ultra, Quickvue Dipstick, Streptatest, Clearview Exact Strep A and IMI Test Pack). To ensure the RADTs were evaluated objectively, a standardized in vitro method using known concentrations of GABHS was used to remove the inherent biases associated with clinical studies. The IMI Test Pack was the easiest RADT to use overall. The ability to detect all positive GABHS (sensitivity) varied considerably between kits from 95% [95% confidence interval (CI): 88-98%], for the IMI Test Pack and OSOM, to 62% (95% CI: 51-72%) for Clearview, at the highest GABHS concentration. None of the RADTs gave any false-positive results with commensal flora-they were 100% specific. The IMI Test Pack is most suitable for use in primary care, as it had high sensitivity, high specificity and was easy to use.

Cost-effectiveness of a modified two-step algorithm using a combined glutamate dehydrogenase/toxin enzyme immunoassay and real-time PCR for the diagnosis of Clostridium difficile infection.

PubMed

Vasoo, Shawn; Stevens, Jane; Portillo, Lena; Barza, Ruby; Schejbal, Debra; Wu, May May; Chancey, Christina; Singh, Kamaljit

2014-02-01

The analytical performance and cost-effectiveness of the Wampole Toxin A/B EIA, the C. Diff. Quik Chek Complete (CdQCC) (a combined glutamate dehydrogenase antigen/toxin enzyme immunoassay), two RT-PCR assays (Progastro Cd and BD GeneOhm) and a modified two-step algorithm using the CdQCC reflexed to RT-PCR for indeterminate results were compared. The sensitivity of the Wampole Toxin A/B EIA, CdQCC (GDH antigen), BD GeneOhm and Progastro Cd RT-PCR were 85.4%, 95.8%, 100% and 93.8%, respectively. The algorithm provided rapid results for 86% of specimens and the remaining indeterminate results were resolved by RT-PCR, offering the best balance of sensitivity and cost savings per test (algorithm ∼US$13.50/test versus upfront RT-PCR ∼US$26.00/test). Copyright © 2012. Published by Elsevier B.V.

Vaginal Prostate Specific Antigen (PSA) Is a Useful Biomarker of Semen Exposure Among HIV-Infected Ugandan Women.

PubMed

Woolf-King, Sarah E; Muyindike, Winnie; Hobbs, Marcia M; Kusasira, Adrine; Fatch, Robin; Emenyonu, Nneka; Johnson, Mallory O; Hahn, Judith A

2017-07-01

The practical feasibility of using prostate specific antigen (PSA) as a biomarker of semen exposure was examined among HIV-infected Ugandan women. Vaginal fluids were obtained with self-collected swabs and a qualitative rapid test (ABAcard ® p30) was used to detect PSA. Trained laboratory technicians processed samples on-site and positive PSA tests were compared to self-reported unprotected vaginal sex (UVS) in the last 48 h. A total of 77 women submitted 126 samples for PSA testing at up to three study visits. Of these samples, 31 % (n = 39/126) were PSA positive, and 64 % (n = 25/39) of the positive PSA samples were accompanied by self-report of no UVS at the study visit the PSA was collected. There were no reported difficulties with specimen collection, storage, or processing. These findings provide preliminary data on high levels of misreported UVS among HIV-infected Ugandan women using practically feasible methods for PSA collection and processing.

Detection of Antigenic Variants of Subtype H3 Swine Influenza A Viruses from Clinical Samples.

PubMed

Martin, Brigitte E; Bowman, Andrew S; Li, Lei; Nolting, Jacqueline M; Smith, David R; Hanson, Larry A; Wan, Xiu-Feng

2017-04-01

A large population of genetically and antigenically diverse influenza A viruses (IAVs) are circulating among the swine population, playing an important role in influenza ecology. Swine IAVs not only cause outbreaks among swine but also can be transmitted to humans, causing sporadic infections and even pandemic outbreaks. Antigenic characterizations of swine IAVs are key to understanding the natural history of these viruses in swine and to selecting strains for effective vaccines. However, influenza outbreaks generally spread rapidly among swine, and the conventional methods for antigenic characterization require virus propagation, a time-consuming process that can significantly reduce the effectiveness of vaccination programs. We developed and validated a rapid, sensitive, and robust method, the polyclonal serum-based proximity ligation assay (polyPLA), to identify antigenic variants of subtype H3N2 swine IAVs. This method utilizes oligonucleotide-conjugated polyclonal antibodies and quantifies antibody-antigen binding affinities by quantitative reverse transcription-PCR (RT-PCR). Results showed the assay can rapidly detect H3N2 IAVs directly from nasal wash or nasal swab samples collected from laboratory-challenged animals or during influenza surveillance at county fairs. In addition, polyPLA can accurately separate the viruses at two contemporary swine IAV antigenic clusters (H3N2 swine IAV-α and H3N2 swine IAV-ß) with a sensitivity of 84.9% and a specificity of 100.0%. The polyPLA can be routinely used in surveillance programs to detect antigenic variants of influenza viruses and to select vaccine strains for use in controlling and preventing disease in swine. Copyright © 2017 American Society for Microbiology.

[Measures taken by a university hospital for the prevention and control of the 2009 H1N1 influenza].

PubMed

Hayashi, Jun; Murata, Masayuki; Furusyo, Norihiro; Hoshina, Takayuki; Shimono, Nobuyuki

2010-09-01

After extensive discussion with the Fukuoka City government of measures for the prevention and control of the 2009 H1N1 influenza pandemic, Kyushu University Hospital organized the infection control teams of 39 hospitals in the Fukuoka City area in preparation for a possible outbreak. A facility was set up at Kyushu University Hospital for the screening of outpatients with fever, and those with influenza and an underlying disease or severe symptoms were admitted to the hospital. 37 (22%) of the 171 outpatients with fever were infected with the new strain of influenza, confirmed by rapid influenza antigen test and PCR: Of these 37 patients, 17 (45.9%) were negative by influenza antigen test. Other 37 patients (5 adults, 32 children) were admitted, all of whom were successfully treated with neuraminidase inhibitors and discharged with no aftereffects.

Empirical validation of guidelines for the management of pharyngitis in children and adults.

PubMed

McIsaac, Warren J; Kellner, James D; Aufricht, Peggy; Vanjaka, Anita; Low, Donald E

2004-04-07

Recent guidelines for management of pharyngitis vary in their recommendations concerning empirical antibiotic treatment and the need for laboratory confirmation of group A streptococcus (GAS). To assess the impact of guideline recommendations and alternative approaches on identification and treatment of GAS pharyngitis in children and adults. Throat cultures and rapid antigen tests were performed on 787 children and adults aged 3 to 69 years with acute sore throat attending a family medicine clinic in Calgary, Alberta, from September 1999 to August 2002. Recommendations from 2 guidelines (those of the Infectious Diseases Society of America and of the American College of Physicians-American Society of Internal Medicine/American Academy of Family Physicians/US Centers for Disease Control and Prevention) were compared with rapid testing alone, a clinical prediction rule (ie, the modified Centor score), and a criterion standard of treatment for positive throat culture results only. Sensitivity and specificity of each strategy for identifying GAS pharyngitis, total antibiotics recommended, and unnecessary antibiotic prescriptions. In children, sensitivity for streptococcal infection ranged from 85.8% (133/155; 95% confidence interval [CI], 79.3%-90.0%) for rapid testing to 100% for culturing all. In adults, sensitivity ranged from 76.7% (56/73; 95% CI, 65.4%-85.8%) for rapid testing without culture confirmation of negative results to 100% for culturing all. In children, specificity ranged from 90.3% (270/299; 95% CI, 86.4%-93.4%) for use of modified Centor score and throat culture to 100% for culturing all. In adults, specificity ranged from 43.8% (114/260; 95% CI, 37.7%-50.1%) for empirical treatment based on a modified Centor score of 3 or 4 to 100% for culturing all. Total antibiotic prescriptions were lowest with rapid testing (24.7% [194/787]; 95% CI, 21.7%-27.8%) and highest with empirical treatment of high-risk adults (45.7% [360/787]; 95% CI, 42.2%-49.3%), due to a high rate of unnecessary prescriptions in adults (43.8% [146/333]; 95% CI, 38.4%-49.4%). Guideline recommendations for the selective use of throat cultures but antibiotic treatment based only on positive rapid test or throat culture results can reduce unnecessary use of antibiotics for treatment of pharyngitis. However, empirical treatment of adults having a Centor score of 3 or 4 is associated with a high rate of unnecessary antibiotic use. In children, strategies incorporating throat culture or throat culture confirmation of negative rapid antigen test results are highly sensitive and specific. Throat culture of all adults or those selected on the basis of a clinical prediction rule had the highest sensitivity and specificity.

Molecular diagnosis of Legionella infections--Clinical utility of front-line screening as part of a pneumonia diagnostic algorithm.

PubMed

Gadsby, Naomi J; Helgason, Kristjan O; Dickson, Elizabeth M; Mills, Jonathan M; Lindsay, Diane S J; Edwards, Giles F; Hanson, Mary F; Templeton, Kate E

2016-02-01

Urinary antigen testing for Legionella pneumophila serogroup 1 is the leading rapid diagnostic test for Legionnaires' Disease (LD); however other Legionella species and serogroups can also cause LD. The aim was to determine the utility of front-line L. pneumophila and Legionella species PCR in a severe respiratory infection algorithm. L. pneumophila and Legionella species duplex real-time PCR was carried out on 1944 specimens from hospitalised patients over a 4 year period in Edinburgh, UK. L. pneumophila was detected by PCR in 49 (2.7%) specimens from 36 patients. During a LD outbreak, combined L. pneumophila respiratory PCR and urinary antigen testing had optimal sensitivity and specificity (92.6% and 98.3% respectively) for the detection of confirmed cases. Legionella species was detected by PCR in 16 (0.9%) specimens from 10 patients. The 5 confirmed and 1 probable cases of Legionella longbeachae LD were both PCR and antibody positive. Front-line L. pneumophila and Legionella species PCR is a valuable addition to urinary antigen testing as part of a well-defined algorithm. Cases of LD due to L. longbeachae might be considered laboratory-confirmed when there is a positive Legionella species PCR result and detection of L. longbeachae specific antibody response. Copyright © 2015 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Preparation of immunochromatographic strips for rapid detection of early secreted protein ESAT-6 and culture filtrate protein CFP-10 from Mycobacterium tuberculosis

PubMed Central

Wu, Xiaoxin; Wang, Yeping; Weng, Tianhao; Hu, Chenyu; Wang, Frederick X.C.; Wu, Zhigang; Yu, Dongshan; Lu, Huoquan; Yao, Hangping

2017-01-01

Abstract The early secreted protein early secretory antigenic target 6(ESAT-6) and the culture filtrate protein 10 (CFP-10) are 2 antigens that are specific to Mycobacterium tuberculosis. These 2 antigens are good targets for tuberculosis (TB) detection. To rapidly diagnose TB across a variety of samples, we developed colloidal gold immunochromatographic strips (ICSs) based on ESAT-6 and CFP-10. The strips were evaluated using 233 samples, including sputum, plasma, and pleural effusion samples. The positive detection rates for ICSs for ESAT-6 and CFP-10 in sputum (culture-positive for M tuberculosis) were 100% and 91.2%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in plasma were 34.1% and 29.4%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in pleural effusion were 64.7% and 55.9%, respectively. Experimental analysis of culture supernatant showing that the ICS developed for ESAT-6 had a sensitivity of 100% and a specificity of 91.2%. While the ICS developed for CFP-10 had a sensitivity of 91.2% and a specificity of 88.2%. The validity of the test is limited by source of sample. The technique is sensitive and specific for samples in sputum and culture media but not for plasma or pleural effusion samples. Detection of M tuberculosis using ICSs is rapid, simple, and relatively effective; thus, ICSs are a potential screening tool for TB. PMID:29390519

Preparation of immunochromatographic strips for rapid detection of early secreted protein ESAT-6 and culture filtrate protein CFP-10 from Mycobacterium tuberculosis.

PubMed

Wu, Xiaoxin; Wang, Yeping; Weng, Tianhao; Hu, Chenyu; Wang, Frederick X C; Wu, Zhigang; Yu, Dongshan; Lu, Huoquan; Yao, Hangping

2017-12-01

The early secreted protein early secretory antigenic target 6(ESAT-6) and the culture filtrate protein 10 (CFP-10) are 2 antigens that are specific to Mycobacterium tuberculosis. These 2 antigens are good targets for tuberculosis (TB) detection.To rapidly diagnose TB across a variety of samples, we developed colloidal gold immunochromatographic strips (ICSs) based on ESAT-6 and CFP-10.The strips were evaluated using 233 samples, including sputum, plasma, and pleural effusion samples.The positive detection rates for ICSs for ESAT-6 and CFP-10 in sputum (culture-positive for M tuberculosis) were 100% and 91.2%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in plasma were 34.1% and 29.4%, respectively. The positive detection rates for ICSs for ESAT-6 and CFP-10 in pleural effusion were 64.7% and 55.9%, respectively. Experimental analysis of culture supernatant showing that the ICS developed for ESAT-6 had a sensitivity of 100% and a specificity of 91.2%. While the ICS developed for CFP-10 had a sensitivity of 91.2% and a specificity of 88.2%.The validity of the test is limited by source of sample. The technique is sensitive and specific for samples in sputum and culture media but not for plasma or pleural effusion samples. Detection of M tuberculosis using ICSs is rapid, simple, and relatively effective; thus, ICSs are a potential screening tool for TB. Copyright © 2017 The Authors. Published by Wolters Kluwer Health, Inc. All rights reserved.

Multicenter Clinical Evaluation of the Novel Alere i Strep A Isothermal Nucleic Acid Amplification Test.

PubMed

Cohen, Daniel M; Russo, Michael E; Jaggi, Preeti; Kline, Jennifer; Gluckman, William; Parekh, Amisha

2015-07-01

Rapid detection of group A beta-hemolytic streptococcus (GAS) is used routinely to help diagnose and treat pharyngitis. However, available rapid antigen detection tests for GAS have relatively low sensitivity, and backup testing is recommended in children. Newer assays are more sensitive yet require excessive time for practical point-of-care use as well as laboratory personnel. The Alere i strep A test is an isothermal nucleic acid amplification test designed to offer highly sensitive results at the point of care within 8 min when performed by nonlaboratory personnel. The performance of the Alere i strep A test was evaluated in a multicenter prospective trial in a Clinical Laboratory Improvement Amendments (CLIA)-waived setting in comparison to bacterial culture in 481 children and adults. Compared to culture, the Aleri i strep A test had 96.0% sensitivity and 94.6% specificity. Discrepant results were adjudicated by PCR and found the Alere i strep A test to have 98.7% sensitivity and 98.5% specificity. Overall, the Alere i strep A test could provide a one-step, rapid, point-of-care testing method for GAS pharyngitis and obviate backup testing on negative results. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Ten years of treating necrotizing fasciitis.

PubMed

Nordqvist, Gunnar; Walldén, Axel; Brorson, Håkan; Tham, Johan

2015-05-01

Necrotizing fasciitis is a soft tissue infection characterized by rapid progression and a high mortality rate. The objective of this study was to investigate diagnosis, causative microbial agents, comorbidities, antibiotic regimen and outcome regarding this disease at Skåne University Hospital in Malmö, Sweden. From medical records, we identified 33 patients treated from January 2003 to January 2013, 31 of whom could be included in our investigation. The infections were monomicrobial in 87% of the cases, and most were caused by group A streptococci. The rate of polymicrobial infections was lower than in other studies. In addition to blood and wound cultures, a rapid antigen detection test for group A streptococci was used in a majority of the cases as a supplement to other diagnostic tools. The time from onset of symptoms to surgery proved to be significantly shorter for patients infected with group A streptococci than for other patients. The mortality rate among all patients was 19%, which is lower than much of the historical material but in line with some more recent studies of this disease. Our results indicate that low mortality rates can be achieved by surgery, appropriate antibiotics and good supportive care. Furthermore, we show that the use of the rapid antigen detection test for group A streptococci, in this setting, helps to shorten the time to surgical intervention in patients suffering from necrotizing fasciitis. This also helps to guide the antibiotic treatment into a narrower spectrum.

Malaria rapid diagnostic tests in endemic settings.

PubMed

Maltha, J; Gillet, P; Jacobs, J

2013-05-01

Malaria rapid diagnostic tests (RDTs) are instrument-free tests that provide results within 20 min and can be used by community health workers. RDTs detect antigens produced by the Plasmodium parasite such as Plasmodium falciparum histidine-rich protein-2 (PfHPR2) and Plasmodium lactate dehydrogenase (pLDH). The accuracy of RDTs for the diagnosis of uncomplicated P. falciparum infection is equal or superior to routine microscopy (but inferior to expert microscopy). Sensitivity for Plasmodium vivax is 75-100%; for Plasmodium ovale and Plasmodium malariae, diagnostic performance is poor. Design limitations of RDTs include poor sensitivity at low parasite densities, susceptibility to the prozone effect (PfHRP2-detecting RDTs), false-negative results due to PfHRP2 deficiency in the case of pfhrp2 gene deletions (PfHRP2-detecting RDTs), cross-reactions between Plasmodium antigens and detection antibodies, false-positive results by other infections and susceptibility to heat and humidity. End-user's errors relate to safety, procedure (delayed reading, incorrect sample and buffer volumes) and interpretation (not recognizing invalid test results, disregarding faint test lines). Withholding antimalarial treatment in the case of negative RDT results tends to be infrequent and tendencies towards over-prescription of antibiotics have been noted. Numerous shortcomings in RDT kits' labelling, instructions for use (correctness and readability) and contents have been observed. The World Health Organization and partners actively address quality assurance of RDTs by comparative testing of RDTs, inspections of manufacturing sites, lot testing and training tools but no formal external quality assessment programme of end-user performance exists. Elimination of malaria requires RDTs with lower detection limits, for which nucleic acid amplification tests are under development. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

Multi-centre field evaluation of the performance of the Trinity Biotech Uni-Gold HIV 1/2 rapid test as a first-line screening assay for gay and bisexual men compared with 4th generation laboratory immunoassays.

PubMed

Keen, P; Conway, D P; Cunningham, P; McNulty, A; Couldwell, D L; Davies, S C; Smith, D E; Gray, J; Holt, M; O'Connor, C C; Read, P; Callander, D; Prestage, G; Guy, R

2017-01-01

The Trinity Biotech Uni-Gold HIV test (Uni-Gold) is often used as a supplementary rapid test in testing algorithms. To evaluate the operational performance of the Uni-Gold as a first-line screening test among gay and bisexual men (GBM) in a setting where 4th generation HIV laboratory assays are routinely used. We compared the performance of Uni-Gold with conventional HIV serology conducted in parallel among GBM attending 22 testing sites. Sensitivity was calculated separately for acute and established infection, defined using 4th generation screening Ag/Ab immunoassay (EIA) and Western blot results. Previous HIV testing history and results of supplementary 3rd generation HIV Ab EIA, and p24 antigen EIA were used to further characterise cases of acute infection. Of 10,793 specimens tested with Uni-Gold and conventional serology, 94 (0.90%, 95%CI:0.70-1.07) were confirmed as HIV-positive by conventional serology, and 37 (39.4%) were classified as acute infection. Uni-Gold sensitivity was 81.9% overall (77/94, 95%CI:72.6-89.1); 56.8% for acute infection (21/37, 95%CI:39.5-72.9) and 98.2% for established infection (56/57, 95%CI:90.6-100.0). Of 17 false non-reactive Uni-Gold results, 16 were acute infections, and of these seven were p24 antigen reactive but antibody negative. Uni-Gold specificity was 99.9% (10,692/10,699, 95%CI:99.9-100.0), PPV was 91.7% (95%CI:83.6-96.6) and NPV was 99.8% (95%CI:99.7-99.9), respectively. In this population, Uni-Gold had good specificity and sensitivity was high for established infections when compared to 4th generation laboratory assays, however sensitivity was lower in acute infections. Where rapid tests are used in populations with a high proportion of acute infections, additional testing strategies are needed to detect acute infections. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparative Study of Malaria Prevalence among Travellers in Nigeria (West Africa) Using Slide Microscopy and a Rapid Diagnosis Test.

PubMed

Dougnon, T V; Bankole, H S; Hounmanou, Y M G; Echebiri, S; Atchade, P; Mohammed, J

2015-01-01

Malaria is a major disease in Africa and leads to various public health problems. A study was carried out at the Aviation Medical Clinic Laboratory, Murtala Mohammed Airport, Ikeja, Lagos State, Nigeria, in 2014. The work aimed to determine the prevalence of malaria among patients attending the laboratory. Blood samples were therefore collected from 51 patients and subjected to both blood smear microscopy and a rapid immunochromatographic diagnostic test (SD BIOLINE Malaria Ag) for detection of, respectively, malaria parasites and antigens. At the end of the study, 22% of the patients were detected positive by the microscopic examination while 9.8% were tested positive when using SD BIOLINE Malaria Ag. The outcomes of the study show a high prevalence of malaria at the airport. This represents a serious risk factor leading to a high likelihood of spread and occurrence of malaria in other countries including Western countries whereby the disease is nonendemic. It also pointed out that the blood smear microscopy seems to be better than Rapid Diagnosis Test (RDT) for malaria diagnosis.

Evaluation of a rapid quantitative determination method of PSA concentration with gold immunochromatographic strips.

PubMed

Wu, Cheng-Ching; Lin, Hung-Yu; Wang, Chao-Ping; Lu, Li-Fen; Yu, Teng-Hung; Hung, Wei-Chin; Houng, Jer-Yiing; Chung, Fu-Mei; Lee, Yau-Jiunn; Hu, Jin-Jia

2015-11-03

Prostate cancer remains the most common cancer in men. Qualitative or semi-quantitative immunochromatographic measurements of prostate specific antigen (PSA) have been shown to be simple, noninvasive and feasible. The aim of this study was to evaluate an optimized gold immunochromatographic strip device for the detection of PSA, in which the results can be analysed using a Chromogenic Rapid Test Reader to quantitatively assess the test results. This reader measures the reflectance of the signal line via a charge-coupled device camera. For quantitative analysis, PSA concentration was computed via a calibration equation. Capillary blood samples from 305 men were evaluated, and two independent observers interpreted the test results after 12 min. Blood samples were also collected and tested with a conventional quantitative assay. Sensitivity, specificity, positive and negative predictive values, and accuracy of the PSA rapid quantitative test system were 100, 96.6, 89.5, 100, and 97.4 %, respectively. Reproducibility of the test was 99.2, and interobserver variation was 8 % with a false positive rate of 3.4 %. The correlation coefficient between the ordinary quantitative assay and the rapid quantitative test was 0.960. The PSA rapid quantitative test system provided results quickly and was easy to use, so that tests using this system can be easily performed at outpatient clinics or elsewhere. This system may also be useful for initial cancer screening and for point-of-care testing, because results can be obtained within 12 min and at a cost lower than that of conventional quantitative assays.

Simultaneous detection and serotyping of dengue infection using single tube multiplex CDC Dengue Real-Time RT-PCR from India.

PubMed

Sharma, Shashi; Tandel, Kundan; Danwe, Surabhi; Bhatt, Puneet; Dash, P K; Ranjan, Praveer; Rathi, K R; Gupta, Rajiv Mohan; Parida, M M

2018-03-01

Four antigenically different dengue virus serotypes (DENV-1, DENV-2, DENV-3 and DENV-4) are known to cause infections in humans. Some of these are known to cause more severe disease than the others. Chances for developing Dengue hemorrhagic fever-dengue shock syndrome (DHF-DSS) increases significantly with history of previous infection with one of the four serotypes. Therefore, early diagnosis, serotyping and providing early warning of dengue fever epidemics to concerned authorities becomes very important for better patient outcome and to curb the rapid spread in the community. During the 2014 outbreak, a total of 100 samples from suspected cases of dengue were collected. NS1 antigen based rapid test was used for serological diagnosis. Dengue complex one step reverse transcription-polymerase chain reaction was performed to look for presence of viral RNA. Single tube multiplex RT-PCR was also performed to look for infecting serotype. CDC Dengue Multiplex Real Time PCR assay was performed for rapid diagnosis and simultaneous serotyping of the dengue virus. Out of the 100 samples screened, 69 were found to be positive by NS1Ag Rapid test. 34 samples were found positive by dengue consensus RT-PCR assay. 22 samples were found to be positive by single tube Dengue multiplex RT-PCR assay. Serotype DEN-2 was present in maximum numbers followed by DEN-3. 44 samples were found positive by DENV CDC Multiplex Real time PCR assay. DEN-2 was found in maximum numbers followed by DEN-1. Dengue remains to be an important health problem in India and across the globe. Few serotypes of dengue are more dangerous than the others. Rapid diagnosis and serotyping remains the key for better patient management and prevention of disease spreading in the community. Highly sensitive, specific and rapid CDC real time RT-PCR assay was found to be most promising tool among all available molecular diagnostic methods. This will serve a rapid and reliable simultaneous dengue virus detection as well serotyping assay in near future for rapid identification of dengue suspected sample screening.

A survey of feline leukaemia virus infection of domestic cats from selected areas in Harare, Zimbabwe.

PubMed

Muchaamba, Francis; Mutiringindi, Takudzwa H; Tivapasi, Musavenga T; Dhliwayo, Solomon; Matope, Gift

2014-11-14

A cross-sectional study was conducted to detect the feline leukaemia virus (FeLV) p27 antigen and to determine risk factors and the haematological changes associated with infection in domestic cats in Zimbabwe. Sera were collected for detection of the p27 antigen, urea, creatinine, alanine aminotransferase and gamma-glutamyl transferase levels, whilst whole blood was collected for haematology. FeLV p27 antigen was detected using a rapid enzyme-linked immunosorbent assay (ELISA) test kit. Data on risk factors were analysed using a logistic regression model. Of the 100 cats tested, 41% (95% CI: 31.19% - 50.81%) (41/100) were positive for the FeLV p27 antigen. Sex and health status of cats were not significantly (p > 0.05) associated with infection. Intact cats (OR = 9.73), those living in multicat housing (OR = 5.23) and cats that had access to outdoor life (OR = 35.5) were found to have higher odds of infection compared with neutered cats, those living in single-cat housing, and without access to outdoor life, respectively. Biochemistry and haematology revealed no specific changes. The results showed that FeLV infection was high in sampled cats, providing evidence of active infection. Thus, it would be prudent to introduce specific control measures for FeLV infection in Zimbabwe.

Detection of hMPV antigen by EIA in clinical specimens.

PubMed

Pancer, Katarzyna; Ciaćka, Agnieszka; Gut, Włodzimierz; Lipka, Bozena; Mierzejewska, Justyna; Milewska-Bobula, Bogumiła; Smorczewska-Kiljan, Anna; Jahnz-Rózyk, Karina; Litwińska, Bogumiła

2011-01-01

Human Metapneumovirus (hMPV) is one of the latest discovered viruses. It has been classified to Paramyxoviridae family. It is the second viral etiological agent, after RSV, which causes respiratory tract infections (RTI) in children, especially children below 5 years old. It is estimated that 5-25% of RTI in children is due to hMPV. In adults hMPV reinfections are bounded to upper respiratory tract infections. The aim of the study was to establish usefulness of ELISA test in detecting hMPV antigen and to analyze hMPV infection in connection to clinical diagnosis. 273 nasopharyngeal swabs from children (189 swabs) and adults (84 swabs) with respiratory tract infections collected from 2008 to 2010 were examined. Due to similarity of hMPV and RSV viruses and overlapping of their epidemic season rapid immunochromatographic test for RSV antigen detection was also performed in case of 120 samples, hMPV antigen was detected in 24.5% of all swabs (n = 67): in 0.0% probes in 2008, 29.0% in 2009 and 36.8% in first quarter of 2010. The highest rate ofhMPV infection was detected from summer of 2009 till the end of March 2010 (VIII-IX 2009 - 62.5%, X-XII 2009 - 44.1% and I-III 2010 -36.8%). We analyzed respiratory tract diseases reported in patients with hMPV infection. Infection due to hMPV was found in 26.5% of children and 24.0% of adults with recognized pneumonia, respectively in 28.4 and 17.6% of patients with bronchitis. Bronchiolitis was diagnosed in two children with hMPV. RSV and hMPV coinfections were confirmed in 15 out of 120 examined probes. Cross reaction pattern was excluded thanks to ELISA hMPV antigen test which was performed with suspension of RSV and thanks to statistical analysis. Coinfections were confirmed in 8% of pneumonia, 11% of bronchitis and 24.2% of the rest concomitant diagnoses. We found hMPV infection as the significant agent ofpneumonia not only in children but also in adults. ELISA hMPV antigen test can be used in diagnosis of etiological agent of respiratory infections in children and adults and in coinfections as well.

Recovery from desensitization of IgE-dependent responses in human lung mast cells.

PubMed

Lewis, A; MacGlashan, D W; Suvarna, S K; Peachell, P T

2017-08-01

Clinical desensitization and oral food immunotherapy are therapeutic interventions that allow individuals who react adversely to an allergen (drug or food) to be made tolerant to the allergen. However, tolerance is brief, and allergen hypersensitivity can recur within days following allergen withdrawal. We hypothesize that the reason these treatments are temporary reflects rapid recovery of mast cells from a desensitized state. We sought to test this. Desensitization of IgE-mediated histamine release from human lung mast cells was explored by methods that partially replicate the pattern of treatment during clinical desensitization. Specific and non-specific desensitization and changes in surface IgE were examined following desensitization. Recovery from desensitization was also studied. Desensitization of mast cell responses was readily induced with concentrations of antigen or anti-IgE that were suboptimal for secretion. There was little or no non-specific desensitization when lung mast cells were exposed to antigens. There was no loss of cell surface IgE following desensitization. Removing the desensitizing stimulus from the media following desensitization allowed the cells to recover with half-point of recovery of ~1.5 days and complete recovery after 5 days. Both the functional response and histamine content recovered within this time frame. The recovery appeared possible because both antigens and anti-IgE dissociated rapidly from cells after washing to remove excess stimulus. Human lung mast cells readily recover from a desensitized state following removal of desensitizing antigen. This finding provides a potential explanation for the ephemeral nature of clinical desensitization. © 2017 John Wiley & Sons Ltd.

Evaluation of a new in-clinic test system to detect feline immunodeficiency virus and feline leukemia virus infection.

PubMed

Sand, Christina; Englert, Theresa; Egberink, Herman; Lutz, Hans; Hartmann, Katrin

2010-06-01

Many in-house tests for the diagnosis of feline immunodeficiency virus (FIV) and feline leukemia virus (FeLV) infection are licensed for use in veterinary practice. A new test with unknown performance has recently appeared on the market. The aims of this study were to define the efficacy of a new in-clinic test system, the Anigen Rapid FIV Ab/FeLV Ag Test, and to compare it with the current leading in-clinic test, the SNAP Kombi Plus FeLV Antigen/FIB Antibody Test. Three-hundred serum samples from randomly selected healthy and diseased cats presented to the Clinic of Small Animal Medicine at Ludwig Maximilian University were tested using both the Anigen Rapid Test and the SNAP Kombi Plus Test. Diagnostic sensitivity, specificity, and positive and negative predictive values were calculated for both tests using Western blot as the gold standard for verification of FIV infection and PCR as the gold standard for FeLV infection. The presence of antibodies against FIV was confirmed by Western blot in 9/300 samples (prevalence 3%). FeLV DNA was detected by PCR in 15/300 samples (prevalence 5%). For FIV infection the Anigen Rapid Test had a sensitivity of 88.9%, specificity of 99.7%, positive predictive value of 88.9%, and negative predictive value of 99.7%. For FeLV infection, the Anigen Rapid Test had a sensitivity of 40.0%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 96.9%. Diagnostic accuracy was similar to that of the SNAP Kombi Plus Test. The new Anigen Rapid FIV Ab/FeLV Ag Test performed very well and can be recommended for use in veterinary practice.

Development and Evaluation of an Enzyme-Linked Immunosorbent Assay To Detect Histoplasma capsulatum Antigenuria in Immunocompromised Patients▿

PubMed Central

Scheel, Christina M.; Samayoa, Blanca; Herrera, Alejandro; Lindsley, Mark D.; Benjamin, Lynette; Reed, Yvonne; Hart, John; Lima, Sandra; Rivera, Blanca E.; Raxcaco, Gabriella; Chiller, Tom; Arathoon, Eduardo; Gómez, Beatriz L.

2009-01-01

Histoplasma capsulatum infection causes significant morbidity and mortality in human immunodeficiency virus-infected individuals, particularly those in countries with limited access to rapid diagnostics or antiretroviral therapies. The fungus easily disseminates in persons with AIDS, resulting in progressive disseminated histoplasmosis (PDH), which can progress rapidly to death if undiagnosed. The availability of a simple, rapid method to detect H. capsulatum infection in less developed countries where the infection is endemic would dramatically decrease the time to diagnosis and treatment of PDH. We have developed an antigen-capture enzyme-linked immunosorbent assay (ELISA) to detect PDH antigenuria in infected patients. The assay uses polyclonal antibodies against H. capsulatum as both capture and detection reagents, and a standard reference curve is included to quantify antigenuria and ensure reproducibility. We evaluated this assay using specimens collected from patients with AIDS and culture-proven histoplasmosis in a Guatemalan clinic (n = 48), from healthy persons (n = 83), and from patients with other, nonhistoplasmosis diseases (n = 114). The ELISA demonstrated a sensitivity of 81% and a specificity of 95% in detecting H. capsulatum antigen in urine. This assay relies on simple technology that can be performed in institutions with limited resources. Use of this test will facilitate rapid diagnosis of PDH in countries where mortality is high, expediting treatment and likely reducing PDH-related mortality. PMID:19357311

Conventional Rapid Latex Agglutination in Estimation of von Willebrand Factor: Method Revisited and Potential Clinical Applications

PubMed Central

Che Hussin, Che Maraina

2014-01-01

Measurement of von Willebrand factor antigen (VWF : Ag) levels is usually performed in a specialised laboratory which limits its application in routine clinical practice. So far, no commercial rapid test kit is available for VWF : Ag estimation. This paper discusses the technical aspect of latex agglutination method which was established to suit the purpose of estimating von Willebrand factor (VWF) levels in the plasma sample. The latex agglutination test can be performed qualitatively and semiquantitatively. Reproducibility, stability, linearity, limit of detection, interference, and method comparison studies were conducted to evaluate the performance of this test. Semiquantitative latex agglutination test was strongly correlated with the reference immunoturbidimetric assay (Spearman's rho = 0.946, P < 0.001, n = 132). A substantial agreement (κ = 0.77) was found between qualitative latex agglutination test and the reference assay. Using the scoring system for the rapid latex test, no agglutination is with 0% VWF : Ag (control negative), 1+ reaction is equivalent to <20% VWF : Ag, and 4+ reaction indicates >150% VWF : Ag (when comparing with immunoturbidimetric assay). The findings from evaluation studies suggest that latex agglutination method is suitable to be used as a rapid test kit for the estimation of VWF : Ag levels in various clinical conditions associated with high levels and low levels of VWF : Ag. PMID:25759835

Assessing the relationship between antigenicity and immunogenicity of human rabies vaccines when administered by intradermal route

PubMed Central

Bilagumba, Gangaboraiah; Ravish, Haradanahalli Shankarappa; Narayana, Hanumanthappa Ashwath Doddabele

2010-01-01

The metadata of 10 published studies and 3 vaccine trial reports comprising of 19 vaccine cohorts from four countries conducted over a period of 23 years (1986–2009) was used for metaanalysis. The vaccines studied were purified chick embryo cell vaccine (Rabipur, India and Germany), purified vero cell rabies vaccine (Verorab, France; Indirab, India) and human diploid cell vaccine (MIRV, France). The potency of these vaccines varied from 0.55 IU to 2.32 IU per intradermal dose of 0.1 ml per site. The vaccines were administered to 1,011 subjects comprising of 19 cohorts and using five different ID regimens. The immunogenicity was measured by assays of rabies virus neutralizing antibody (RVNA) titres using rapid fluorescent focus inhibition test (RFFIT) [15 cohorts] and mouse neutralization test (MNT) [4 cohorts]. The statistical analysis of the data was done by Karl Pearson's correlation coefficient to measure the relationship between antigenicity and immunogenicity. It was revealed that, there was no significant linear relationship between antigenicity and immunogenicity of rabies vaccines when administered by intradermal route (p > 0.230 and p > 0.568). PMID:20523131

Protection conferred by recombinant Yersinia pestis antigens produced by a rapid and highly scalable plant expression system

PubMed Central

Santi, Luca; Giritch, Anatoli; Roy, Chad J.; Marillonnet, Sylvestre; Klimyuk, Victor; Gleba, Yuri; Webb, Robert; Arntzen, Charles J.; Mason, Hugh S.

2006-01-01

Plague is still an endemic disease in different regions of the world. Increasing reports of incidence, the discovery of antibiotic resistance strains, and concern about a potential use of the causative bacteria Yersinia pestis as an agent of biological warfare have highlighted the need for a safe, efficacious, and rapidly producible vaccine. The use of F1 and V antigens and the derived protein fusion F1-V has shown great potential as a protective vaccine in animal studies. Plants have been extensively studied for the production of pharmaceutical proteins as an inexpensive and scalable alternative to common expression systems. In the current study the recombinant plague antigens F1, V, and fusion protein F1-V were produced by transient expression in Nicotiana benthamiana by using a deconstructed tobacco mosaic virus-based system that allowed very rapid and extremely high levels of expression. All of the plant-derived purified antigens, administered s.c. to guinea pigs, generated systemic immune responses and provided protection against an aerosol challenge of virulent Y. pestis. PMID:16410352

Streptococcus group B typing: comparison of counter-immunoelectrophoresis with the precipitin method.

PubMed

Kubín, V; Jelínková, J; Franêk, J

1977-07-01

The method of counter-immunoelectrophoresis (CIE) was tested for its applicability to group B streptococcus typing. The results obtained were compared with the typing by the ring precipitin test. Identical antigens and identical hyperimmune typing serum batches had been used in both methods. A large majority of 75 freshly isolated strains were typed identically by both methods. Five strains with a weak antigenic outfit were untypable by the ring precipitin test but were typed by CIE owing to a higher sensitivity of CIE method. Two strains were typable by the precipitin test but not by CIE; an explanation for this phenomenon is lacking. The CIE method in group B typing is specific, rapid, highly sensitive and relatively simple. It requires strict maintenance of standard conditions. The method is economical with respect to manipulation and material, requires small amounts of diagnostic antisera. Potent antisera may be used diluted. Moreover, sera for CIE typing need not be absorbed to remove group B antibodies. CIE method is practicable for group B streptococcus typing, especially in laboratories carrying out routine large scale type identification.

Validation of a urine circulating cathodic antigen cassette test for detection of Schistosoma haematobiumin uMkhanyakude district of South Africa.

PubMed

Rubaba, O; Chimbari, M J; Soko, W; Manyangadze, T; Mukaratirwa, S

2018-06-01

Circulating cathodic antigen (CCA) tests for schistosomiasis are fast and less complicated allowing making them good candidates for routine qualitative screening for schistosomiasis at point of care. The urine-CCA has been evaluated for detection of S. mansoni with promising results. Its specificity and consistency in detecting S. haematobium infection in different endemic regions has been variable. This study validated a rapid urine-CCA cassette test for qualitative detection of S. haematobium infection in an S. haematobium endemic area with low S. mansoni prevalence. Microscopic examination for the standard urine filtration technique was used to validate the commercially available urine-CCA cassette test (rapid medical diagnostics ® ). The validation was done in a sample of primary school pupils (n = 420) aged 10-15 years in schools in the Jozini Municipality, KZN. There was a relationship between infection intensity and a positive urine-CCA test. Using the urine filtration method as the gold standard, the prevalence for S. haematobium was 40%, the accuracy of the CCA kit was 54.8%, sensitivity was 68.1% while the specificity was 45.8%. The positive predictive value was 45.82% while the negative predictive value was 68.05%. Both the urine filtration and the urine-CCA methods detected heavy (≥50 eggs/10 mL urine) and light infections at statistically significant levels. The overall accuracy, sensitivity and specificity of the urine-CCA cassette test were low. The urine-CCA cassette test performed much better for heavy infections than low infections (p < 0.05) implying that the kit may not be suitable for low endemic areas. Copyright © 2018 Elsevier B.V. All rights reserved.

Additional Progress in the Development and Application of a Direct, Rapid Immunohistochemical Test for Rabies Diagnosis.

PubMed

Rupprecht, Charles E; Xiang, Zhiquan; Servat, Alexandre; Franka, Richard; Kirby, Jordona; Ertl, Hildegund C J

2018-06-20

Laboratory-based surveillance is fundamental to effective rabies prevention and control. The direct fluorescent antibody (AB) test (FAT) is the gold standard for rabies diagnosis. Recently, additional tests besides the FAT have been developed, such as the direct rapid immunohistochemical test (DRIT). In this study, our objective was to further refine technical aspects of the DRIT using a combination of two monoclonal ABs (MABs), 502 and 802, conduct additional testing among rabies reference laboratories using a diversity of animal species and rabies virus (RV) variants and compare the potential utility of the DRIT for end users via proficiency testing (PT) against the FAT. Considering the ideal molar ratios of biotin to AB in formulation of the DRIT conjugate, 3.9 was found to be superior to 7.4, for detection of RV antigens in the brain of a naturally infected raccoon. Optimization of the DRIT conjugate may also be dependent upon the apparent choice of specific viral antigens for testing, as a gray fox RV variant reacted less strongly than a raccoon RV variant in determining the working dilution of the MAB cocktail. Using the same MABs and protocol, the DRIT was compared to the FAT using more than 800 samples of mammalian brains, representative of more than 25 taxa, including in excess of 250 animal rabies cases from Europe and North America. Sensitivity was determined at 98% (96⁻100%, 95% CI) and specificity was calculated at 95% (92⁻96%, 95% CI). In a comparison among end users, PT of laboratory personnel resulted in values of 77⁻100% sensitivity and 86-100% specificity. Based upon these and previously reported results, the DRIT appears to be a suitable alternative to the FAT for use in lyssavirus diagnosis.

Heteroassembled gold nanoparticles with sandwich-immunoassay LSPR chip format for rapid and sensitive detection of hepatitis B virus surface antigen (HBsAg).

PubMed

Kim, Jinwoon; Oh, Seo Yeong; Shukla, Shruti; Hong, Seok Bok; Heo, Nam Su; Bajpai, Vivek K; Chun, Hyang Sook; Jo, Cheon-Ho; Choi, Bong Gill; Huh, Yun Suk; Han, Young-Kyu

2018-06-01

This study aimed to develop a more sensitive method for the detection of hepatitis B surface antigen (HBsAg) using heteroassembled gold nanoparticles (AuNPs). A single layered localized surface plasmon resonance (LSPR) chip format was developed with antigen-antibody reaction-based detection symmetry using AuNPs, which detected HBsAg at 10 pg/mL. To further improve the detection limit, a modified detection format was fabricated by fixing a secondary antibody (to form a heteroassembled sandwich format) to the AuNP monolayer, which enhanced the detection sensitivity by about 100 times. The developed heteroassembled AuNPs sandwich-immunoassay LSPR chip format was able to detect as little as 100 fg/mL of HBsAg within 10-15 min. In addition, the heteroassembled AuNPs sandwich-immunoassay LSPR chip format did not show any non-specific binding to other tested antigens, including alpha fetoprotein (AFP), C-reactive protein (CRP), and prostate-specific antigen (PSA). These findings confirm that the proposed detection strategy of heteroassembled AuNPs sandwich-immunoassay LSPR chip format may provide a new platform for early diagnosis of various human diseases. Copyright © 2018 Elsevier B.V. All rights reserved.

The diagnostic accuracy of three rapid diagnostic tests for typhoid fever at Chittagong Medical College Hospital, Chittagong, Bangladesh.

PubMed

Maude, Rapeephan R; de Jong, Hanna K; Wijedoru, Lalith; Fukushima, Masako; Ghose, Aniruddha; Samad, Rasheda; Hossain, Mohammed Amir; Karim, Mohammed Rezaul; Faiz, Mohammed Abul; Parry, Christopher M

2015-10-01

To determine the diagnostic accuracy of three rapid diagnostic tests (RDTs) for typhoid fever in febrile hospitalised patients in Bangladesh. Febrile adults and children admitted to Chittagong Medical College Hospital, Bangladesh, were investigated with Bact/Alert(®) blood cultures and real-time PCR to detect Salmonella enterica Typhi and Paratyphi A and assays for Rickettsia, leptospirosis and dengue fever. Acute serum samples were examined with the LifeAssay (LA) Test-it™ Typhoid IgM lateral flow assay detecting IgM antibodies against S. Typhi O antigen, CTKBiotech Onsite Typhoid IgG/IgM Combo Rapid-test cassette lateral flow assay detecting IgG and IgM antibodies against S. Typhi O and H antigens and SD Bioline line assay for IgG and IgM antibodies against S. Typhi proteins. In 300 malaria smear-negative febrile patients [median (IQR) age of 13.5 (5-31) years], 34 (11.3%) had confirmed typhoid fever: 19 positive by blood culture for S. Typhi (three blood PCR positive) and 15 blood culture negative but PCR positive for S. Typhi in blood. The respective sensitivity and specificity of the three RDTs in patients using a composite reference standard of blood culture and/or PCR-confirmed typhoid fever were 59% and 61% for LifeAssay, 59% and 74% for the CTK IgM and/or IgG, and 24% and 96% for the SD Bioline RDT IgM and/or IgG. The LifeAssay RDT had a sensitivity of 63% and a specificity of 91% when modified with a positive cut-off of ≥2+ and analysed using a Bayesian latent class model. These typhoid RDTs demonstrated moderate diagnostic accuracies, and better tests are needed. © 2015 The Authors. Tropical Medicine & International Health Published by John Wiley & Sons Ltd.

BRUCELLOSIS IN TEREKEKA COUNTY, CENTRAL EQUATORIA STATE, SOUTHERN SUDAN.

PubMed

Lado, D; Maina, N; Lado, M; Abade, A; Amwayi, S; Omolo, J; Oundo, J

2012-01-01

To identify factors associated with Brucellosis in patients attending Terekeka Health Facility, Terekeka County, Central Equatoria State, Southern Sudan and to evaluate the utility of the rapid test kit Euracil®. A facility based case-control study. Terekeka Health Facility, Terekeka County, Central Equatoria State, Southern Sudan. Cases were patients presenting at the Terekeka Health Facility with clinical symptoms suggestive of Brucellosis and tested positive for Brucellosis by rapid antigen test while controls were selected from individuals attending Terekeka Health facility with health problems unrelated to brucellosis or febrile illness. A total of fifty eight cases with clinical symptoms suggestive of and tested positive for Brucellosis by rapid antigen test presented. A total of 116 consented controls were recruited into the study. Males accounted for 52% of the cases and 53% of the controls. The mean age was 31 years for both groups. Cases without formal education were 84% while 40% had no source of income, 20% of the cases and 14% of the controls were cattle keepers while 5% of the cases and 13% of the controls were students. In multivariate analysis there were many factors associated with Brucellosis like consumption of raw meat, living with animals at the same place, raising of goats, farm cleaning contact, eating of aborted and wild animals. Logistic regression revealed two factors associated with the disease; consumption of raw milk (OR=3.9, P-value 0.001, 95% CI 1.6666-9.0700) was a risk factor while drinking boiled milk was protective (OR = 0.09, p-value 0.000, 95% CI, 0.1-0.2). The main age-groups affected were 20-30 years with males being affected more than females. Drinking of raw milk was significantly associated with Brucellosis while drinking boiled milk was protective. There should be active public health education on the benefits of boiling milk before consumption. Further studies to elucidate the extent and epidemiology of brucellosis in humans and animals in Southern Sudan are recommended.

Laboratory Validation of the Sand Fly Fever Virus Antigen Assay

DTIC Science & Technology

2015-12-01

TOSV), sandfly fever Sicilian virus (SFSV), sandfly fever Naples virus (SFNV), and Punta Toro virus (Tesh 1988 , Alkan et al . 2013). These viruses pose a...of meningitis in Mediter- ranean and southern European countries during the vector season (Braito et al . 1997). Sandfly fever Sicilian virus also...stationed there (Peralta et al . 1965). Rapid field assessments of sand flies for phleboviruses have been previously unavailable. The available tests are

Molecular Diagnostics in Transfusion Medicine: In Capillary, on a Chip, in Silico, or in Flight?

PubMed Central

Garritsen, Henk S.P.; Xiu-Cheng Fan, Alex; Lenz, Daniela; Hannig, Horst; Yan Zhong, Xiao; Geffers, Robert; Lindenmaier, Werner; Dittmar, Kurt E.J.; Wörmann, Bernhard

2009-01-01

Summary Serology, defined as antibody-based diagnostics, has been regarded as the diagnostic gold standard in transfusion medicine. Nowadays however the impact of molecular diagnostics in transfusion medicine is rapidly growing. Molecular diagnostics can improve tissue typing (HLA typing), increase safety of blood products (NAT testing of infectious diseases), and enable blood group typing in difficult situations (after transfusion of blood products or prenatal non-invasive RhD typing). Most of the molecular testing involves the determination of the presence of single nucleotide polymorphisms (SNPs). Antigens (e.g. blood group antigens) mostly result from single nucleotide differences in critical positions. However, most blood group systems cannot be determined by looking at a single SNP. To identify members of a blood group system a number of critical SNPs have to be taken into account. The platforms which are currently used to perform molecular diagnostics are mostly gel-based, requiring time-consuming multiple manual steps. To implement molecular methods in transfusion medicine in the future the development of higher-throughput SNP genotyping non-gel-based platforms which allow a rapid, cost-effective screening are essential. Because of its potential for automation, high throughput and cost effectiveness the special focus of this paper is a relative new technique: SNP genotyping by MALDI-TOF MS analysis. PMID:21113259

Rapid screening and identification of dominant B cell epitopes of HBV surface antigen by quantum dot-based fluorescence polarization assay

NASA Astrophysics Data System (ADS)

Meng, Zhongji; Song, Ruihua; Chen, Yue; Zhu, Yang; Tian, Yanhui; Li, Ding; Cui, Daxiang

2013-03-01

A method for quickly screening and identifying dominant B cell epitopes was developed using hepatitis B virus (HBV) surface antigen as a target. Eleven amino acid fragments from HBV surface antigen were synthesized by 9-fluorenylmethoxy carbonyl solid-phase peptide synthesis strategy, and then CdTe quantum dots were used to label the N-terminals of all peptides. After optimizing the factors for fluorescence polarization (FP) immunoassay, the antigenicities of synthetic peptides were determined by analyzing the recognition and combination of peptides and standard antibody samples. The results of FP assays confirmed that 10 of 11 synthetic peptides have distinct antigenicities. In order to screen dominant antigenic peptides, the FP assays were carried out to investigate the antibodies against the 10 synthetic peptides of HBV surface antigen respectively in 159 samples of anti-HBV surface antigen-positive antiserum. The results showed that 3 of the 10 antigenic peptides may be immunodominant because the antibodies against them existed more widely among the samples and their antibody titers were higher than those of other peptides. Using three dominant antigenic peptides, 293 serum samples were detected for HBV infection by FP assays; the results showed that the antibody-positive ratio was 51.9% and the sensitivity and specificity were 84.3% and 98.2%, respectively. In conclusion, a quantum dot-based FP assay is a very simple, rapid, and convenient method for determining immunodominant antigenic peptides and has great potential in applications such as epitope mapping, vaccine designing, or clinical disease diagnosis in the future.

A strategy to identify linker-based modules for the allosteric regulation of antibody-antigen binding affinities of different scFvs

PubMed Central

Thie, Holger

2017-01-01

ABSTRACT Antibody single-chain variable fragments (scFvs) are used in a variety of applications, such as for research, diagnosis and therapy. Essential for these applications is the extraordinary specificity, selectivity and affinity of antibody paratopes, which can also be used for efficient protein purification. However, this use is hampered by the high affinity for the protein to be purified because harsh elution conditions, which may impair folding, integrity or viability of the eluted biomaterials, are typically required. In this study, we developed a strategy to obtain structural elements that provide allosteric modulation of the affinities of different antibody scFvs for their antigen. To identify suitable allosteric modules, a complete set of cyclic permutations of calmodulin variants was generated and tested for modulation of the affinity when substituting the linker between VH and VL. Modulation of affinity induced by addition of different calmodulin-binding peptides at physiologic conditions was demonstrated for 5 of 6 tested scFvs of different specificities and antigens ranging from cell surface proteins to haptens. In addition, a variety of different modulator peptides were tested. Different structural solutions were found in respect of the optimal calmodulin permutation, the optimal peptide and the allosteric effect for scFvs binding to different antigen structures. Significantly, effective linker modules were identified for scFvs with both VH-VL and VL-VH architecture. The results suggest that this approach may offer a rapid, paratope-independent strategy to provide allosteric regulation of affinity for many other antibody scFvs. PMID:28055297

Influenza A virus hemagglutinin glycosylation compensates for antibody escape fitness costs.

PubMed

Kosik, Ivan; Ince, William L; Gentles, Lauren E; Oler, Andrew J; Kosikova, Martina; Angel, Matthew; Magadán, Javier G; Xie, Hang; Brooke, Christopher B; Yewdell, Jonathan W

2018-01-01

Rapid antigenic evolution enables the persistence of seasonal influenza A and B viruses in human populations despite widespread herd immunity. Understanding viral mechanisms that enable antigenic evolution is critical for designing durable vaccines and therapeutics. Here, we utilize the primerID method of error-correcting viral population sequencing to reveal an unexpected role for hemagglutinin (HA) glycosylation in compensating for fitness defects resulting from escape from anti-HA neutralizing antibodies. Antibody-free propagation following antigenic escape rapidly selected viruses with mutations that modulated receptor binding avidity through the addition of N-linked glycans to the HA globular domain. These findings expand our understanding of the viral mechanisms that maintain fitness during antigenic evolution to include glycan addition, and highlight the immense power of high-definition virus population sequencing to reveal novel viral adaptive mechanisms.

Prostate-Specific Antigen (PSA) Test: MedlinePlus Lab Test Information

MedlinePlus

... gov/labtests/prostatespecificantigenpsatest.html Prostate-Specific Antigen (PSA) Test To use the sharing features on this page, ... JavaScript. What is a prostate-specific antigen (PSA) test? A prostate-specific antigen (PSA) test measures the ...

Canine parvovirus--a review of epidemiological and diagnostic aspects, with emphasis on type 2c.

PubMed

Decaro, Nicola; Buonavoglia, Canio

2012-02-24

Canine parvovirus type 2 (CPV-2) emerged in late 1970s causing severe epizootics in kennels and dog shelters worldwide. Soon after its emergence, CPV-2 underwent genetic evolution giving rise consecutively to two antigenic variants, CPV-2a and CPV-2b that replaced progressively the original type. In 2000, a new antigenic variant, CPV-2c, was detected in Italy and rapidly spread to several countries. In comparison to the original type CPV-2, the antigenic variants display increased pathogenicity in dogs and extended host range, being able to infect and cause disease in cats. Epidemiological survey indicate that the newest type CPV-2c is becoming prevalent in different geographic regions and is often associated to severe disease in adult dogs and also in dogs that have completed the vaccination protocols. However, the primary cause of failure of CPV vaccination is interference by maternally derived immunity. Diagnosis of CPV infection by traditional methods has been shown to be poorly sensitive, especially in the late stages of infections. New diagnostic approaches based on molecular methods have been developed for sensitive detection of CPV in clinical samples and rapid characterisation of the viral type. Continuous surveillance will help assess whether there is a real need to update currently available vaccines and diagnostic tests. Copyright © 2011 Elsevier B.V. All rights reserved.

The limitations of point of care testing for pandemic influenza: what clinicians and public health professionals need to know.

PubMed

Hatchette, Todd F; Bastien, Nathalie; Berry, Jody; Booth, Tim F; Chernesky, Max; Couillard, Michel; Drews, Steven; Ebsworth, Anthony; Fearon, Margaret; Fonseca, Kevin; Fox, Julie; Gagnon, Jean-Nicolas; Guercio, Steven; Horsman, Greg; Jorowski, Cathy; Kuschak, Theodore; Li, Yan; Majury, Anna; Petric, Martin; Ratnam, Sam; Smieja, Marek; Van Caeseele, Paul

2009-01-01

As the world prepares for the next influenza pandemic, governments have made significant funding commitments to vaccine development and antiviral stockpiling. While these are essential components to pandemic response, rapid and accurate diagnostic testing remains an often neglected cornerstone of pandemic influenza preparedness. Clinicians and Public Health Practitioners need to understand the benefits and drawbacks of different influenza tests in both seasonal and pandemic settings. Culture has been the traditional gold standard for influenza diagnosis but requires from 1-10 days to generate a positive result, compared to nucleic acid detection methods such as real time reverse transcriptase polymerase chain reaction (RT-PCR). Although the currently available rapid antigen detection kits can generate results in less than 30 minutes, their sensitivity is suboptimal and they are not recommended for the detection of novel influenza viruses. Until point-of-care (POC) tests are improved, PILPN recommends that the best option for pandemic influenza preparation is the enhancement of nucleic acid-based testing capabilities across Canada.

An adenovirus-vectored nasal vaccine confers rapid and sustained protection against anthrax in a single-dose regimen.

PubMed

Zhang, Jianfeng; Jex, Edward; Feng, Tsungwei; Sivko, Gloria S; Baillie, Leslie W; Goldman, Stanley; Van Kampen, Kent R; Tang, De-chu C

2013-01-01

Bacillus anthracis is the causative agent of anthrax, and its spores have been developed into lethal bioweapons. To mitigate an onslaught from airborne anthrax spores that are maliciously disseminated, it is of paramount importance to develop a rapid-response anthrax vaccine that can be mass administered by nonmedical personnel during a crisis. We report here that intranasal instillation of a nonreplicating adenovirus vector encoding B. anthracis protective antigen could confer rapid and sustained protection against inhalation anthrax in mice in a single-dose regimen in the presence of preexisting adenovirus immunity. The potency of the vaccine was greatly enhanced when codons of the antigen gene were optimized to match the tRNA pool found in human cells. In addition, an adenovirus vector encoding lethal factor can confer partial protection against inhalation anthrax and might be coadministered with a protective antigen-based vaccine.

An Adenovirus-Vectored Nasal Vaccine Confers Rapid and Sustained Protection against Anthrax in a Single-Dose Regimen

PubMed Central

Jex, Edward; Feng, Tsungwei; Sivko, Gloria S.; Baillie, Leslie W.; Goldman, Stanley; Van Kampen, Kent R.; Tang, De-chu C.

2013-01-01

Bacillus anthracis is the causative agent of anthrax, and its spores have been developed into lethal bioweapons. To mitigate an onslaught from airborne anthrax spores that are maliciously disseminated, it is of paramount importance to develop a rapid-response anthrax vaccine that can be mass administered by nonmedical personnel during a crisis. We report here that intranasal instillation of a nonreplicating adenovirus vector encoding B. anthracis protective antigen could confer rapid and sustained protection against inhalation anthrax in mice in a single-dose regimen in the presence of preexisting adenovirus immunity. The potency of the vaccine was greatly enhanced when codons of the antigen gene were optimized to match the tRNA pool found in human cells. In addition, an adenovirus vector encoding lethal factor can confer partial protection against inhalation anthrax and might be coadministered with a protective antigen-based vaccine. PMID:23100479

Immunodiagnosis of Paracoccidioidomycosis due to Paracoccidioides brasiliensis Using a Latex Test: Detection of Specific Antibody Anti-gp43 and Specific Antigen gp43

PubMed Central

dos Santos, Priscila Oliveira; Rodrigues, Anderson Messias; Fernandes, Geisa Ferreira; da Silva, Silvia Helena Marques; Burger, Eva; de Camargo, Zoilo Pires

2015-01-01

Background Paracoccidioidomycosis (PCM) is a life-threatening systemic disease and is a neglected public health problem in many endemic regions of Latin America. Though several diagnostic methods are available, almost all of them present with some limitations. Method/Principle Findings A latex immunoassay using sensitized latex particles (SLPs) with gp43 antigen, the immunodominant antigen of Paracoccidioides brasiliensis, or the monoclonal antibody mAb17c (anti-gp43) was evaluated for antibody or antigen detection in sera, cerebrospinal fluid (CSF), and bronchoalveolar lavage (BAL) from patients with PCM due to P. brasiliensis. The gp43-SLPs performed optimally to detect specific antibodies with high levels of sensitivity (98.46%, 95% CI 91.7–100.0), specificity (93.94%, 95% CI 87.3–97.7), and positive (91.4%) and negative (98.9%) predictive values. In addition, we propose the use of mAb17c-SLPs to detect circulating gp43, which would be particularly important in patients with immune deficiencies who fail to produce normal levels of immunoglobulins, achieving good levels of sensitivity (96.92%, 95% CI 89.3–99.6), specificity (88.89%, 95% CI 81.0–94.3), and positive (85.1%) and negative (97.8%) predictive values. Very good agreement between latex tests and double immune diffusion was observed for gp43-SLPs (k = 0.924) and mAb17c-SLPs (k = 0.850), which reinforces the usefulness of our tests for the rapid diagnosis of PCM in less than 10 minutes. Minor cross-reactivity occurred with sera from patients with other fungal infections. We successfully detected antigens and antibodies from CSF and BAL samples. In addition, the latex test was useful for monitoring PCM patients receiving therapy. Conclusions/Significance The high diagnostic accuracy, low cost, reduced assay time, and simplicity of this new latex test offer the potential to be commercialized and makes it an attractive diagnostic assay for use not only in clinics and medical mycology laboratories, but mainly in remote locations with limited laboratory infrastructure and/or minimally trained community health workers. PMID:25679976

Diagnostic performance of influenza viruses and RSV rapid antigen detection tests in children in tertiary care.

PubMed

Moesker, F M; van Kampen, J J A; Aron, G; Schutten, M; van de Vijver, D A M C; Koopmans, M P G; Osterhaus, A D M E; Fraaij, P L A

2016-06-01

Rapid antigen detection tests (RADTs) are increasingly used to detect influenza viruses and respiratory syncytial virus (RSV). However, their sensitivity and specificity are a matter of debate, challenging their clinical usefulness. Comparing diagnostic performances of BinaxNow Influenza AB(®) (BNI) and BinaxNow RSV(®) (BNR), to those of real-time reverse transcriptase PCR (RT-PCR), virus isolation and direct immunofluorescence (D-IF) in paediatric patients. Between November 2005 and September 2013, 521 nasal washings from symptomatic children (age <5 years) attending our tertiary care centre were tested, with a combination of the respective assays using RT-PCR as gold standard. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of BNI were 69% (confidence interval [CI] [51-83]), 96% [94-97], 55% [39-70] and 98% [96-99] respectively. Of eleven false-negative samples, RT-PCR Ct-values were higher than all RT-PCR positive test results (27 vs 22, p=0.012). Of twenty false-positive samples, none were culture positive and two tested positive in D-IF. Sensitivity, specificity, PPV and NPV for BNR were 79% [73-85], 98% [96-99], 97% [93-99] and 88% [84-91]. Of the 42 false-negative samples the median Ct-value was higher than that of all RT-PCR positive samples (31 vs 23, p<0.0001). Five false-positive samples were detected. Three of these tested positive for RSV in virus isolation and D-IF. RADTs have a high specificity with BNR being superior to BNI. However, their relative low sensitivity limits their usefulness for clinical decision making in a tertiary care paediatric hospital. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Health Workers' Use of Malaria Rapid Diagnostic Tests (RDTs) to Guide Clinical Decision Making in Rural Dispensaries, Tanzania

PubMed Central

Masanja, M. Irene; McMorrow, Meredith; Kahigwa, Elizeus; Kachur, S. Patrick; McElroy, Peter D.

2010-01-01

Rapid diagnostic tests (RDTs) were developed as an alternative to microscopy for malaria diagnosis. The RDTs detect malaria parasite antigen(s) in whole blood with high sensitivity and specificity. We assessed health worker malaria treatment practices after the introduction of RDTs in peripheral health facilities without microscopy. From December 2007 to October 2008, we introduced histidine-rich protein II (HRP-2)-based ParaHIT RDTs for routine use in 12 health facilities in Rufiji District, Tanzania. Health workers received training on how to perform RDTs for patients 5 years of age or older with fever or suspected malaria. Children < 5 years of age were to be treated empirically per national guidelines. Among the 30,195 patients seen at these 12 health facilities, 10,737 (35.6%) were tested with an RDT for malaria. 88.3% (9,405/10,648) of tested patients reported fever or history of fever and 2.7% (289/10,677) of all tested individuals were children < 5 years of age. The RDT results were recorded for 10,650 patients (99.2%). Among the 5,488 (51.5%) RDT-positive patients, 5,256 (98.6%) were treated with an appropriate first-line antimalarial per national guidelines (artemether-lumefantrine or quinine). Among the 5,162 RDT-negative patients, only 205 (4.0%) were treated with an antimalarial. Other reported treatments included antibiotics and antipyretics. Implementation of RDTs in rural health facilities resulted in high adherence to national treatment guidelines. Patients testing negative by RDT were rarely treated with antimalarials. Unapproved antimalarials were seldom used. Health workers continued to follow guidelines for the empiric treatment of febrile children. PMID:21118927

Diagnostic Tests to Support Late-Stage Control Programs for Schistosomiasis and Soil-Transmitted Helminthiases.

PubMed

Hawkins, Kenneth R; Cantera, Jason L; Storey, Helen L; Leader, Brandon T; de Los Santos, Tala

2016-12-01

Global efforts to address schistosomiasis and soil-transmitted helminthiases (STH) include deworming programs for school-aged children that are made possible by large-scale drug donations. Decisions on these mass drug administration (MDA) programs currently rely on microscopic examination of clinical specimens to determine the presence of parasite eggs. However, microscopy-based methods are not sensitive to the low-intensity infections that characterize populations that have undergone MDA. Thus, there has been increasing recognition within the schistosomiasis and STH communities of the need for improved diagnostic tools to support late-stage control program decisions, such as when to stop or reduce MDA. Failure to adequately address the need for new diagnostics could jeopardize achievement of the 2020 London Declaration goals. In this report, we assess diagnostic needs and landscape potential solutions and determine appropriate strategies to improve diagnostic testing to support control and elimination programs. Based upon literature reviews and previous input from experts in the schistosomiasis and STH communities, we prioritized two diagnostic use cases for further exploration: to inform MDA-stopping decisions and post-MDA surveillance. To this end, PATH has refined target product profiles (TPPs) for schistosomiasis and STH diagnostics that are applicable to these use cases. We evaluated the limitations of current diagnostic methods with regards to these use cases and identified candidate biomarkers and diagnostics with potential application as new tools. Based on this analysis, there is a need to develop antigen-detecting rapid diagnostic tests (RDTs) with simplified, field-deployable sample preparation for schistosomiasis. Additionally, there is a need for diagnostic tests that are more sensitive than the current methods for STH, which may include either a field-deployable molecular test or a simple, low-cost, rapid antigen-detecting test.

Uniform cell-autonomous tumorigenesis of the choroid plexus by papovavirus large T antigens.

PubMed Central

Chen, J D; Van Dyke, T

1991-01-01

The simian virus 40 (SV40) large tumor antigen (T antigen) under its natural regulatory elements induces choroid plexus papillomas in transgenic mice. Because these tumors develop focally after several months, it has been suggested that secondary cellular alterations are required to induce a tumor in this tissue. In contrast to SV40, the related lymphotropic papovavirus early region induces rapid nonfocal choroid plexus neoplasia in transgenic mice. Here, using hybrid gene constructs, we showed that T antigen from either virus in in fact sufficient to induce these tumors. Their abilities to induce proliferative abnormalities in other tissues, such as kidney and thymus, were also indistinguishable. Differences in the rate of choroid plexus tumorigenesis reflected differences in the control regions of the two viruses, rather than differences in T antigen per se. Under SV40 regulation, expression was limited to a fraction of the choroid plexus cells prior to the formation of focal tumors. When SV40 T antigen was placed under lymphotropic papovavirus control, in contrast, expression was generally uniform in the choroid plexus and rapid expansion of the tissue ensued. We found a direct relationship between T-antigen expression, morphological transformation, and proliferation of the choroid plexus epithelial cells. Analysis of mosaic transgenic mice indicated further that T antigen exerts its mitogenic effect cell autonomously. These studies form the foundation for elucidating the role of various T-antigen subactivities in tumorigenesis. Images PMID:1658622

Comparison of two antigen assays for rapid intrapartum detection of vaginal group B streptococcal colonization.

PubMed

Green, M; Dashefsky, B; Wald, E R; Laifer, S; Harger, J; Guthrie, R

1993-01-01

As part of a clinical investigation evaluating the efficacy of intrapartum antigen detection for screening for heavy vaginal colonization with group B streptococci (GBS), we compared the performance of modified Bactigen and Directigen GBS latex particle agglutination (LPA) kits. Paired vaginal swabs obtained from women in labor were rapidly transported to the laboratory and used for culturing (both swabs) and LPA testing (one swab by each method). GBS growth was estimated semiquantitatively and further designated as light or heavy growth. Performance specifications for each method were determined by comparing LPA and culture results from the same swab. A total of 4,251 paired swabs were evaluated during the study period. The performance specifications for detecting GBS growth of any degree for Bactigen and Directigen, respectively, were as follows: sensitivity, 20 and 24%; specificity, 99 and 99%. The performance specifications for heavy colonization for Bactigen and Directigen, respectively, were as follows: sensitivity, 57 and 62%; specificity, 99 and 99%. Neither LPA kit was a sensitive indicator of vaginal colonization with GBS or neonatal infection.

A large Legionnaires' disease outbreak in Pamplona, Spain: early detection, rapid control and no case fatality

PubMed Central

CASTILLA, J.; BARRICARTE, A.; ALDAZ, J.; GARCÍA CENOZ, M.; FERRER, T.; PELAZ, C.; PINEDA, S.; BALADRÓN, B.; MARTÍN, I.; GOÑI, B.; ARATAJO, P.; CHAMORRO, J.; LAMEIRO, F.; TORROBA, L.; DORRONSORO, I.; MARTÍNEZ-ARTOLA, V.; ESPARZA, M. J.; GASTAMINZA, M. A.; FRAILE, P.; ALDAZ, P.

2008-01-01

SUMMARY An outbreak of Legionnaire's disease was detected in Pamplona, Spain, on 1 June 2006. Patients with pneumonia were tested to detect Legionella pneumophila antigen in urine (Binax Now; Binax Inc., Scarborough, ME, USA), and all 146 confirmed cases were interviewed. The outbreak was related to district 2 (22 012 inhabitants), where 45% of the cases lived and 50% had visited; 5% lived in neighbouring districts. The highest incidence was found in the resident population of district 2 (3/1000 inhabitants), section 2 (14/1000). All 31 cooling towers of district 2 were analysed. L. pneumophila antigen (Binax Now) was detected in four towers, which were closed on 2 June. Only the strain isolated in a tower situated in section 2 of district 2 matched all five clinical isolates, as assessed by mAb and two genotyping methods, AFLP and PFGE. Eight days after closing the towers, new cases ceased appearing. Early detection and rapid coordinated medical and environmental actions permitted immediate control of the outbreak and probably contributed to the null case fatality. PMID:17662166

A large Legionnaires' disease outbreak in Pamplona, Spain: early detection, rapid control and no case fatality.

PubMed

Castilla, J; Barricarte, A; Aldaz, J; García Cenoz, M; Ferrer, T; Pelaz, C; Pineda, S; Baladrón, B; Martín, I; Goñi, B; Aratajo, P; Chamorro, J; Lameiro, F; Torroba, L; Dorronsoro, I; Martínez-Artola, V; Esparza, M J; Gastaminza, M A; Fraile, P; Aldaz, P

2008-06-01

An outbreak of Legionnaire's disease was detected in Pamplona, Spain, on 1 June 2006. Patients with pneumonia were tested to detect Legionella pneumophila antigen in urine (Binax Now; Binax Inc., Scarborough, ME, USA), and all 146 confirmed cases were interviewed. The outbreak was related to district 2 (22 012 inhabitants), where 45% of the cases lived and 50% had visited; 5% lived in neighbouring districts. The highest incidence was found in the resident population of district 2 (3/1000 inhabitants), section 2 (14/1000). All 31 cooling towers of district 2 were analysed. L. pneumophila antigen (Binax Now) was detected in four towers, which were closed on 2 June. Only the strain isolated in a tower situated in section 2 of district 2 matched all five clinical isolates, as assessed by mAb and two genotyping methods, AFLP and PFGE. Eight days after closing the towers, new cases ceased appearing. Early detection and rapid coordinated medical and environmental actions permitted immediate control of the outbreak and probably contributed to the null case fatality.

Correlation between the immunological condition and the results of immunoenzymatic tests in diagnosing infectious mononucleosis.

PubMed

Tamaro, Giorgio; Donato, Michela; Princi, Tanja; Parco, Sergio

2009-04-01

A symptom-based diagnosis of infectious mononucleosis is not sufficiently accurate, since some clinical symptoms of infectious mononucleosis are also detected in other virally induced diseases. Moreover, not all patients suffering from infectious mononucleosis show circulating atypical lymphocytes, which are considered characteristic of this disease. Therefore, when this disorder is suspected, serum analyses are carried out to detect the presence of certain immunoglobulins associated with infectious mononucleosis in the patient's blood. The aim of this study was to evaluate the sensitivity and the specificity of a rapid test detecting heterophil antibodies in diagnosing infectious mononucleosis in a paediatric population. We considered 163 paediatric patients with suspected infectious mononucleosis and we tested their serums to detect heterophil antibodies (using an inexpensive and rapid test) and specific immunoglobulins directed against Epstein-Barr virus (EBV) (these assays are known to be characterized by high sensitivity and specificity, but are more expensive and time-consuming). By comparing the results of the rapid test with those of the other assays, we found that the sensitivity of the first test was 61.8%, whereas its specificity was sufficiently high (about 90%). We show that, in paediatric patients, the detection of heterophil antibodies is not a very sensitive test, therefore the determination of immunoglobulins against specific antigens of EBV is recommended.

Field validation of recombinant antigen immunoassays for diagnosis of Lassa fever.

PubMed

Boisen, Matthew L; Hartnett, Jessica N; Shaffer, Jeffrey G; Goba, Augustine; Momoh, Mambu; Sandi, John Demby; Fullah, Mohamed; Nelson, Diana K S; Bush, Duane J; Rowland, Megan M; Heinrich, Megan L; Koval, Anatoliy P; Cross, Robert W; Barnes, Kayla G; Lachenauer, Anna E; Lin, Aaron E; Nekoui, Mahan; Kotliar, Dylan; Winnicki, Sarah M; Siddle, Katherine J; Gbakie, Michael; Fonnie, Mbalu; Koroma, Veronica J; Kanneh, Lansana; Kulakosky, Peter C; Hastie, Kathryn M; Wilson, Russell B; Andersen, Kristian G; Folarin, Onikepe O; Happi, Christian T; Sabeti, Pardis C; Geisbert, Thomas W; Saphire, Erica Ollmann; Khan, S Humarr; Grant, Donald S; Schieffelin, John S; Branco, Luis M; Garry, Robert F

2018-04-12

Lassa fever, a hemorrhagic fever caused by Lassa virus (LASV), is endemic in West Africa. It is difficult to distinguish febrile illnesses that are common in West Africa from Lassa fever based solely on a patient's clinical presentation. The field performance of recombinant antigen-based Lassa fever immunoassays was compared to that of quantitative polymerase chain assays (qPCRs) using samples from subjects meeting the case definition of Lassa fever presenting to Kenema Government Hospital in Sierra Leone. The recombinant Lassa virus (ReLASV) enzyme-linked immunosorbant assay (ELISA) for detection of viral antigen in blood performed with 95% sensitivity and 97% specificity using a diagnostic standard that combined results of the immunoassays and qPCR. The ReLASV rapid diagnostic test (RDT), a lateral flow immunoassay based on paired monoclonal antibodies to the Josiah strain of LASV (lineage IV), performed with 90% sensitivity and 100% specificity. ReLASV immunoassays performed better than the most robust qPCR currently available, which had 82% sensitivity and 95% specificity. The performance characteristics of recombinant antigen-based Lassa virus immunoassays indicate that they can aid in the diagnosis of LASV Infection and inform the clinical management of Lassa fever patients.

[Diagnostic tools for canine parvovirus infection].

PubMed

Proksch, A L; Hartmann, K

2015-01-01

Canine parvovirus (CPV) infection is one of the most important and common infectious diseases in dogs, in particular affecting young puppies when maternal antibodies have waned and vaccine-induced antibodies have not yet developed. The mortality rate remains high. Therefore, a rapid and safe diagnostic tool is essential to diagnose the disease to 1) provide intensive care treatment and 2) to identify virus-shedding animals and thus prevent virus spread. Whilst the detection of antibodies against CPV is considered unsuitable to diagnose the disease, there are several different methods to directly detect complete virus, virus antigen or DNA. Additionally, to test in commercial laboratories, rapid in-house tests based on ELISA are available worldwide. The specificity of the ELISA rapid in-house tests is reported to be excellent. However, results on sensitivity vary and high numbers of false-negative results are commonly reported, which potentially leads to misdiagnosis. Polymerase chain reaction (PCR) is a very sensitive and specific diagnostic tool. It also provides the opportunity to differentiate vaccine strains from natural infection when sequencing is performed after PCR.

Immune response to measles vaccine in Peruvian children.

PubMed Central

Bautista-López, N. L.; Vaisberg, A.; Kanashiro, R.; Hernández, H.; Ward, B. J.

2001-01-01

OBJECTIVE: To evaluate the immune response in Peruvian children following measles vaccination. METHODS: Fifty-five Peruvian children received Schwarz measles vaccine (about 10(3) plaque forming units) at about 9 months of age. Blood samples were taken before vaccination, then twice after vaccination: one sample at between 1 and 4 weeks after vaccination and the final sample 3 months post vaccination for evaluation of immune cell phenotype and lymphoproliferative responses to measles and non-measles antigens. Measles-specific antibodies were measured by plaque reduction neutralization. FINDINGS: The humoral response developed rapidly after vaccination; only 4 of the 55 children (7%) had plaque reduction neutralization titres <200 mlU/ml 3 months after vaccination. However, only 8 out of 35 children tested (23%) had lymphoproliferative responses to measles antigens 3-4 weeks after vaccination. Children with poor lymphoproliferative responses to measles antigens had readily detectable lymphoproliferative responses to other antigens. Flow cytometric analysis of peripheral blood mononuclear cells revealed diffuse immune system activation at the time of vaccination in most children. The capacity to mount a lymphoproliferative response to measles antigens was associated with expression of CD45RO on CD4+ T-cells. CONCLUSION: The 55 Peruvian children had excellent antibody responses after measles vaccination, but only 23% (8 out of 35) generated detectable lymphoproliferative responses to measles antigens (compared with 55-67% in children in the industrialized world). This difference may contribute to the less than uniform success of measles vaccination programmes in the developing world. PMID:11731811

Direct Competitive Enzyme-Linked Immunosorbent Assay (ELISA).

PubMed

Kohl, Thomas O; Ascoli, Carl A

2017-07-05

The competitive enzyme-linked immunosorbent assay (ELISA) (cELISA; also called an inhibition ELISA) is designed so that purified antigen competes with antigen in the test sample for binding to an antibody that has been immobilized in microtiter plate wells. The same concept works if the immobilized molecule is antigen and the competing molecules are purified labeled antibody versus antibody in a test sample. Direct cELISAs incorporate labeled antigen or antibody, whereas indirect assay configurations use reporter-labeled secondary antibodies. The cELISA is very useful for determining the concentration of small-molecule antigens in complex sample mixtures. In the direct cELISA, antigen-specific capture antibody is adsorbed onto the microtiter plate before incubation with either known standards or unknown test samples. Enzyme-linked antigen (i.e., labeled antigen) is also added, which can bind to the capture antibody only when the antibody's binding site is not occupied by either the antigen standard or antigen in the test samples. Unbound labeled and unlabeled antigens are washed away and substrate is added. The amount of antigen in the standard or the test sample determines the amount of reporter-labeled antigen bound to antibody, yielding a signal that is inversely proportional to antigen concentration within the sample. Thus, the higher the antigen concentration in the test sample, the less labeled antigen is bound to the capture antibody, and hence the weaker is the resultant signal. © 2017 Cold Spring Harbor Laboratory Press.

Evaluation of the rapid influenza detection tests GOLD SIGN FLU and Quick Navi-Flu for the detection of influenza A and B virus antigens in adults during the influenza season.

PubMed

Akaishi, Yu; Matsumoto, Tetsuya; Harada, Yoshimi; Hirayama, Yoji

2016-11-01

As the characteristics and accuracy of rapid influenza detection tests (RIDTs) vary, the development of a high-performance RIDT has been eagerly anticipated. In this study, the new RIDT GOLD SIGN FLU and the existing RIDT Quick Navi-Flu were evaluated in terms of detecting the antigens of influenza viruses A and B in Japanese adults with influenza-like symptoms. The study was performed from December 2013 to March 2014. Among the 123 patients from whom nasopharyngeal swab specimens were collected, 59 tested positive by viral isolation as the gold standard method (influenza A, n=38; influenza B, n=21). For GOLD SIGN FLU, the sensitivities were 73.7% and 81.0%, and the specificities were 97.6% and 98.0% for influenza A and B, respectively. For Quick Navi-Flu, the sensitivities were 86.8% and 85.7%, and the specificities were 98.8% and 100% for influenza A and B, respectively. The time to the appearance of the line on the test strip was less than 3min for influenza A and less than 2min for influenza B with both RIDTs in more than 90% of cases. GOLD SIGN FLU was useful for diagnosing influenza A, and the result was readily available for influenza B particularly among adult patients. Quick Navi-Flu showed better sensitivities and specificities than GOLD SIGN FLU. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Standardization and validation of a cytometric bead assay to assess antibodies to multiple Plasmodium falciparum recombinant antigens.

PubMed

Ondigo, Bartholomew N; Park, Gregory S; Gose, Severin O; Ho, Benjamin M; Ochola, Lyticia A; Ayodo, George O; Ofulla, Ayub V; John, Chandy C

2012-12-21

Multiplex cytometric bead assay (CBA) have a number of advantages over ELISA for antibody testing, but little information is available on standardization and validation of antibody CBA to multiple Plasmodium falciparum antigens. The present study was set to determine optimal parameters for multiplex testing of antibodies to P. falciparum antigens, and to compare results of multiplex CBA to ELISA. Antibodies to ten recombinant P. falciparum antigens were measured by CBA and ELISA in samples from 30 individuals from a malaria endemic area of Kenya and compared to known positive and negative control plasma samples. Optimal antigen amounts, monoplex vs multiplex testing, plasma dilution, optimal buffer, number of beads required were assessed for CBA testing, and results from CBA vs. ELISA testing were compared. Optimal amounts for CBA antibody testing differed according to antigen. Results for monoplex CBA testing correlated strongly with multiplex testing for all antigens (r = 0.88-0.99, P values from <0.0001 - 0.004), and antibodies to variants of the same antigen were accurately distinguished within a multiplex reaction. Plasma dilutions of 1:100 or 1:200 were optimal for all antigens for CBA testing. Plasma diluted in a buffer containing 0.05% sodium azide, 0.5% polyvinylalcohol, and 0.8% polyvinylpyrrolidone had the lowest background activity. CBA median fluorescence intensity (MFI) values with 1,000 antigen-conjugated beads/well did not differ significantly from MFI with 5,000 beads/well. CBA and ELISA results correlated well for all antigens except apical membrane antigen-1 (AMA-1). CBA testing produced a greater range of values in samples from malaria endemic areas and less background reactivity for blank samples than ELISA. With optimization, CBA may be the preferred method of testing for antibodies to P. falciparum antigens, as CBA can test for antibodies to multiple recombinant antigens from a single plasma sample and produces a greater range of values in positive samples and lower background readings for blank samples than ELISA.

Dual Electrophoresis Detection System for Rapid and Sensitive Immunoassays with Nanoparticle Signal Amplification

NASA Astrophysics Data System (ADS)

Zhang, Fangfang; Ma, Junjie; Watanabe, Junji; Tang, Jinlong; Liu, Huiyu; Shen, Heyun

2017-02-01

An electrophoretic technique was combined with an enzyme-linked immunosorbent assay (ELISA) system to achieve a rapid and sensitive immunoassay. A cellulose acetate filter modified with polyelectrolyte multilayer (PEM) was used as a solid substrate for three-dimensional antigen-antibody reactions. A dual electrophoresis process was used to induce directional migration and local condensation of antigens and antibodies at the solid substrate, avoiding the long diffusion times associated with antigen-antibody reactions in conventional ELISAs. The electrophoretic forces drove two steps in the ELISA process, namely the adsorption of antigen, and secondary antibody-labelled polystyrene nanoparticles (NP-Ab). The total time needed for dual electrophoresis-driven detection was just 4 min, nearly 2 h faster than a conventional ELISA system. Moreover, the rapid NP-Ab electrophoresis system simultaneously achieved amplification of the specific signal and a reduction in noise, leading to a more sensitive NP-Ab immunoassay with a limit of detection (LOD) of 130 fM, and wide range of detectable concentrations from 0.13 to 130 pM. These results suggest that the combination of dual electrophoresis detection and NP-Ab signal amplification has great potential for future immunoassay systems.

Rapid diagnostic testing for community-acquired pneumonia: can innovative technology for clinical microbiology be exploited?

PubMed

Yu, Victor L; Stout, Janet E

2009-12-01

Two nonsynchronous events have affected the management of community-acquired pneumonia (CAP): spiraling empiricism for CAP and the "golden era" of clinical microbiology. The development of broad-spectrum antibiotics has led to widespread empiric use without ascertaining the etiology of the infecting microbe. Unfortunately, this approach clashes with the second event, which is the advent of molecular-based microbiology that can identify the causative pathogen rapidly at the point of care. The urinary antigen is a most effective rapid test that has allowed targeted therapy for Legionnaire disease at the point of care. The high specificity (> 90%) allows the clinician to administer appropriate anti-Legionella therapy based on a single rapid test; however, its low sensitivity (76%) means that a notable number of cases of Legionnaire disease will go undiagnosed if other tests, especially culture, are not performed. Further, culture for Legionella is not readily available. If a culture is not performed, epidemiologic identification of the source of the bacterium cannot be ascertained by molecular fingerprinting of the patient and the putative source strain. We recommend resurrection of the basic principles of infectious disease, which are to identify the microbial etiology of the infection and to use narrow, targeted antimicrobial therapy. To reduce antimicrobial overuse with subsequent antimicrobial resistance, these basic principles must be applied in concert with traditional and newer tests in the clinical microbiology laboratory.

Evaluation of two 4th generation point-of-care assays for the detection of Human Immunodeficiency Virus infection.

PubMed

Stafylis, Chrysovalantis; Klausner, Jeffrey D

2017-01-01

Fourth generation assays detect simultaneously antibodies for HIV and the p24 antigen, identifying HIV infection earlier than previous generation tests. Previous studies have shown that the Alere Determine HIV-1/2 Combo has lower than anticipated performance in detecting antibodies for HIV and the p24 antigen. Furthermore, there are currently very few studies evaluating the performance of Standard Diagnostics BIOLINE HIV Ag/Ab Combo. To evaluate the performance of the Alere Determine HIV-1/2 Combo and the Standard Diagnostics BIOLINE HIV Ag/Ab Combo in a panel of frozen serum samples. The testing panel included 133 previously frozen serum specimens from the UCLA Clinical Microbiology & Immunoserology laboratory. Reference testing included testing for HIV antibodies by a 3rd generation enzyme immunoassay followed by HIV RNA detection. Antibody negative and RNA positive sera were also tested by a laboratory 4th generation HIV Ab/Ag enzyme immunoassay. Reference testing yielded 97 positives for HIV infection and 36 negative samples. Sensitivity of the Alere test was 95% (88-98%), while the SD Bioline sensitivity was 91% (83-96%). Both assays showed 100% (90-100%) specificity. No indeterminate or invalid results were recorded. Among 13 samples with acute infection (HIV RNA positive, HIV antibody negative), 12 were found positive by the first assay and 8 by the second. The antigen component of the Alere assay detected 10 acute samples, while the SD Bioline assay detected only one. Both rapid assays showed very good overall performance in detecting HIV infection in frozen serum samples, but further improvements are required to improve the performance in acute infection.

Effects of Different Cell-Detaching Methods on the Viability and Cell Surface Antigen Expression of Synovial Mesenchymal Stem Cells.

PubMed

Tsuji, Kunikazu; Ojima, Miyoko; Otabe, Koji; Horie, Masafumi; Koga, Hideyuki; Sekiya, Ichiro; Muneta, Takeshi

2017-06-09

Flow cytometric analysis of cell surface antigens is a powerful tool for the isolation and characterization of stem cells residing in adult tissues. In contrast to the collection of hematopoietic stem cells, the process of enzymatic digestion is usually necessary to prepare mesenchymal stem cells (MSCs) suspensions, which can influence the expression of cell surface markers. In this study, we examined the effects of various cell-detaching reagents and digestion times on the expression of stem cell-related surface antigens and MSC functions. Human MSCs were detached from dishes using four different reagents: trypsin, TrypLE, collagenase, and a nonenzymatic cell dissociation reagent (C5789; Sigma-Aldrich). Following dissociation reagent incubations ranging from 5 to 120 min, cell surface markers were analyzed by flow cytometry. Trypsin and TrypLE quickly dissociated the cells within 5 min, while collagenase and C5789 required 60 min to obtain maximum cell yields. C5789 significantly decreased cell viability at 120 min. Trypsin treatment significantly reduced CD44+, CD55+, CD73+, CD105+, CD140a+, CD140b+, and CD201+ cell numbers within 30 min. Collagenase treatment reduced CD140a expression by 30 min. In contrast, TrypLE treatment did not affect the expression of any cell surface antigens tested by 30 min. Despite the significant loss of surface antigen expression after 60 min of treatment with trypsin, adverse effects of enzymatic digestion on multipotency of MSCs were limited. Overall, our data indicated that TrypLE is advantageous over other cell dissociation reagents tested for the rapid preparation of viable MSC suspensions.

Nested polymerase chain reaction on blood clots for gene encoding 56 kDa antigen and serology for the diagnosis of scrub typhus.

PubMed

Prakash, J A J; Kavitha, M L; Mathai, E

2011-01-01

Scrub typhus is a zoonotic illness endemic in the Asia-Pacific region. Early diagnosis and appropriate management contribute significantly to preventing adverse outcomes including mortality. Serology is widely used for diagnosing scrub typhus. Recent reports suggest that polymerase chain reaction (PCR) could be a rapid and reliable alternative. This study assessed the utility of these tests for scrub typhus diagnosis. Nested PCR to detect the 56 kDa antigen gene of O. tsutsugamushi was performed on blood clots from 87 individuals with clinically suspected scrub typhus. Weil-Felix test and scrub typhus IgM ELISA were performed on serum samples from the same patients. As a gold standard reference test was not available, latent class analysis (LCA) was used to assess the performance of the three tests. The LCA analysis showed the sensitivity of Weil-Felix test, IgM ELISA and PCR to be 59%, 100% and 58% respectively. The specificity of ELISA was only 73%, whereas those of the Weil-Felix test and PCR were 94% and 100% respectively. Nested PCR using blood clots while specific, lacked sensitivity as compared to IgM ELISA. In resource-poor settings Weil-Felix test still remains valuable despite its moderate sensitivity.

Development and Evaluation of an Immunodiffusion Test for Diagnosis of Systemic Zygomycosis (Mucormycosis): Preliminary Report

PubMed Central

Jones, Kenneth W.; Kaufman, Leo

1978-01-01

An antigen analysis with filtrate and homogenate precipitinogens of single isolates of the zygomycetes Absidia corymbifera, Mucor pusillus, Rhizopus arrhizus, and Rhizopus oryzae demonstrated the presence of common antigens among the three genera as well as antigens which permit their differentiation. Selected homogenate antigens were valuable in developing a diagnostic immunodiffusion (ID) test for systemic zygomycosis. When sera from 43 patients with various proven mycoses other than zygomycosis were tested against each of the antigens, none formed precipitin bands identical to those formed by A. cormybifera, M. pusillus, and the Rhizopus spp. rabbit reference antisera. Sera from 23 normal persons and 25 diabetics did not react with any of the antigens. Homogenate antigens detected antibody in 8 of the 11 sera (73%) from suspected or proven cases of zygomycosis, whereas ID tests with filtrate antigens detected antibody in only 2 of the 11 sera (18%). Of the eight sera that reacted with the homogenate antigens, five only reacted with a specific Rhizopus sp. antigen, two only reacted with a specific M. pusillus antigen, and one only reacted with a specific A. corymbifera antigen. Study results show the ID test with homogenate antigens to be more specific and sensitive than the ID test with filtrate antigens and indicate that the former is a promising technique for diagnosing human zygomycosis. Images PMID:75212

Antigen persistence of rapid diagnostic tests in pregnant women in Nanoro, Burkina Faso, and the implications for the diagnosis of malaria in pregnancy.

PubMed

Kattenberg, Johanna H; Tahita, Christian M; Versteeg, Inge A J; Tinto, Halidou; Traoré-Coulibaly, Maminata; Schallig, Henk D F H; Mens, Petra F

2012-05-01

To evaluate persistence of several Plasmodium antigens in pregnant women after treatment and compare diagnostics during treatment follow-up. Thirty-two pregnant women (N = 32) with confirmed malaria infection by a histidine-rich protein 2 (HRP2)-based rapid diagnostic test (RDT) and microscopy were followed for 28 days after artemisinin-based combination therapy (ACT). A Plasmodium lactate dehydrogenase (pLDH)-based RDT and two ELISAs based on the detection of dihydrofolate reductase-thymidylate synthase (DHFR-TS) and haeme detoxification protein (HDP) were compared with each other and to RT-PCR at each visit. The mean visit number (95% confidence interval) on which the HRP2-based RDT was still positive after treatment was 3.4 (2.7-4.1) visits with some patients still positive at day 28. This is significantly later than the pLDH-based RDT [0.84 (0.55-1.1)], microscopy (median 1, range 1-3), DHFR-TS-ELISA [1.7 (1.1-2.3)] and RT-PCR (median 2, range 1-5) (P < 0.05), but not significantly later than HDP-ELISA [2.1 (1.6-2.7)]. Lower gravidity and higher parasite density at day 0 resulted in significantly longer positive results with most tests (P < 0.05). HRP2 can persist up to 28 days after ACT treatment; therefore, this test is not suitable for treatment follow-up in pregnant women and can generate problems when using this test during intermittent preventive treatment (IPTp). DHFR-TS is less persistent than HRP2, making it a potentially interesting target for diagnosis. © 2012 Blackwell Publishing Ltd.

Urine Antigen Detection as an Aid to Diagnose Invasive Aspergillosis.

PubMed

Marr, Kieren A; Datta, Kausik; Mehta, Seema; Ostrander, Darin B; Rock, Michelle; Francis, Jesse; Feldmesser, Marta

2018-04-19

Establishing rapid diagnoses of invasive aspergillosis (IA) is priority, given poor outcomes of late therapy. Non-culture based tests that detect galactomannan and β-D glucan are available, but are technically cumbersome and rely on invasive sampling (blood or bronchoalveolar lavage). We optimized a lateral flow dipstick assay using the galactofuranose -specific monoclonal antibody (mAb476), which was previously shown to recognize urine antigens after Aspergillus fumigatus pulmonary infection in an`imals. Urine samples were obtained from a cohort of 78 subjects undergoing clinical evaluation for suspected invasive fungal infections, and stored frozen until testing. Urine was processed by centrifugation through desalting columns and exposed to dipsticks. Reviewers blinded to EORTC/MSG clinical diagnoses graded results. Western blots were performed on urines from two subjects to characterize mAb476-reactive antigens. Per-patient sensitivity and specificity for diagnosis of proven or probable IA in the overall cohort was 80% (95% CI: 61.4-92.3) and 92% (95% CI 74-99). In the sub-group with cancer, sensitivity was 89.5% (95% CI 66.7-98.7) and specificity was 90.9% (95% CI 58.7 - 99.8); amongst all others, sensitivity and specificity were 63.6 (95% CI 30.8 - 89.1) and 92.9 (66.1 - 99.8), respectively. Eliminating lung transplant recipients with airway disease increased sensitivity in the non-cancer cohort (85.7%, 95% CI 42.1-99.6%). Semi-quantitative urine assay results correlated with serum galactomannan indices. Western blots demonstrated mAb476-reactive antigens in urine from cases, ranging between 26 - 35kDa in size. Urine testing using mAb476 may be used as an aid to diagnose IA in high-risk patients.

A new paradigm for Aedes spp. surveillance using gravid ovipositing sticky trap and NS1 antigen test kit.

PubMed

Lau, Sai Ming; Chua, Tock H; Sulaiman, Wan-Yussof; Joanne, Sylvia; Lim, Yvonne Ai-Lian; Sekaran, Shamala Devi; Chinna, Karuthan; Venugopalan, Balan; Vythilingam, Indra

2017-03-21

Dengue remains a serious public health problem in Southeast Asia and has increased 37-fold in Malaysia compared to decades ago. New strategies are urgently needed for early detection and control of dengue epidemics. We conducted a two year study in a high human density dengue-endemic urban area in Selangor, where Gravid Ovipositing Sticky (GOS) traps were set up to capture adult Aedes spp. mosquitoes. All Aedes mosquitoes were tested using the NS1 dengue antigen test kit. All dengue cases from the study site notified to the State Health Department were recorded. Weekly microclimatic temperature, relative humidity (RH) and rainfall were monitored. Aedes aegypti was the predominant mosquito (95.6%) caught in GOS traps and 23% (43/187 pools of 5 mosquitoes each) were found to be positive for dengue using the NS1 antigen kit. Confirmed cases of dengue were observed with a lag of one week after positive Ae. aegypti were detected. Aedes aegypti density as analysed by distributed lag non-linear models, will increase lag of 2-3 weeks for temperature increase from 28 to 30 °C; and lag of three weeks for increased rainfall. Proactive strategy is needed for dengue vector surveillance programme. One method would be to use the GOS trap which is simple to setup, cost effective (below USD 1 per trap) and environmental friendly (i.e. use recyclable plastic materials) to capture Ae. aegypti followed by a rapid method of detecting of dengue virus using the NS1 dengue antigen kit. Control measures should be initiated when positive mosquitoes are detected.

Laboratory diagnosis of Chlamydia pneumoniae infections

PubMed Central

Peeling, Rosanna W

1995-01-01

Chlamydia pneumoniae is an important cause of respiratory illness. There is a need for accurate and rapid laboratory diagnostic methods that will lead to improved patient care, appropriate use of antimicrobial therapy and a better understanding of the epidemiology of this emerging pathogen. Culture is highly specific but is technically demanding, expensive, has a long turnaround time and its sensitivity is highly dependent on transport conditions. Antigen detection tests such as enzyme immunoassay and direct fluorescent antibody assay, and molecular detection methods such as the polymerase chain reaction assay, may provide a rapid diagnosis without the requirement for stringent transport conditions. The results of these tests should be interpreted with caution until more thorough evaluation is available. Serology remains the method of choice. The limitations of different serological methods for the laboratory diagnosis of C pneumoniae are discussed. PMID:22514397

Effect of specimen storage, antibiotics, and feminine hygiene products on the detection of group B Streptococcus by culture and the STREP B OIA test.

PubMed

Ostroff, R M; Steaffens, J W

1995-07-01

Agar culture from vaginal swabs is the routine method for diagnosis of maternal Group B Streptococcus (GBS) colonization. Swab specimens are often transported to a clinical laboratory for processing. In these studies, specimen transport was simulated by inoculating swabs with GBS and storing them at selected temperatures and with or without transport medium. The recovery of viable GBS was assessed by agar culture. GBS antigen was detected immunologically with an Optical ImmunoAssay (OIA) method. Swabs that were stored with transport medium harbored viable but rapidly declining numbers of GBS. In contrast, a strong OIA signal was maintained. Recovery of viable GBS organisms declined more quickly when swabs were stored in the absence of transport medium, whereas detection of GBS antigen remained consistent. Both methods were tested for interference from either antibiotics or feminine hygiene products. These compounds inhibited the detection of GBS by culture but had no detrimental effect on the OIA result.

Immunoblotting with human native antigen shows stage-related sensitivity in the serodiagnosis of hepatic cystic echinococcosis.

PubMed

Mariconti, Mara; Bazzocchi, Chiara; Tamarozzi, Francesca; Meroni, Valeria; Genco, Francesca; Maserati, Roberta; Brunetti, Enrico

2014-01-01

The diagnosis of hepatic cystic echinococcosis is based on ultrasonography and confirmed by serology. However, no biological marker of cyst viability is currently available implying years-long patient follow-up, which is not always feasible in endemic areas. We characterized the performance of an immunoblotting test based on human hydatid cyst fluid with particular regard to its ability to distinguish between cyst stages. Sera from patients with cysts in different stages showed distinctive band pattern recognition. Most importantly, the test discriminated in 80% of cases CE3a from CE3b transitional cysts, known to have different viability profiles. Interestingly, we observed a rapid change in band pattern recognition of sera from one patient at time points when his cyst passed from active to transitional to inactive stages. Further identification of different antigens expressed by different cyst stages will support the development of diagnostic tools that could early define cyst viability, to guide clinical decision making, and shorten patient follow-up.

21 CFR 866.6010 - Tumor-associated antigen immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Tumor-associated antigen immunological test system... SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Tumor Associated Antigen immunological Test Systems § 866.6010 Tumor-associated antigen immunological test system. (a) Identification. A...

Use of a three-band HRP2/pLDH combination rapid diagnostic test increases diagnostic specificity for falciparum malaria in Ugandan children.

PubMed

Hawkes, Michael; Conroy, Andrea L; Opoka, Robert O; Namasopo, Sophie; Liles, W Conrad; John, Chandy C; Kain, Kevin C

2014-02-01

Rapid diagnostic tests (RDTs) for malaria provide a practical alternative to light microscopy for malaria diagnosis in resource-limited settings. Three-band RDTs incorporating two parasite antigens may have enhanced diagnostic specificity, relative to two-band RDTs with a single parasite antigen (typically histidine-rich protein 2 [HRP2]). Phase 1: 2,000 children, two months to five years of age, admitted to a referral hospital in Jinja, Uganda, with acute febrile illness were enrolled. A WHO highly rated three-band RDT was compared to light microscopy of thick peripheral blood films read by local expert microscopists.Phase 2: the three-band RDT was used as a screening tool for inclusion of patients in a clinical trial, and subjects with three positive RDT bands were tested by microscopy using blood samples drawn in parallel. Discordant results were adjudicated by PCR. Phase 1: 1,648 children had both a RDT and peripheral blood smear performed. The specificity of a RDT with all three bands positive was 82% (95% CI: 79-85%) compared to 62% (95% CI: 59-66%) for HRP2 alone. The sensitivity was 88% (95% CI: 85-89%) and 94% (95% CI: 92-95%) for three-band positive RDT and HRP2 antigen, respectively. 119 patients (7.2%) had a positive HRP2 band, but negative parasite lactate dehydrogenase (pLHD) band and negative peripheral smear, and 72 (61%) of these had received pre-treatment with anti-malarials, suggesting a false positive HRP2 result (p = 0.002).Phase 2: the positive predictive value (PPV) of the three-band RDT was 94% (95% CI 89%-97%) using microscopy as the reference standard. However, microscopy-discordant results were shown to be positive for P. falciparum by PCR in all cases, suggesting that the PPV was in fact higher. The pLDH antigen on three-band RDTs, used in combination with HRP2, provides added diagnostic specificity for malaria parasitaemia and may be useful to distinguish acute infection from recently treated infection. In situations where diagnostic specificity is desirable (e.g., for selection of malaria-infected participants in clinical trials), a three-band RDT should be considered in a sub-Saharan African setting.

Evaluation of a rapid diagnostic test (CareStart™ Malaria HRP-2/pLDH (Pf/pan) Combo Test) for the diagnosis of malaria in a reference setting

PubMed Central

2010-01-01

Background Malaria Rapid Diagnostic Tests (RDTs) are widely used for diagnosing malaria. The present retrospective study evaluated the CareStart™ Malaria HRP-2/pLDH (Pf/pan) Combo Test targeting the Plasmodium falciparum specific antigen histidine-rich protein (HRP-2) and the pan-Plasmodium antigen lactate dehydrogenase (pLDH) in a reference setting. Methods The CareStart™ Malaria HRP-2/pLDH (Pf/pan) Combo Test was evaluated on a collection of samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included were P. falciparum (n = 320), Plasmodium vivax (n = 76), Plasmodium ovale (n = 76), Plasmodium malariae (n = 23) and Plasmodium negative samples (n = 95). Results Overall sensitivity for the detection of P. falciparum was 88.8%, increasing to 94.3% and 99.3% at parasite densities above 100 and 1,000/μl respectively. For P. vivax, P. ovale and P. malariae, overall sensitivities were 77.6%, 18.4% and 30.4% respectively. For P. vivax sensitivity reached 90.2% for parasite densities above 500/μl. Incorrect species identification occurred in 11/495 samples (2.2%), including 8/320 (2.5%) P. falciparum samples which generated only the pan-pLDH line. For P. falciparum samples, 205/284 (72.2%) HRP-2 test lines had strong or medium line intensities, while for all species the pan-pLDH lines were less intense, especially in the case of P. ovale. Agreement between observers was excellent (kappa values > 0.81 for positive and negative readings) and test results were reproducible. The test was easy to perform with good clearing of the background. Conclusion The CareStart™ Malaria HRP-2/pLDH (Pf/pan) Combo Test performed well for the detection of P. falciparum and P. vivax, but sensitivities for P. ovale and P. malariae were poor. PMID:20565816

Evaluation of a rapid diagnostic test (CareStart Malaria HRP-2/pLDH (Pf/pan) Combo Test) for the diagnosis of malaria in a reference setting.

PubMed

Maltha, Jessica; Gillet, Philippe; Bottieau, Emmanuel; Cnops, Lieselotte; van Esbroeck, Marjan; Jacobs, Jan

2010-06-18

Malaria Rapid Diagnostic Tests (RDTs) are widely used for diagnosing malaria. The present retrospective study evaluated the CareStart Malaria HRP-2/pLDH (Pf/pan) Combo Test targeting the Plasmodium falciparum specific antigen histidine-rich protein (HRP-2) and the pan-Plasmodium antigen lactate dehydrogenase (pLDH) in a reference setting. The CareStart Malaria HRP-2/pLDH (Pf/pan) Combo Test was evaluated on a collection of samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included were P. falciparum (n = 320), Plasmodium vivax (n = 76), Plasmodium ovale (n = 76), Plasmodium malariae (n = 23) and Plasmodium negative samples (n = 95). Overall sensitivity for the detection of P. falciparum was 88.8%, increasing to 94.3% and 99.3% at parasite densities above 100 and 1,000/microl respectively. For P. vivax, P. ovale and P. malariae, overall sensitivities were 77.6%, 18.4% and 30.4% respectively. For P. vivax sensitivity reached 90.2% for parasite densities above 500/microl. Incorrect species identification occurred in 11/495 samples (2.2%), including 8/320 (2.5%) P. falciparum samples which generated only the pan-pLDH line. For P. falciparum samples, 205/284 (72.2%) HRP-2 test lines had strong or medium line intensities, while for all species the pan-pLDH lines were less intense, especially in the case of P. ovale. Agreement between observers was excellent (kappa values > 0.81 for positive and negative readings) and test results were reproducible. The test was easy to perform with good clearing of the background. The CareStart Malaria HRP-2/pLDH (Pf/pan) Combo Test performed well for the detection of P. falciparum and P. vivax, but sensitivities for P. ovale and P. malariae were poor.

Improving the Specificity of Plasmodium falciparum Malaria Diagnosis in High-Transmission Settings with a Two-Step Rapid Diagnostic Test and Microscopy Algorithm.

PubMed

Murungi, Moses; Fulton, Travis; Reyes, Raquel; Matte, Michael; Ntaro, Moses; Mulogo, Edgar; Nyehangane, Dan; Juliano, Jonathan J; Siedner, Mark J; Boum, Yap; Boyce, Ross M

2017-05-01

Poor specificity may negatively impact rapid diagnostic test (RDT)-based diagnostic strategies for malaria. We performed real-time PCR on a subset of subjects who had undergone diagnostic testing with a multiple-antigen (histidine-rich protein 2 and pan -lactate dehydrogenase pLDH [HRP2/pLDH]) RDT and microscopy. We determined the sensitivity and specificity of the RDT in comparison to results of PCR for the detection of Plasmodium falciparum malaria. We developed and evaluated a two-step algorithm utilizing the multiple-antigen RDT to screen patients, followed by confirmatory microscopy for those individuals with HRP2-positive (HRP2 + )/pLDH-negative (pLDH - ) results. In total, dried blood spots (DBS) were collected from 276 individuals. There were 124 (44.9%) individuals with an HRP2 + /pLDH + result, 94 (34.1%) with an HRP2 + /pLDH - result, and 58 (21%) with a negative RDT result. The sensitivity and specificity of the RDT compared to results with real-time PCR were 99.4% (95% confidence interval [CI], 95.9 to 100.0%) and 46.7% (95% CI, 37.7 to 55.9%), respectively. Of the 94 HRP2 + /pLDH - results, only 32 (34.0%) and 35 (37.2%) were positive by microscopy and PCR, respectively. The sensitivity and specificity of the two-step algorithm compared to results with real-time PCR were 95.5% (95% CI, 90.5 to 98.0%) and 91.0% (95% CI, 84.1 to 95.2), respectively. HRP2 antigen bands demonstrated poor specificity for the diagnosis of malaria compared to that of real-time PCR in a high-transmission setting. The most likely explanation for this finding is the persistence of HRP2 antigenemia following treatment of an acute infection. The two-step diagnostic algorithm utilizing microscopy as a confirmatory test for indeterminate HRP2 + /pLDH - results showed significantly improved specificity with little loss of sensitivity in a high-transmission setting. Copyright © 2017 American Society for Microbiology.

[Sensitivity of three inmunocromathographic tests in faeces samples for Campylobacter and Salmonella detection in comparison to culture].

PubMed

Liébana-Martos, Ma del Carmen; Gutierrez, José; Riazzo, Cristina; Navarro, José Ma

2014-06-01

Introduction: Campylobacter sp. and Salmonella enterica are two of the main organisms causing gastroenteritis in our environment. Immunochromatographic tests for antigen detection performed directly on stool samples for its simplicity and rapid results may make them useful diagnostic elements in the context of primary care. During October 2012 we selected all feces in which enteropathogenic bacteria are isolated from those received for stool culture in the laboratory of Microbiology of the University Hospital Virgen de las Nieves of Granada. After standard management of faeces samples and isolation of any enteropathogen, the commercial kits: Campy Leti, Ridaquick Campylobacterscreen and Salmonella Leti were tested for simultaneous research of Campylobacter and Salmonella antigens. Sensitivity and specificity were determined. Two hundred and thirty five stool samples were received in which 8 Salmonella enterica (7 B serogroup and 1 D serogroup), 7 Campylobacter jejuni, 4 Aeromonas hydrophila and 1 Yersinia enterocolitica were isolated. Campy Leti, Ridaquick Campylobacterscreen and Salmonella Leti presented a sensitivity of 100%, 100% and 75%, respectively. Specificities corresponded to 46%, 69% and 100%, respectively. Immunocromatographic tests can be useful for a first screening of enteropathogen in primary care.

Laboratory diagnostics of malaria

NASA Astrophysics Data System (ADS)

Siahaan, L.

2018-03-01

Even now, malaria treatment should only be administered after laboratory confirmation. There are several principal methods for diagnosing malaria. All these methods have their disadvantages.Presumptive treatment of malaria is widely practiced where laboratory tests are not readily available. Microscopy of Giemsa-stained thick and thin blood films remains the gold standard for the diagnosis of malaria infection. The technique of slide preparation, staining and reading are well known and standardized, and so is the estimate of the parasite density and parasite stages. Microscopy is not always available or feasible at primary health services in limited resource settings due to cost, lack of skilled manpower, accessories and reagents required. Rapid diagnostic tests (RDTs) are potential tools for parasite-based diagnosis since the tests are accurate in detecting malaria infections and are easy to use. The test is based on the capture of parasite antigen that released from parasitized red blood cells using monoclonal antibodies prepared against malaria antigen target. Polymerase Chain Reaction (PCR), depend on DNA amplification approaches and have higher sensitivity than microscopy. PCR it is not widely used due to the lack of a standardized methodology, high costs, and the need for highly-trained staff.

Serological Evidence of Lyssavirus Infection among Bats in Nagaland, a North-Eastern State in India.

PubMed

Mani, R S; Dovih, D P; Ashwini, M A; Chattopadhyay, B; Harsha, P K; Garg, K M; Sudarshan, S; Puttaswamaiah, R; Ramakrishnan, U; Madhusudana, S N

2017-06-01

Bats are known to be reservoirs of several medically important viruses including lyssaviruses. However, no systematic surveillance for bat rabies has been carried out in India, a canine rabies endemic country with a high burden of human rabies. Surveillance for rabies virus (RABV) infection in bats was therefore carried out in Nagaland, a north-eastern state in India at sites with intense human-bat interfaces during traditional bat harvests. Brain tissues and sera from bats were tested for evidence of infection due to RABV. Brain tissues were subjected to the fluorescent antibody test for detection of viral antigen and real-time reverse transcriptase PCR for presence of viral RNA. Bat sera were tested for the presence of rabies neutralizing antibodies by the rapid fluorescent focus inhibition test. None of the bat brains tested (n = 164) were positive for viral antigen or viral RNA. However, rabies neutralizing antibodies were detected in 4/78 (5·1%) bat sera tested, suggesting prior exposure to RABV or related lyssaviruses. The serological evidence of lyssaviral infection in Indian bats may have important implications in disease transmission and rabies control measures, and warrant extensive bat surveillance to better define the prevalence of lyssaviral infection in bats.

Peritonsillar abscess and cellulitis and their relation to a positive antigen detection test for streptococcal infection.

PubMed

Risberg, Stefan; Engfeldt, Peter; Hugosson, Svante

2010-10-01

The microbiological cause of peritonsillar abscess and the role of group A β-haemolytic Streptococcus (GAS) are unclear. We performed a retrospective study at the ear, nose and throat clinic (ENT) of Orebro University Hospital, Sweden, and included 376 events of peritonsillitis between 2002 and 2004. We determined if the patients had visited a primary healthcare centre (PHCC) within 30 days prior to inclusion. The results of the rapid antigen detection test for GAS (Strep A) taken at the PHCC were compared with the occurrence of peritonsillar abscess (PTA) and peritonsillar cellulitis (PTC). A Strep A test was performed in 61% (229/376) of the events studied. Strep A was positive in 22% of PTA events and in 35% of PTC events (p = 0.036). Of 48,000 Strep A tests taken in primary healthcare, mainly for sore throat, 22% were positive. We examined the relationship between age, the incidence of PTA, and positive Strep A tests. We also determined if there was a monthly correlation between number of positive Strep A tests and number of PTA events. We found no significant correlations. In conclusion, our findings indicate that GAS does not play a major role in the development of PTA/PTC.

[Use of Rapid Antigen Detection Test (RADT) among general practitioner teachers at the Paris Descartes University: 2005-2007].

PubMed

Cornaglia, C; Robinet, J; Partouche, H

2009-06-01

The distribution of the Rapid Antigen Detection Test (RADT) and the National Health Insurance's information campaign should efficiently reduce the unjustified use antibiotic. However, a preliminary survey among GP trainers at the Paris Descartes University indicated that the RADT was seldom used. This study had for aim to describe the RADT use trend among trainers since 2005 and the main obstacles to its widespread use, and to assess the Mac Isaac score use and antibiotic prescriptions. Between February and May 2007, a survey was carried out among 66 GPs who were required to report their first ten patients over three years of age presenting with pharyngitis. RADT use and antibiotic prescriptions were compared with those of the 2005 survey. RADT use had decreased (52.5% [48.2-56.8] versus 57.5% [52.1-68.8], p<0.05). GPs did not use the RADT because they considered it "useless in decision making". Clinical findings were sufficient in most cases. The Mac Isaac score was not widely used by GPs (28.3%) and antibiotic prescription had increased except for macrolides which had decreased (10% vs 15%). Among patients with a negative RADT, 11.9% (vs 10.5% en 2005, p<0.001) were prescribed antibiotics. The RADT use decreased in two years among GP trainers. GPs still prescribe treatment according to clinical findings, most without using diagnostic tools.

Standardization and validation of a cytometric bead assay to assess antibodies to multiple Plasmodium falciparum recombinant antigens

PubMed Central

2012-01-01

Background Multiplex cytometric bead assay (CBA) have a number of advantages over ELISA for antibody testing, but little information is available on standardization and validation of antibody CBA to multiple Plasmodium falciparum antigens. The present study was set to determine optimal parameters for multiplex testing of antibodies to P. falciparum antigens, and to compare results of multiplex CBA to ELISA. Methods Antibodies to ten recombinant P. falciparum antigens were measured by CBA and ELISA in samples from 30 individuals from a malaria endemic area of Kenya and compared to known positive and negative control plasma samples. Optimal antigen amounts, monoplex vs multiplex testing, plasma dilution, optimal buffer, number of beads required were assessed for CBA testing, and results from CBA vs. ELISA testing were compared. Results Optimal amounts for CBA antibody testing differed according to antigen. Results for monoplex CBA testing correlated strongly with multiplex testing for all antigens (r = 0.88-0.99, P values from <0.0001 - 0.004), and antibodies to variants of the same antigen were accurately distinguished within a multiplex reaction. Plasma dilutions of 1:100 or 1:200 were optimal for all antigens for CBA testing. Plasma diluted in a buffer containing 0.05% sodium azide, 0.5% polyvinylalcohol, and 0.8% polyvinylpyrrolidone had the lowest background activity. CBA median fluorescence intensity (MFI) values with 1,000 antigen-conjugated beads/well did not differ significantly from MFI with 5,000 beads/well. CBA and ELISA results correlated well for all antigens except apical membrane antigen-1 (AMA-1). CBA testing produced a greater range of values in samples from malaria endemic areas and less background reactivity for blank samples than ELISA. Conclusion With optimization, CBA may be the preferred method of testing for antibodies to P. falciparum antigens, as CBA can test for antibodies to multiple recombinant antigens from a single plasma sample and produces a greater range of values in positive samples and lower background readings for blank samples than ELISA. PMID:23259607

Development of a rapid immunoassay system: Luminescent detection of antigen-associated antibody-luciferase in the presence of a dye that absorbs light from free antibody-luciferase.

PubMed

Mori, Akihiro; Ojima-Kato, Teruyo; Fuchi, Shingo; Kaiya, Shinichi; Kojima, Takaaki; Nakano, Hideo

2017-12-01

In this report, we developed a rapid immunoassay system, designated the bioluminescent interference gathering optical (BINGO) assay, which required no time-consuming washing steps for removal of unbound antibodies. This system employed a luciferase (Luc)-conjugated antibody (LucAb) and a dye that absorbed light from the LucAb. The antigen-associated LucAb was localized by transfer of an antigen to the detector-side of a chamber where a detector photomultiplier tube (PMT) was installed. In contrast, the free LucAb was distributed throughout the solution, and the light emitted by the free LucAb was absorbed by the dye. Therefore, only light from LucAb associated with antigen could be detected by the PMT. The new system could be used to rapidly detect the amount of antigen-antibody-Luc complex by collecting steps, such as centrifugation or magnetic collection of antibody-coated magnetic beads. Proof-of-principle experiments were performed using a model system with streptavidin beads and biotinylated Luc. The feasibility of the system was demonstrated using magnetic beads coated with anti-Escherichia coli O157 antibody, enabling detection of 4 × 10 3  cells in only 15 min. Thus, this system may have applications in a variety of biomedical fields. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Diagnostic performance of a rapid in-clinic test for the detection of Canine Parvovirus under different storage conditions and vaccination status.

PubMed

Kantere, Maria C; Athanasiou, Labrini V; Spyrou, Vassiliki; Kyriakis, Constantinos S; Kontos, Vassilios; Chatzopoulos, Dimitrios C; Tsokana, Constantina N; Billinis, Charalambos

2015-04-01

Canine parvovirus (CPV) is one of the most common causes of acute haemorrhagic enteritis in young dogs, while clinical diagnosis is often indecisive. The aim of our study was to evaluate the diagnostic accuracy of an in-clinic rapid test in the detection of CPV infection in dogs. To this end, we compared the Rapid Diagnostic Kit of Canine Parvovirus, Coronavirus and Rotavirus antigen (Quicking(®)) to PCR, which is considered as the most reliable diagnostic method. A total of 78 duplicated faecal samples were collected from diarrhoeic dogs. Vaccination history within a month prior to the onset of diarrhoea was reported for 12 of the sampled dogs. The rapid diagnostic test was performed in 23 of the faecal samples directly, while the rest were placed into a sterile cotton tipped swab suitable for collection and transportation of viruses (Sigma Σ-VCM(®)) and stored at -20 °C. The sensitivity of the Quicking rapid diagnostic test compared to PCR in the total number of samples, in samples from non-vaccinated dogs and in samples tested directly after collection were 22.22% (95% CI: 13.27-33.57%), 26.67% (95% CI: 16.08-39.66%) and 76.47% (95% CI: 50.10-93.04%) respectively, while the specificity of the test was 100% in any case. In conclusion, negative results do not exclude parvoenteritis from the differential diagnosis, especially in dogs with early vaccination history, but a positive result almost certainly indicates CPV infection. An improved sensitivity may be expected when the test is performed immediately. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of a rapid antibody test for point-of-care diagnosis of animal African trypanosomosis.

PubMed

Boulangé, Alain; Pillay, Davita; Chevtzoff, Cyrille; Biteau, Nicolas; Comé de Graça, Vanessa; Rempeters, Leonie; Theodoridis, Dimitrios; Baltz, Théo

2017-01-15

Trypanosoma congolense and T. vivax are the main causative agents of animal African trypanosomosis (AAT), a disease which hinders livestock production throughout sub-Saharan Africa and in some parts of South America. Although two trypanocidal drugs are currently available, the level of treatment is low due to the difficulty in diagnosing the disease in the field. The major clinical signs of AAT such as anaemia, weight loss, and infertility, are common to several other endemic livestock diseases. Current diagnostic methods, based on the visualization of the parasite in the blood, or on the detection of its DNA or the antibodies it triggers in the host, are not suitable for direct use in the field as they require specialized equipment and personnel. Thus, we developed a quick-format diagnostic test (15min) based on the recombinant TcoCB and TvGM6 antigens for detection of T. congolense and T. vivax, respectively, aimed at providing farmers and veterinarians in the field with the means to conduct a quick diagnosis. The specificity and sensitivity of the test were evaluated using sera from experimentally infected cattle, and fresh blood when possible. The prototype, which includes both antigens, shows a specificity of 95.9 (95% C.I., 90.4%-100%) and a sensitivity of 92.0% (95% C.I., 85.9%-98.1%) for T. congolense and 98.2% (95% C.I., 94.7%-100%) for T. vivax. The high levels of sensitivity and specificity of this rapid test, the possibility of using directly whole blood, and the ease of interpreting the result, all contribute to make of this test a valuable candidate to contribute to the control of AAT in the field. However, further tests with more representative, numerous and fresh reference samples are necessary in order to compare this test with the ELISA, the current gold standard serological test for trypanosomosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Isolation of the most immunoreactive antigenes of echinococcus granulosus from sheep hydatid fluid.

PubMed

Pozzuoli, R; Piantelli, M; Perucci, C; Arru, E; Musiani, P

1975-11-01

This paper describes a simplified procedure for obtaining purified Echinococcus granulosus antigens from sheep hydatid fluid by using affinity chromatography on concanavalin A-Sepharose. The presence of two "major" antigens (4 and 5) was confirmed. Antigen 5 was isolated by preparative polyacrylamide gel electrophoresis. Antigen 4, eluted by diffusion from the gel, was seen to be "contaminated" by antigen 5 and was isolated by using anti-5 Sepharose-linked serum. These two major antigens were then tested separately against the sera of hydatidosis patients by using very simple immunolgic tests. The best results were obtained in passive hemagglutination with antigen 4. Antigen 4 is the most immunoreactive parasitic antigen; antibodies against it were found in the sera of all hydatidosis patients showing positive reaction. Apart from the direct use of this antigen in serologic tests, it appears possible to standarize the most frequently used and commerically available antigenic materials by titrating this component.

Misdiagnosis of HIV infection during a South African community-based survey: implications for rapid HIV testing

PubMed Central

Kufa, Tendesayi; Kharsany, Ayesha BM; Cawood, Cherie; Khanyile, David; Lewis, Lara; Grobler, Anneke; Chipeta, Zawadi; Bere, Alfred; Glenshaw, Mary; Puren, Adrian

2017-01-01

Abstract Introduction: We describe the overall accuracy and performance of a serial rapid HIV testing algorithm used in community-based HIV testing in the context of a population-based household survey conducted in two sub-districts of uMgungundlovu district, KwaZulu-Natal, South Africa, against reference fourth-generation HIV-1/2 antibody and p24 antigen combination immunoassays. We discuss implications of the findings on rapid HIV testing programmes. Methods: Cross-sectional design: Following enrolment into the survey, questionnaires were administered to eligible and consenting participants in order to obtain demographic and HIV-related data. Peripheral blood samples were collected for HIV-related testing. Participants were offered community-based HIV testing in the home by trained field workers using a serial algorithm with two rapid diagnostic tests (RDTs) in series. In the laboratory, reference HIV testing was conducted using two fourth-generation immunoassays with all positives in the confirmatory test considered true positives. Accuracy, sensitivity, specificity, positive predictive value, negative predictive value and false-positive and false-negative rates were determined. Results: Of 10,236 individuals enrolled in the survey, 3740 were tested in the home (median age 24 years (interquartile range 19–31 years), 42.1% males and HIV positivity on RDT algorithm 8.0%). From those tested, 3729 (99.7%) had a definitive RDT result as well as a laboratory immunoassay result. The overall accuracy of the RDT when compared to the fourth-generation immunoassays was 98.8% (95% confidence interval (CI) 98.5–99.2). The sensitivity, specificity, positive predictive value and negative predictive value were 91.1% (95% CI 87.5–93.7), 99.9% (95% CI 99.8–100), 99.3% (95% CI 97.4–99.8) and 99.1% (95% CI 98.8–99.4) respectively. The false-positive and false-negative rates were 0.06% (95% CI 0.01–0.24) and 8.9% (95% CI 6.3–12.53). Compared to true positives, false negatives were more likely to be recently infected on limited antigen avidity assay and to report antiretroviral therapy (ART) use. Conclusions: The overall accuracy of the RDT algorithm was high. However, there were few false positives, and the sensitivity was lower than expected with high false negatives, despite implementation of quality assurance measures. False negatives were associated with recent (early) infection and ART exposure. The RDT algorithm was able to correctly identify the majority of HIV infections in community-based HIV testing. Messaging on the potential for false positives and false negatives should be included in these programmes. PMID:28872274

Multiplex real-time PCR for identification of canine parvovirus antigenic types.

PubMed

Kaur, Gurpreet; Chandra, Mudit; Dwivedi, P N; Narang, Deepti

2016-07-01

Canine parvovirus (CPV) is an important disease causing gastroenteritis and/or haemorrhagic gastroenteritis in dogs. There are four antigenic types of CPV reported worldwide viz. CPV 2, CPV 2a, CPV 2b and CPV 2c. The diagnosis of CPV with the identification of the antigen type responsible remains problematic. In the present study, identification as well as antigenic typing of CPV was done using a de novo multiplex real time PCR to combat the problem of antigenic type identification. From the study it could be concluded that the here developed multiplex real time PCR assay could be used for rapid detection of CPV as well as typing of its three antigenic types. Copyright © 2016 Elsevier B.V. All rights reserved.

Novel Treponema pallidum Recombinant Antigens for Syphilis Diagnostics: Current Status and Future Prospects

PubMed Central

Kubanov, Aleksey; Runina, Anastassia

2017-01-01

The recombinant protein technology considerably promoted the development of rapid and accurate treponema-specific laboratory diagnostics of syphilis infection. For the last ten years, the immunodominant recombinant inner membrane lipoproteins are proved to be sensitive and specific antigens for syphilis screening. However, the development of an enlarged T. pallidum antigen panel for diagnostics of early and late syphilis and differentiation of syphilis stages or cured syphilis remains as actual goal of multidisciplinary expertise. Current review revealed novel recombinant antigens: surface-exposed proteins, adhesins, and periplasmic and flagellar proteins, which are promising candidates for the improved syphilis serological diagnostics. The opportunities and limitations of diagnostic usage of these antigens are discussed and the criteria for selection of optimal antigens panel summarized. PMID:28523273

Performance of ARCHITECT HCV core antigen test with specimens from US plasma donors and injecting drug users.

PubMed

Mixson-Hayden, Tonya; Dawson, George J; Teshale, Eyasu; Le, Thao; Cheng, Kevin; Drobeniuc, Jan; Ward, John; Kamili, Saleem

2015-05-01

Hepatitis C virus (HCV) core antigen is a serological marker of current HCV infection. The aim of this study was mainly to evaluate the performance characteristics of the ARCHITECT HCV core antigen assay with specimens from US plasma donors and injecting drug users. A total of 551 serum and plasma samples with known anti-HCV and HCV RNA status were tested for HCV core antigen using the Abbott ARCHITECT HCV core antigen test. HCV core antigen was detectable in 100% of US plasma donor samples collected during the pre-seroconversion phase of infection (anti-HCV negative/HCV RNA positive). Overall sensitivity of the HCV core antigen assay was 88.9-94.3% in samples collected after seroconversion. The correlation between HCV core antigen and HCV RNA titers was 0.959. HCV core antigen testing may be reliably used to identify current HCV infection. Published by Elsevier B.V.

Implementation of broad screening with Ebola rapid diagnostic tests in Forécariah, Guinea

PubMed Central

Nebie, Yacouba K.; Koivogui, Lamine; Abiola, Nadine; Vansteelandt, Amanda; Worrel, Mary C.; Shang, Judith; Murphy, Louise B.; Fitter, David L.; Marston, Barbara J.; Martel, Lise

2017-01-01

Background Laboratory-enhanced surveillance is critical for rapidly detecting the potential re-emergence of Ebola virus disease. Rapid diagnostic tests (RDT) for Ebola antigens could expand diagnostic capacity for Ebola virus disease. Objectives The Guinean National Coordination for Ebola Response conducted a pilot implementation to determine the feasibility of broad screening of patients and corpses with the OraQuick® Ebola RDT. Methods The implementation team developed protocols and trained healthcare workers to screen patients and corpses in Forécariah prefecture, Guinea, from 15 October to 30 November 2015. Data collected included number of consultations, number of fevers reported or measured, number of tests performed for patients or corpses and results of confirmatory RT-PCR testing. Data on malaria RDT results were collected for comparison. Feedback from Ebola RDT users was collected informally during supervision visits and forums. Results There were 3738 consultations at the 15 selected healthcare facilities; 74.6% of consultations were for febrile illness. Among 2787 eligible febrile patients, 2633 were tested for malaria and 1628 OraQuick® Ebola RDTs were performed. A total of 322 OraQuick® Ebola RDTs were conducted on corpses. All Ebola tests on eligible patients were negative. Conclusions Access to Ebola testing was expanded by the implementation of RDTs in an emergency situation. Feedback from Ebola RDT users and lessons learned will contribute to improving quality for RDT expansion. PMID:28879148

Six rapid tests for direct detection of Clostridium difficile and its toxins in fecal samples compared with the fibroblast cytotoxicity assay.

PubMed

Turgeon, David K; Novicki, Thomas J; Quick, John; Carlson, LaDonna; Miller, Pat; Ulness, Bruce; Cent, Anne; Ashley, Rhoda; Larson, Ann; Coyle, Marie; Limaye, Ajit P; Cookson, Brad T; Fritsche, Thomas R

2003-02-01

Clostridium difficile is one of the most frequent causes of nosocomial gastrointestinal disease. Risk factors include prior antibiotic therapy, bowel surgery, and the immunocompromised state. Direct fecal analysis for C. difficile toxin B by tissue culture cytotoxin B assay (CBA), while only 60 to 85% sensitive overall, is a common laboratory method. We have used 1,003 consecutive, nonduplicate fecal samples to compare six commercially available immunoassays (IA) for C. difficile detection with CBA: Prima System Clostridium difficile Tox A and VIDAS Clostridium difficile Tox A II, which detect C. difficile toxin A; Premier Cytoclone A/B and Techlab Clostridium difficile Tox A/B, which detect toxins A and B; and ImmunoCard Clostridium difficile and Triage Micro C. difficile panels, which detect toxin A and a species-specific antigen. For all tests, Triage antigen was most sensitive (89.1%; negative predictive value [NPV] = 98.7%) while ImmunoCard was most specific (99.7%; positive predictive value [PPV] = 95.0%). For toxin tests only, Prima System had the highest sensitivity (82.2%; NPV = 98.0%) while ImmunoCard had the highest specificity (99.7%; PPV = 95.0%). Hematopoietic stem cell transplant (HSCT) patients contributed 44.7% of all samples tested, and no significant differences in sensitivity or specificity were noted between HSCT and non-HSCT patients. IAs, while not as sensitive as direct fecal CBA, produce reasonable predictive values, especially when both antigen and toxin are detected. They also offer significant advantages over CBA in terms of turnaround time and ease of use.

Microfluidic immunosensor with integrated liquid core waveguides for sensitive Mie scattering detection of avian influenza antigens in a real biological matrix.

PubMed

Heinze, Brian C; Gamboa, Jessica R; Kim, Keesung; Song, Jae-Young; Yoon, Jeong-Yeol

2010-11-01

This work presents the use of integrated, liquid core, optical waveguides for measuring immunoagglutination-induced light scattering in a microfluidic device, towards rapid and sensitive detection of avian influenza (AI) viral antigens in a real biological matrix (chicken feces). Mie scattering simulations were performed and tested to optimize the scattering efficiency of the device through proper scatter angle waveguide geometry. The detection limit is demonstrated to be 1 pg mL(-1) in both clean buffer and real biological matrix. This low detection limit is made possible through on-chip diffusional mixing of AI target antigens and high acid content microparticle assay reagents, coupled with real-time monitoring of immunoagglutination-induced forward Mie scattering via high refractive index liquid core optical waveguides in close proximity (100 μm) to the sample chamber. The detection time for the assay is <2 min. This device could easily be modified to detect trace levels of any biological molecules that antibodies are available for, moving towards a robust platform for point-of-care disease diagnostics.

Multiplex detection of pathogen biomarkers in human blood, serum, and saliva using silicon photonic microring resonators

NASA Astrophysics Data System (ADS)

Estrada, I. A.; Burlingame, R. W.; Wang, A. P.; Chawla, K.; Grove, T.; Wang, J.; Southern, S. O.; Iqbal, M.; Gunn, L. C.; Gleeson, M. A.

2015-05-01

Genalyte has developed a multiplex silicon photonic chip diagnostics platform (MaverickTM) for rapid detection of up to 32 biological analytes from a drop of sample in just 10 to 20 minutes. The chips are manufactured with waveguides adjacent to ring resonators, and probed with a continuously variable wavelength laser. A shift in the resonant wavelength as mass binds above the ring resonators is measured and is directly proportional to the amount of bound macromolecules. We present here the ability to multiplex the detection of hemorrhagic fever antigens in whole blood, serum, and saliva in a 16 minute assay. Our proof of concept testing of a multiplex antigencapture chip has the ability to detect Zaire Ebola (ZEBOV) recombinant soluble glycoprotein (rsGP), Marburg virus (MARV) Angola recombinant glycoprotein (rGP) and dengue nonstructural protein I (NS1). In parallel, detection of 2 malaria antigens has proven successful, but has yet to be incorporated into multiplex with the others. Each assay performs with sensitivity ranging from 1.6 ng/ml to 39 ng/ml depending on the antigen detected, and with minimal cross-reactivity.

Disruption of SMIM1 causes the Vel− blood type

PubMed Central

Ballif, Bryan A; Helias, Virginie; Peyrard, Thierry; Menanteau, Cécile; Saison, Carole; Lucien, Nicole; Bourgouin, Sébastien; Le Gall, Maude; Cartron, Jean-Pierre; Arnaud, Lionel

2013-01-01

Here, we report the biochemical and genetic basis of the Vel blood group antigen, which has been a vexing mystery for decades, especially as anti-Vel regularly causes severe haemolytic transfusion reactions. The protein carrying the Vel blood group antigen was biochemically purified from red blood cell membranes. Mass spectrometry-based de novo peptide sequencing identified this protein to be small integral membrane protein 1 (SMIM1), a previously uncharacterized single-pass membrane protein. Expression of SMIM1 cDNA in Vel− cultured cells generated anti-Vel cell surface reactivity, confirming that SMIM1 encoded the Vel blood group antigen. A cohort of 70 Vel− individuals was found to be uniformly homozygous for a 17 nucleotide deletion in the coding sequence of SMIM1. The genetic homogeneity of the Vel− blood type, likely having a common origin, facilitated the development of two highly specific DNA-based tests for rapid Vel genotyping, which can be easily integrated into blood group genotyping platforms. These results answer a 60-year-old riddle and provide tools of immediate assistance to all clinicians involved in the care of Vel− patients. PMID:23505126

[Extramembranous glomerulonephritis of acquired syphilis in a patient recently infected by the hepatitis B virus. Demonstration of the treponemal antigen in the kidney by indirect immunofluorescence].

PubMed

Schillinger, F; Montagnac, R; Goclowski, C; Dine, G; Alessandri, E; Hopfner, C; Birembaut, P

1983-01-22

A 38 years old male homosexual with active secondary syphilis presented with pure nephrotic syndrome while HBs and HBe tests were positive without clinical hepatitis. He had circulating immune complexes, IgG--IgM cryoglobulinemia and high IgA, IgM and IgE levels; the C3 and C4 complement constituents were normal. Examination of renal biopsy sections under light, fluorescent and electronic microscopy showed stage I membranous glomerulonephritis the syphilitic origin of which was confirmed by indirect immunofluorescence and by rapid cure under penicillin treatment. This case calls for the following comments: (1) glomerular deposits are extramembranous rather than subendothelial in syphilitic nephrosis, a disease now classified among circulating immune complexes diseases; (2) in the kidney, the treponema antigen can be demonstrated by indirect immunofluorescence and the anti-treponema antibody, by elution; (3) the outcome of the nephrotic syndrome is always favourable, either spontaneously or after penicillin treatment; (4) syphilis and HBs antigens are frequently associated, particularly in homosexual patients; one should be looked for when the other is discovered.

Joining the in vitro immunization of alpaca lymphocytes and phage display: rapid and cost effective pipeline for sdAb synthesis.

PubMed

Comor, Lubos; Dolinska, Saskia; Bhide, Katarina; Pulzova, Lucia; Jiménez-Munguía, Irene; Bencurova, Elena; Flachbartova, Zuzana; Potocnakova, Lenka; Kanova, Evelina; Bhide, Mangesh

2017-01-23

Camelids possess unique functional heavy chain antibodies, which can be produced and modified in vitro as a single domain antibody (sdAb or nanobody) with full antigen binding ability. Production of sdAb in conventional manner requires active immunization of Camelidae animal, which is laborious, time consuming, costly and in many cases not feasible (e.g. in case of highly toxic or infectious antigens). In this study, we describe an alternative pipeline that includes in vitro stimulation of naïve alpaca B-lymphocytes by antigen of interest (in this case endothelial cell binding domain of OspA of Borrelia) in the presence of recombinant alpaca interleukins 2 and 4, construction of sdAb phage library, selection of antigen specific sdAb expressed on phages (biopanning) and confirmation of binding ability of sdAb to the antigen. By joining the in vitro immunization and the phage display ten unique phage clones carrying sdAb were selected. Out of ten, seven sdAb showed strong antigen binding ability in phage ELISA. Furthermore, two soluble forms of sdAb were produced and their differential antigen binding affinity was measured with bio-layer interferometry. A proposed pipeline has potential to reduce the cost substantially required for maintenance of camelid herd for active immunization. Furthermore, in vitro immunization can be achieved within a week to enrich mRNA copies encoding antigen-specific sdAbs in B cell. This rapid and cost effective pipeline can help researchers to develop efficiently sdAb for diagnostic and therapeutic purposes.

CdSe/ZnS Quantum Dot-Labeled Lateral Flow Strips for Rapid and Quantitative Detection of Gastric Cancer Carbohydrate Antigen 72-4

NASA Astrophysics Data System (ADS)

Yan, Xinyu; Wang, Kan; Lu, Wenting; Qin, Weijian; Cui, Daxiang; He, Jinghua

2016-03-01

Carbohydrate antigen 72-4 (CA72-4) is an important biomarker associated closely with diagnosis and prognosis of early gastric cancer. How to realize quick, sensitive, specific, and quantitative detection of CA72-4 in clinical specimens has become a great requirement. Herein, we reported a CdSe/ZnS quantum dot-labeled lateral flow test strip combined with a charge-coupled device (CCD)-based reader was developed for rapid, sensitive, and quantitative detection of CA72-4. Two mouse monoclonal antibodies (mAbs) against CA72-4 were employed. One of them was coated as a test line, while another mAb was labeled with quantum dots and coated onto conjugate pad. The goat anti-mouse IgG was immobilized as a control line. After sample was added, a sandwich structure was formed with CA72-4 and these two mAbs. The fluorescent signal from quantum dots (QD)-labeled mAb in sandwich structure was related to the amount of detected CA72-4. A CCD-based reader was used to realize quantitative detection of CA72-4. Results showed that developed QD-labeled lateral flow strips to detect CA72-4 biomarker with the sensitivity of 2 IU/mL and 10 min detection time. One hundred sera samples from clinical patients with gastric cancer and healthy people were used to confirm specificity of this strip method; results showed that established strip method own 100 % reproducibility and 100 % specificity compared with Roche electrochemiluminescence assay results. In conclusion, CdSe/ZnS quantum dot-labeled lateral flow strips for detection of CA72-4 could realize rapid, sensitive, and specific detection of clinical samples and could own great potential in clinical translation in near future.

Development of a blocking latex agglutination test for the detection of antibodies to chicken anemia virus.

PubMed

Trinh, Dai Quang; Ogawa, Haruko; Bui, Vuong Nghia; Nguyen, Tham Thi Hong; Gronsang, Dulyatad; Baatartsogt, Tugsbaatar; Kizito, Mugimba Kahoza; AboElkhair, Mohammed; Yamaguchi, Shigeo; Nguyen, Viet Khong; Imai, Kunitoshi

2015-09-01

A blocking latex agglutination test (b-LAT) developed in this study was evaluated for the detection of antibodies against chicken anemia virus (CAV) in chickens. Polystyrene latex beads were coupled with a neutralizing monoclonal antibody (mAb) to CAV (mAb-beads). When mAb-beads were mixed with antigens prepared from the lysate of MDCC-MSB1 cells infected with CAV, agglutination occurred. A short pre-incubation of CAV antigens with CAV-specific antiserum inhibited the agglutination of mAb-beads. The test results were obtained within 5min. The specificity of b-LAT was evaluated using sera from specific pathogen-free chickens and sera containing antibodies to avian influenza virus, Newcastle disease virus, infectious bursal disease virus, and Marek's disease virus; nonspecific agglutination and cross-reactivity with antibodies to unrelated viruses were not observed. The examination of 94 serum samples collected from commercial breeder chickens of various ages (17-63 weeks) revealed good agreement (93.6%, Kappa value=0.82) between b-LAT and a virus neutralization test, known to be most sensitive and specific in the detection of antibodies to CAV. These results indicate that b-LAT, a simple and rapid test, is a useful and reliable tool in CAV serology. Copyright © 2015 Elsevier B.V. All rights reserved.

Ability of immunodiagnostic tests to differentiate between dogs naturally infected with Leishmania infantum and Leishmune(®)-vaccinated dogs.

PubMed

Ribeiro, R A N; Teixeira-Neto, R G; Belo, V S; Ferreira, E C; Schallig, H D F H; Silva, E S

2015-06-01

Visceral leishmaniasis (VL) is a serious chronic disease with a lethality rate of up to 10% in humans. In urban areas of Brazil, dogs are the main reservoirs of the etiological agent (Leishmania infantum) of VL, and the Brazilian Ministry of Health recommends the euthanasia of animals that are seropositive in both the immunochromatographic dual path platform rapid test (DPP(®); Bio-Manguinhos) and the enzyme-linked immunosorbent assay (ELISA) with an L. major-like antigen (Bio-Manguinhos). Vaccination is an additional tool in the control of canine VL, but the use of Leishmune(®) (Zoetis Indústria de Produtos Veterinários, São Paulo, SP, Brazil), which contains the fucose mannose ligand (FML) isolated from L. donovani, is not currently recommended by the Brazilian Ministry of Health because vaccinated animals may exhibit positive serology and there are reservations regarding the efficacy of the vaccine. The aims of the present study were: (i) to verify the abilities of the fast agglutination screening test (FAST), the direct agglutination test (DAT), the indirect fluorescent-antibody test (IFAT), the DPP rapid test, and ELISA tests with L. major-like and FML antigens to differentiate between L. infantum-infected and Leishmune(®)-vaccinated dogs, and (ii) to analyze the sensitivities and specificities of the different methods. The reactivities to these tests of Leishmune(®)-vaccinated dogs (n = 71), asymptomatic (n = 20) and symptomatic (n = 20) naturally infected dogs, and unvaccinated healthy control dogs (n = 5) were compared. None of the Leishmune(®)-vaccinated dogs tested seropositive in FAST and DAT, although one dog was reactive to DPP and four dogs to ELISA/L. major-like and IFAT tests. While 69 (97%) of vaccinated dogs reacted to ELISA/FML, only one was seropositive in both ELISA/L. major-like and IFAT tests. Individually, all immunodiagnostic tests presented high specificities and positive likelihood ratios (LR+), and high specificity values were obtained when the tests were considered in pairs. However, sensitivity and LR- values were low for ELISA/L. major-like and IFAT tests individually, and for all pair combinations of tests except for FAST with DPP.

The future of viral hepatitis testing: innovations in testing technologies and approaches.

PubMed

Peeling, Rosanna W; Boeras, Debrah I; Marinucci, Francesco; Easterbrook, Philippa

2017-11-01

A large burden of undiagnosed hepatitis virus cases remains globally. Despite the 257 million people living with chronic hepatitis B virus infection, and 71 million with chronic viraemic HCV infection, most people with hepatitis remain unaware of their infection. Advances in rapid detection technology have created new opportunities for enhancing access to testing and care, as well as monitoring of treatment. This article examines a range of other technological innovations that can be leveraged to provide more affordable and simplified approaches to testing for HBV and HCV infection and monitoring of treatment response. These include improved access to testing through alternative sampling methods (use of dried blood spots, oral fluids, self-testing) and combination rapid diagnostic tests for detection of HIV, HBV and HCV infection; more affordable options for confirmation of virological infection (HBV DNA and HCV RNA) such as point-of-care molecular assays, HCV core antigen and multi-disease polyvalent molecular platforms that make use of existing centralised laboratory based or decentralised TB and HIV instrumentation for viral hepatitis testing; and finally health system improvements such as integration of laboratory services for procurement and sample transportation and enhanced data connectivity to support quality assurance and supply chain management.

Analytical sensitivity of current best-in-class malaria rapid diagnostic tests.

PubMed

Jimenez, Alfons; Rees-Channer, Roxanne R; Perera, Rushini; Gamboa, Dionicia; Chiodini, Peter L; González, Iveth J; Mayor, Alfredo; Ding, Xavier C

2017-03-24

Rapid diagnostic tests (RDTs) are today the most widely used method for malaria diagnosis and are recommended, alongside microscopy, for the confirmation of suspected cases before the administration of anti-malarial treatment. The diagnostic performance of RDTs, as compared to microscopy or PCR is well described but the actual analytical sensitivity of current best-in-class tests is poorly documented. This value is however a key performance indicator and a benchmark value needed to developed new RDTs of improved sensitivity. Thirteen RDTs detecting either the Plasmodium falciparum histidine rich protein 2 (HRP2) or the plasmodial lactate dehydrogenase (pLDH) antigens were selected from the best performing RDTs according to the WHO-FIND product testing programme. The analytical sensitivity of these products was evaluated using a range of reference materials including P. falciparum and Plasmodium vivax whole parasite samples as well as recombinant proteins. The best performing HRP2-based RDTs could detect all P. falciparum cultured samples at concentrations as low as 0.8 ng/mL of HRP2. The limit of detection of the best performing pLDH-based RDT specifically detecting P. vivax was 25 ng/mL of pLDH. The analytical sensitivity of P. vivax and Pan pLDH-based RDTs appears to vary considerably from product to product, and improvement of the limit-of-detection for P. vivax detecting RDTs is needed to match the performance of HRP2 and Pf pLDH-based RDTs for P. falciparum. Different assays using different reference materials produce different values for antigen concentration in a given specimen, highlighting the need to establish universal reference assays.

Surface grafted antibodies: controlled architecture permits enhanced antigen detection.

PubMed

Sebra, Robert P; Masters, Kristyn S; Bowman, Christopher N; Anseth, Kristi S

2005-11-22

The attachment of antibodies to substrate surfaces is useful for achieving specific detection of antigens and toxins associated with clinical and field diagnostics. Here, acrylated whole antibodies were produced through conjugation chemistry, with the goal of covalently photografting these proteins from surfaces in a controlled fashion, to facilitate rapid and sensitive antigenic detection. A living radical photopolymerization chemistry was used to graft the acrylated whole antibodies on polymer surfaces at controlled densities and spatial locations by controlling the exposure time and area, respectively. Copolymer grafts containing these antibodies were synthesized to demonstrate two principles. First, PEG functionalities were introduced to prevent nonspecific protein interactions and improve the reaction kinetics by increasing solvation and mobility of the antibody-containing chains. Both of these properties lead to sensitive (pM) and rapid (<20 min) detection of antigens with this surface modification technique. Second, graft composition was tailored to include multiple antibodies on the same grafted chains, establishing a means for simultaneously detecting multiple antigens on one grafted surface area. Finally, the addition of PEG spacers between the acrylate functionality and the pendant detection antibodies was tuned to enhance the detection of a short-half-life molecule, glucagon, in a complex biological environment, plasma.

[Examination about utility of a Streptococcus pneumoniae capsular antigen swiftness search kit urine in a pneumonia patient].

PubMed

Hashikita, Giichi; Yamaguti, Toshiyuki; Tachi, Yoshimi; Kishi, Etsuko; Kawamura, Toru; Takahashi, Shun; Arai, Yukie; Koyama, Sachie; Huruhata, Toshihumi; Itabashi, Akira; Oka, Yoko; Yamazaki, Tsutomu; Maesaki, Sigefumi

2005-01-01

We investigated the usefullness of Binax NOW urine antigen test, an immunochromatographic assay that binds any soluble Streptococcus pneumoniae antigen (C polysaccharide) for the diagnosis of penumoniae form September 2003 to March 2005. We used 372 samples form the patinets with pneumoniae diagnosed for blood or sputum cultuter or gram-stained sputum smear. Out of 24 culture positive specimens, Binax NOW urine antigen test, showed positive in 18 (75%) specimens. The sensitivity of sputum and blood culture was 71.7% and 83.3%, respectively. Binax NOW urine antigen test was seemed false positives in 55 samples, false negatives in 6 samples. The specificity of Binax NOW urine antigen test was evaluated 84.1%. Overall agreement among tests was 83.6%. When compared to culture, false negative urine antigen may be the result of colonizing S. pneumoniae in sputum or pneumonia caused by an agent other than S. pneumoniae. CRP values for cases were both urine antigen and culture were positive ranged from 40 mg/dl to 10 mg/dl while urine antigen and culture negative cases were predominantly less than 10 mg/dl. Positive blood and pleural fluid culture cases were consistently associated with strongly positive urine antigen tests. Non-agreement between urine antigen, culture, and microscopy may be the result of specimen quality, labile nature of S. pneumoniae and antimicrobial therapy.

Rapid Detection of Hepatitis B Virus Surface Antigen by an Agglutination Assay Mediated by a Bispecific Diabody against Both Human Erythrocytes and Hepatitis B Virus Surface Antigen▿

PubMed Central

Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi

2007-01-01

Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens. PMID:17442848

Rapid antigen test use for the management of group A streptococcal pharyngitis in community pharmacies.

PubMed

Demoré, Béatrice; Tebano, Gianpiero; Gravoulet, Julien; Wilcke, Christophe; Ruspini, Eric; Birgé, Jacques; Boivin, Jean-Marc; Hénard, Sandrine; Dieterling, Annick; Munerol, Lidiana; Husson, Julie; Rabaud, Christian; Pulcini, Céline; Malblanc, Sophie

2018-06-06

Despite group A streptococci being an infrequent cause of pharyngitis in adult outpatients, sore throat remains a common indication for antibiotic prescription. This prospective multicentre non-randomised study describes a community pharmacy-based antimicrobial stewardship intervention consisting in the implementation of rapid antigen testing (RAT) for the management of adults with sore throat. Trained pharmacists triaged patients presenting with symptoms of pharyngitis using the modified Centor score. Those at risk for streptococcal infection were tested with RAT. Patients with a positive RAT were invited to consult a physician, whereas others were offered a symptomatic treatment. All patients received educational leaflets and were asked to fill in a follow-up form 7 days later. Ninety-eight pharmacies in one French region participated, and 559 patients were included over 6 months. RAT was proposed in 367 (65.7%) cases, and it was positive in 28 (8.3%). The follow-up form was returned by 140 (38.5%) participants. Of these, 10/10 patients with positive RAT further consulted a physician and were prescribed an antibiotic treatment, whereas 96.5% (110/114) of patients with negative results and not having any other reason to seek for doctor's advice did not consult. All participants found the intervention useful. Pharmacists spent 6-15 min to perform the intervention, and 98.6% (73/74) of pharmacists giving a feedback declared to be ready to implement this intervention in daily practice, if endorsed and reimbursed. Our results suggest that a pharmacy-based programme for the management of sore throat is feasible and could increase adherence to guidelines.

Requirements for diagnosis of malaria at different levels of the laboratory network in Africa.

PubMed

Long, Earl G

2009-06-01

The rapid increase of resistance to cheap, reliable antimalarials, the increasing cost of effective drugs, and the low specificity of clinical diagnosis has increased the need for more reliable diagnostic methods for malaria. The most commonly used and most reliable remains microscopic examination of stained blood smears, but this technique requires skilled personnel, precision instruments, and ideally a source of electricity. Microscopy has the advantage of enabling the examiner to identify the species, stage, and density of an infection. An alternative to microscopy is the rapid diagnostic test (RDT), which uses a labeled monoclonal antibody to detect circulating parasitic antigens. This test is most commonly used to detect Plasmodium falciparum infections and is available in a plastic cassette format. Both microscopy and RDTs should be available at all levels of laboratory service in endemic areas, but in peripheral laboratories with minimally trained staff, the RDT may be a more practical diagnostic method.

Rapid, ultrasensitive detection of microorganisms based on interferometry and lab-on-a-chip nanotechnology

NASA Astrophysics Data System (ADS)

Ymeti, Aurel; Nederkoorn, Paul H. J.; Dudia, Alma; Subramaniam, Vinod; Kanger, Johannes S.

2009-05-01

Future viral outbreaks are a major threat to societal and economic development throughout the world. A rapid, sensitive, and easy-to-use test for viral infections is essential to prevent and to control such viral pandemics. Furthermore, a compact, portable device is potentially very useful in remote or developing regions without easy access to sophisticated laboratory facilities. We have developed a rapid, ultrasensitive sensor that could be used in a handheld device to detect various viruses and measure their concentration. The essential innovation in this technique is the combination of an integrated optical interferometric sensor with antibody-antigen recognition approaches to yield a very sensitive, very rapid test for virus detection. The sensor is able to spot the herpes virus at concentrations of just 850 particles per milliliter under physiological conditions. The sensitivity of the sensor approaches detection of a single virus particle, yielding a sensor of unprecedented sensitivity with wide applications for viral diagnostics. The sensor's detection principle can be extended to any biological target such as bacteria, cells and proteins and for which there are specific antibodies. The nature of the sensor enables multiplexed detection of several analytes at the same time.

Commercial Molecular Tests for Fungal Diagnosis from a Practical Point of View.

PubMed

Lackner, Michaela; Lass-Flörl, Cornelia

2017-01-01

The increasing interest in molecular diagnostics is a result of tremendously improved knowledge on fungal infections in the past 20 years and the rapid development of new methods, in particular polymerase chain reaction. High expectations have been placed on molecular diagnostics, and the number of laboratories now using the relevant technology is rapidly increasing-resulting in an obvious need for standardization and definition of laboratory organization. In the past 10 years, multiple new molecular tools were marketed for the detection of DNA, antibodies, cell wall components, or other antigens. In contrast to classical culture methods, molecular methods do not detect a viable organisms, but only molecules which indicate its presence; this can be nucleic acids, cell components (antigens), or antibodies (Fig. 1). In this chapter, an overview is provided on commercially available detection tools, their strength and how to use them. A main focus is laid on providing tips and tricks that make daily life easier. We try to focus and mention methodical details which are not highlighted in the manufacturer's instructions of these test kits, but are based on our personal experience in the laboratory. Important to keep in mind is that molecular tools cannot replace culture, microscopy, or a critical view on patients' clinical history, signs, and symptoms, but provide a valuable add on tool. Diagnosis should not be based solely on a molecular test, but molecular tools might deliver an important piece of information that helps matching the diagnostic puzzle to a diagnosis, in particular as few tests are in vitro diagnostic tests (IVD) or only part of the whole test carries the IVD certificate (e.g., DNA extraction is often not included). Please be aware that the authors do not claim to provide a complete overview on all commercially available diagnostic assays being currently marketed for fungal detection, as those are subject to constant change. A main focus is put on commonly used panfungal assays and pathogen-specific assays, including Aspergillus-specific, Candida-specific, Cryptococcus specific, Histoplasma-specific, and Pneumocystis-specific assays. Assays are categorized according to their underlying principle in either antigen-detecting or antibody-detecting or DNA-detecting (Fig. 1). Other non-DNA-detecting nucleic acid methods such as FISH and PNA FISH are not summarized in this chapter and an overview on test performance, common false positives, and the clinical evaluation of commercial tests in studies is provided already in a previous book series by Javier Yugueros Marcos and David H. Pincus (Marcos and Pincus, Methods Mol Biol 968:25-54, 2013).

Use of synthetic, crystalline, L-α-dimyristoyl lecithin in cardiolipin antigens

PubMed Central

Reyn, Alice; Bentzon, Michael Weis

1956-01-01

Experiments were carried out by the authors to determine whether synthetic, crystalline, L-α-dimyristoyl lecithin could replace natural purified lecithins in the preparation of cardiolipin antigens. These experiments were designed specifically to find out whether it was possible to obtain the same serological reactions, qualitatively and quantitatively, with the test antigen as with a reference antigen containing natural lecithin, and whether the test antigen had the same keeping qualities as the reference antigen. The tests used were the quantitative complement-fixation test as modified by Mørch in 1933, and the VDRL slide flocculation test. The results showed that synthetic, crystalline, L-α-dimyristoyl lecithin could replace natural lecithin in the preparation of cardiolipin antigens, but that the antigens prepared with the synthetic lecithin were significantly less sensitive than those prepared with an equimolar amount of natural lecithin. The authors consider that further investigation is required before the use of synthetic lecithin is finally adopted. PMID:13342931

Polyvalent heat-killed antigen for the diagnosis of infection with Legionella pneumophila.

PubMed Central

Fallon, R J; Abraham, W H

1982-01-01

A polyvalent antigen composed of heat-killed agar-grown Legionella pneumophila serogroups 1-4 suspended in a suspension of yolk-sac from embryonated hens' eggs has been examined for use in the indirect fluorescent antibody test for Legionella infection. The serological response detected by monovalent antigen correlated well with that detected by polyvalent antigen. The use of polyvalent antigen forms a useful screening test for the detection of antibody to L pneumophila, but positive results must be confirmed by test using monovalent antigen. PMID:7042762

Antibodies Targeting Closely Adjacent or Minimally Overlapping Epitopes Can Displace One Another.

PubMed

Abdiche, Yasmina Noubia; Yeung, Andy Yik; Ni, Irene; Stone, Donna; Miles, Adam; Morishige, Winse; Rossi, Andrea; Strop, Pavel

2017-01-01

Here we describe how real-time label-free biosensors can be used to identify antibodies that compete for closely adjacent or minimally overlapping epitopes on their specific antigen via a mechanism of antibody displacement. By kinetically perturbing one another's binding towards their antigen via the formation of a transient trimolecular complex, antibodies can displace one another in a fully reversible and dose-dependent manner. Displacements can be readily identified when epitope binning assays are performed in a classical sandwich assay format whereby a solution antibody (analyte) is tested for binding to its antigen that is first captured via an immobilized antibody (ligand) because an inverted sandwiching response is observed when an analyte displaces a ligand, signifying the antigen's unusually rapid dissociation from its ligand. In addition to classifying antibodies within a panel in terms of their ability to block or sandwich pair with one another, displacement provides a hybrid mechanism of competition. Using high-throughput epitope binning studies we demonstrate that displacements can be observed on any target, if the antibody panel contains appropriate epitope diversity. Unidirectional displacements occurring between disparate-affinity antibodies can generate apparent asymmetries in a cross-blocking experiment, confounding their interpretation. However, examining competition across a wide enough concentration range will often reveal that these displacements are reversible. Displacement provides a gentle and efficient way of eluting antigen from an otherwise high affinity binding partner which can be leveraged in designing reagents or therapeutic antibodies with unique properties.

Laboratory Diagnosis of Zika Virus Infection.

PubMed

Landry, Marie Louise; St George, Kirsten

2017-01-01

-The rapid and accurate diagnosis of Zika virus infection is an international priority. -To review current recommendations, methods, limitations, and priorities for Zika virus testing. -Sources include published literature, public health recommendations, laboratory procedures, and testing experience. -Until recently, the laboratory diagnosis of Zika infection was confined to public health or research laboratories that prepared their own reagents, and test capacity has been limited. Furthermore, Zika cross-reacts serologically with other flaviviruses, such as dengue, West Nile, and yellow fever. Current or past infection, or even vaccination with another flavivirus, will often cause false-positive or uninterpretable Zika serology results. Detection of viral RNA during acute infection using nucleic acid amplification tests provides more specific results, and a number of commercial nucleic acid amplification tests have received emergency use authorization. In addition to serum, testing of whole blood and urine is recommended because of the higher vial loads and longer duration of shedding. However, nucleic acid amplification testing has limited utility because many patients are asymptomatic or present for testing after the brief period of Zika shedding has passed. Thus, the greatest need and most difficult challenge is development of accurate antibody tests for the diagnosis of recent Zika infection. Research is urgently needed to identify Zika virus epitopes that do not cross-react with other flavivirus antigens. New information is emerging at a rapid pace and, with ongoing public-private and international collaborations and government support, it is hoped that rapid progress will be made in developing robust and widely applicable diagnostic tools.

Application and Comparative Evaluation of Fluorescent Antibody, Immunohistochemistry and Reverse Transcription Polymerase Chain Reaction Tests for the Detection of Rabies Virus Antigen or Nucleic Acid in Brain Samples of Animals Suspected of Rabies in India.

PubMed

Prabhu, K Nithin; Isloor, Shrikrishna; Veeresh, B Hanchinal; Rathnamma, Doddamane; Sharada, R; Das, Lekshmi J; Satyanarayana, M L; Hegde, Nagendra R; Rahman, Sira Abdul

2018-02-28

Accurate and early diagnosis of animal rabies is critical for undertaking public health measures. Whereas the direct fluorescent antibody (DFA) technique is the recommended test, the more convenient, direct rapid immunochemistry test (dRIT), as well as the more sensitive, reverse transcription polymerase chain reaction (RT-PCR), have recently been employed for the laboratory diagnosis of rabies. We compared the three methods on brain samples from domestic (dog, cat, cattle, buffalo, horse, pig and goat) and wild (leopard, wolf and jackal) animals from various parts of India. Of the 257 samples tested, 167 were positive by all the three tests; in addition, 35 of the 36 decomposed samples were positive by RT-PCR. This is the first study in which such large number of animal samples have been subjected to the three tests simultaneously. The results confirm 100% corroboration between DFA and dRIT, buttress the applicability of dRIT in the simple and rapid diagnosis of rabies in animals, and reaffirm the suitability of RT-PCR for samples unfit for testing either by DFA or dRIT.

Application of Protein Microarrays for Multiplexed Detection of Antibodies to Tumor Antigens in Breast Cancer

PubMed Central

Anderson, Karen S.; Ramachandran, Niroshan; Wong, Jessica; Raphael, Jacob V.; Hainsworth, Eugenie; Demirkan, Gokhan; Cramer, Daniel; Aronzon, Diana; Hodi, F. Stephen; Harris, Lyndsay; Logvinenko, Tanya; LaBaer, Joshua

2012-01-01

There is strong preclinical evidence that cancer, including breast cancer, undergoes immune surveillance. This continual monitoring, by both the innate and the adaptive immune systems, recognizes changes in protein expression, mutation, folding, glycosylation, and degradation. Local immune responses to tumor antigens are amplified in draining lymph nodes, and then enter the systemic circulation. The antibody response to tumor antigens, such as p53 protein, are robust, stable, and easily detected in serum, may exist in greater concentrations than their cognate antigens, and are potential highly specific biomarkers for cancer. However, antibodies have limited sensitivities as single analytes, and differences in protein purification and assay characteristics have limited their clinical application. For example, p53 autoantibodies in the sera are highly specific for cancer patients, but are only detected in the sera of 10-20% of patients with breast cancer. Detection of p53 autoantibodies is dependent on tumor burden, p53 mutation, rapidly decreases with effective therapy, but is relatively independent of breast cancer subtype. Although antibodies to hundreds of other tumor antigens have been identified in the sera of breast cancer patients, very little is known about the specificity and clinical impact of the antibody immune repertoire to breast cancer. Recent advances in proteomic technologies have the potential for rapid identification of immune response signatures for breast cancer diagnosis and monitoring. We have adapted programmable protein microarrays for the specific detection of autoantibodies in breast cancer. Here, we present the first demonstration of the application of programmable protein microarray ELISAs for the rapid identification of breast cancer autoantibodies. PMID:18311903

Improving the Immunogenicity of the Mycobacterium bovis BCG Vaccine by Non-Genetic Bacterial Surface Decoration Using the Avidin-Biotin System.

PubMed

Liao, Ting-Yu Angela; Lau, Alice; Joseph, Sunil; Hytönen, Vesa; Hmama, Zakaria

2015-01-01

Current strategies to improve the current BCG vaccine attempt to over-express genes encoding specific M. tuberculosis (Mtb) antigens and/or regulators of antigen presentation function, which indeed have the potential to reshape BCG in many ways. However, these approaches often face serious difficulties, in particular the efficiency and stability of gene expression via nucleic acid complementation and safety concerns associated with the introduction of exogenous DNA. As an alternative, we developed a novel non-genetic approach for rapid and efficient display of exogenous proteins on bacterial cell surface. The technology involves expression of proteins of interest in fusion with a mutant version of monomeric avidin that has the feature of reversible binding to biotin. Fusion proteins are then used to decorate the surface of biotinylated BCG. Surface coating of BCG with recombinant proteins was highly reproducible and stable. It also resisted to the freeze-drying shock routinely used in manufacturing conventional BCG. Modifications of BCG surface did not affect its growth in culture media neither its survival within the host cell. Macrophages phagocytized coated BCG bacteria, which efficiently delivered their surface cargo of avidin fusion proteins to MHC class I and class II antigen presentation compartments. Thereafter, chimeric proteins corresponding to a surrogate antigen derived from ovalbumin and the Mtb specific ESAT6 antigen were generated and tested for immunogenicity in vaccinated mice. We found that BCG displaying ovalbumin antigen induces an immune response with a magnitude similar to that induced by BCG genetically expressing the same surrogate antigen. We also found that BCG decorated with Mtb specific antigen ESAT6 successfully induces the expansion of specific T cell responses. This novel technology, therefore, represents a practical and effective alternative to DNA-based gene expression for upgrading the current BCG vaccine.

Development of a quantitative rapid diagnostic test for multibacillary leprosy using smart phone technology.

PubMed

Paula Vaz Cardoso, Ludimila; Dias, Ronaldo Ferreira; Freitas, Aline Araújo; Hungria, Emerith Mayra; Oliveira, Regiane Morillas; Collovati, Marco; Reed, Steven G; Duthie, Malcolm S; Martins Araújo Stefani, Mariane

2013-10-23

Despite efforts to eliminate leprosy as public health problem, delayed diagnosis and disabilities still occur in many countries. Leprosy diagnosis remains based on clinical manifestations and the number of clinicians with expertise in leprosy diagnosis is in decline. We have developed a new immunochromatographic test with the goal of producing a simple and rapid system that can be used, with a minimal amount of training, to provide an objective and consistent diagnosis of multibacillary leprosy. The test immobilizes two antigens that have been recognized as excellent candidates for serologic diagnosis (the PGL-I mimetic, ND-O, and LID-1), on a nitrocellulose membrane. This allows the detection of specific IgM and IgG antibodies within 20 minutes of the addition of patient sera. Furthermore, we coupled the NDO-LID® rapid tests with a new cell phone-based test reader platform (Smart Reader®) to provide objective interpretation that was both quantifiable and consistent. Direct comparison of serologic responses indicated that the rapid test detected a greater proportion of leprosy patients than a lab-based PGL-I ELISA. While positive responses were detected by PGL-I ELISA in 83.3% of multibacillary patients and 15.4% of paucibacillary patients, these numbers were increased to 87% and 21.2%, respectively, when a combination of the NDO-LID® test and Smart Reader® was used. Among multibacillary leprosy the sensitivity of NDO-LID® test assessed by Smart Reader® was 87% (95% CI, 79.2-92.7%) and the specificity was 96.1% (95% CI, 91.7- 98.6%). The positive predictive value and the negative predictive value of NDO-LID® tests were 94% (95% CI, 87.4-97.8%) and 91.4% (95% CI, 85.9-95.2%), respectively. The widespread provision of rapid diagnostic tests to facilitate the diagnosis or prognosis of multibacillary leprosy could impact on leprosy control programs by aiding early detection, directing appropriate treatment and potentially interrupting Mycobacterium leprae transmission.

Microbial ecology measurement system

NASA Technical Reports Server (NTRS)

1972-01-01

The sensitivity and potential rapidity of the PIA test that was demonstrated during the feasibility study warranted continuing the effort to examine the possibility of adapting this test to an automated procedure that could be used during manned missions. The effort during this program has optimized the test conditions for two important respiratory pathogens, influenza virus and Mycoplasma pneumoniae, developed a laboratory model automated detection system, and investigated a group antigen concept for virus detection. Preliminary tests on the handling of oropharygeal clinical samples for PIA testing were performed using the adenovirus system. The results obtained indicated that the PIA signal is reduced in positive samples and is increased in negative samples. Treatment with cysteine appeared to reduce nonspecific agglutination in negative samples but did not maintain the signal in positive samples.

[Physicians with access to point-of-care tests significantly reduce the antibiotic prescription for common cold].

PubMed

Llor, Carles; Hernández, Silvia; Cots, Josep María; Bjerrum, Lars; González, Beatriz; García, Guillermo; Alcántara, Juan de Dios; Guerra, Gloria; Cid, Marina; Gómez, Manuel; Ortega, Jesús; Pérez, Carolina; Arranz, Javier; Monedero, María José; Paredes, José; Pineda, Vicenta

2013-03-01

This study was aimed at evaluating the effect of two levels of intervention on the antibiotic prescribing in patients with common cold. Before and after audit-based study carried out in primary healthcare centres in Spain. General practitioners registered all the episodes of common cold during 15 working days in January and February in 2008 (preintervention). Two types of intervention were considered: full intervention, consisting in individual feedback based on results from the first registry, courses in rational antibiotic prescribing, guidelines, patient information leaflets, workshops on rapid tests -rapid antigen detection and C-reactive protein tests- and provision of these tests in the surgeries; and partial intervention, consisting of all the above intervention except for the workshop and they did not have access to rapid tests. The same registry was repeated in 2009 (postintervention). In addition, new physicians filled out only the registry in 2009 (control group). 210 physicians underwent the full intervention, 71 the partial intervention and 59 were assigned to the control group. The 340 doctors prescribed antibiotics in 274 episodes of a total of 12,373 cases registered (2.2%).The greatest percentage of antibiotic prescription was found in the control group (4.6%). The partial intervention increased the antibiotic prescription percentage from 1.1% to 2.7% while only doctors who underwent the complete intervention lead to a significant reduction of antibiotics prescribed, from 2.9% before to 0.7% after the intervention (p<0.001). Only physicians with access to rapid tests significantly reduced antibiotic prescription in patients with common cold.

Accuracy of dengue clinical diagnosis with and without NS1 antigen rapid test: Comparison between human and Bayesian network model decision.

PubMed

Sa-Ngamuang, Chaitawat; Haddawy, Peter; Luvira, Viravarn; Piyaphanee, Watcharapong; Iamsirithaworn, Sopon; Lawpoolsri, Saranath

2018-06-18

Differentiating dengue patients from other acute febrile illness patients is a great challenge among physicians. Several dengue diagnosis methods are recommended by WHO. The application of specific laboratory tests is still limited due to high cost, lack of equipment, and uncertain validity. Therefore, clinical diagnosis remains a common practice especially in resource limited settings. Bayesian networks have been shown to be a useful tool for diagnostic decision support. This study aimed to construct Bayesian network models using basic demographic, clinical, and laboratory profiles of acute febrile illness patients to diagnose dengue. Data of 397 acute undifferentiated febrile illness patients who visited the fever clinic of the Bangkok Hospital for Tropical Diseases, Thailand, were used for model construction and validation. The two best final models were selected: one with and one without NS1 rapid test result. The diagnostic accuracy of the models was compared with that of physicians on the same set of patients. The Bayesian network models provided good diagnostic accuracy of dengue infection, with ROC AUC of 0.80 and 0.75 for models with and without NS1 rapid test result, respectively. The models had approximately 80% specificity and 70% sensitivity, similar to the diagnostic accuracy of the hospital's fellows in infectious disease. Including information on NS1 rapid test improved the specificity, but reduced the sensitivity, both in model and physician diagnoses. The Bayesian network model developed in this study could be useful to assist physicians in diagnosing dengue, particularly in regions where experienced physicians and laboratory confirmation tests are limited.

Porous silicon photoluminescence biosensor for rapid and sensitive detection of toxins

NASA Astrophysics Data System (ADS)

Melnyk, Yulia; Pavlova, Karyna; Myndrul, Valerii; Viter, Roman; Smyntyna, Valentyn; Iatsunskyi, Igor

2017-08-01

A rapid and low cost photoluminescence (PL) immunosensor for the determination of low concentrations of Ochratoxin A(OTA) and Aflatoxine B1 (AfB1) has been developed. This biosensor was based on porous silicon (PSi) fabricated by metal-assisted chemical etching (MACE) and modified by antibodies against OTA/AfB1 (anti-OTA/anti-AfB1). Biofunctionalization method of the PSi surface by anti-OTA/ anti-AfB1 was developed. The changes of the PL intensity after interaction of the immobilized anti-OTA/anti-AfB1with OTA/AfB1 antigens were used as biosensor signal, allowing sensitive and selective detection of OTA/AfB1 antigens in BSA solution. The sensitivity of the reported optical biosensor towards OTA/AfB1 antigens is in the range from 10-3 to 102 ng/ml.

Personalized Therapy: Tumor Antigen Discovery for Adoptive Cellular Therapy.

PubMed

Yee, Cassian; Lizee, Gregory A

Adoptive cell therapy using endogenous T cells involves the ex vivo isolation and expansion of antigen-specific T cells from the peripheral blood and is uniquely suited for validating and translating antigen discovery. Endogenous T-cell therapy does not require accessible tumor as a source of infiltrating T cells and is free of regulatory and logistical constraints associated with engineering T cells. Candidate epitope peptides identified through antigen discovery may be rapidly implemented as targets in clinical trials of endogenous T-cell therapy and even incorporated as an "ad hoc" approach to personalized treatment when autologous tumor is available. Several first-in-human studies using a uniform population of antigen-specific T cells defined by phenotype and specificity have provided a means to confirm candidate antigens as potential tumor rejection antigens and to evaluate the reasons for success or failure using as a "transferrable cellular biomarker" the adoptively transferred T cells.

Evaluation of the Kodak Surecell Chlamydia test for the laboratory diagnosis of adult inclusion conjunctivitis.

PubMed

Tantisira, J G; Kowalski, R P; Gordon, Y J

1995-07-01

The Kodak Surecell Chlamydia test, a rapid enzyme immunoassay, has been reported to be highly sensitive (93%) and specific (96%) for detecting chlamydial lipopolysaccharide antigen in conjunctival specimens from infants, but has not been evaluated previously in adult conjunctival specimens. This study was designed to determine the efficacy of the Kodak Surecell Chlamydia test for the laboratory diagnosis of adult inclusion conjunctivitis. Twenty Chlamydia culture-positive conjunctival specimens from adults (true-positives) and 20 true-negative specimens were tested with the Kodak Surecell Chlamydia test. The Kodak Surecell Chlamydia test was 40% (8/20) sensitive, 100% (20/20) specific, and 70% (28/40) efficient. This study indicates that the Kodak Surecell Chlamydia test, though highly specific, is less sensitive in its ability to diagnose chlamydial conjunctivitis in adults than has been reported previously in infants.

Serotype determination of Salmonella by xTAG assay.

PubMed

Zheng, Zhibei; Zheng, Wei; Wang, Haoqiu; Pan, Jincao; Pu, Xiaoying

2017-10-01

Currently, no protocols or commercial kits are available to determine the serotypes of Salmonella by using Luminex MAGPIX®. In this study, an xTAG assay for serotype determination of Salmonella suitable for Luminex MAGPIX® is described and 228 Salmonella isolates were serotype determined by this xTAG assay. The xTAG assay consists of two steps: 1) Multiplex PCR to amplify simultaneously O, H and Vi antigen genes of Salmonella, and 2) Magplex-TAG™ microsphere hybridization to identify accurately the specific PCR products of different antigens. Compared with the serotyping results of traditional serum agglutination test, the sensitivity and specificity of the xTAG assay were 95.1% and 100%, respectively. The agreement rate of these two assays was 95.2%. Compared with Luminex xMAP® Salmonella Serotyping Assay (SSA) kit, the advantages of this xTAG assay are: First, the magnetic beads make it applicable to both the Luminex®100/200™ and MAGPIX® systems. Second, only primers rather than both primers and probes are needed in the xTAG assay, and the process of coupling antigen-specific oligonucleotide probes to beads is circumvented, which make the xTAG assay convenient to be utilized by other laboratories. The xTAG assay may serve as a rapid alternative or complementary method for traditional Salmonella serotyping tests, especially for laboratories that utilize the MAGPIX® systems. Copyright © 2017 Elsevier B.V. All rights reserved.

Rapid Determination of Salmonella in Samples of Egg Noodles, Cake Mixes, and Candies

PubMed Central

Banwart, George J.; Kreitzer, Madeleine J.

1969-01-01

A glass apparatus system was compared with a standard enrichment broth-selective agar method to test samples of egg noodles, cake mixes, and candy for the presence or absence of salmonellae. The glass apparatus system used fermentation of mannitol, production of H2S, or motility, in conjunction with a serological test of flagellar antigens, to detect salmonellae. No salmonellae were detected in 173 samples of food products. Of these samples, 171 were found to be Salmonella-negative after 48 hr with the glass apparatus system. After 72 hr, the standard Salmonella procedure yielded 38 samples which produced Salmonella false-positive results on selective agars. Inoculation of samples with cultures of Salmonella showed that approximately one inoculated cell could be detected after 48 hr of incubation with the glass apparatus. The standard Salmonella test requires a minimum of 72 hr for completion. Compared with the standard Salmonella test, the glass apparatus system is a more rapid and simple system that can be used to determine the presence or absence of Salmonella in these food products. Images PMID:5370460

Ultrasensitive Immunochromatographic Strip for Fast Screening of 27 Sulfonamides in Honey and Pork Liver Samples Based on a Monoclonal Antibody.

PubMed

Chen, Yanni; Guo, Lingling; Liu, Liqiang; Song, Shanshan; Kuang, Hua; Xu, Chuanlai

2017-09-20

Group-specific monoclonal antibodies (Mabs) with selectivity for 27 sulfonamides were developed based on new combinations of immunogen and coating antigen. The Mab was able to recognize 27 sulfonamides with 50% inhibition concentration (IC 50 ) values ranging from 0.15 to 15.38 μg/L. In particular, the IC 50 values for five sulfonamides (sulfamethazine, sulfaquinoxaline, sulfamonomethoxine, sulfadimethoxine, and sulfamethoxazole) were 0.51, 0.15, 0.56, 0.54, and 2.14 μg/L, respectively. On the basis of the Mab, an immunochromatographic lateral flow strip test was established for rapid screening of sulfonamides in honey samples. The visual limit of detection of the strip test for most sulfonamides in spiked honey samples was below 10 μg/kg, satisfying the requirements of authorities. Positive honey and pork liver samples, which had been confirmed by high-performance liquid chromatography/mass spectrometry, were used to validate the reliability of the proposed strip test. The immunochromatographic lateral flow strip test provides a rapid and convenient method for fast screening of sulfonamides in honey samples.

Use of immunochromatographic assay for rapid identification of Mycobacterium tuberculosis complex from liquid culture

PubMed Central

Považan, Anika; Vukelić, Anka; Savković, Tijana; Kurucin, Tatjana

2012-01-01

A new, simple immunochromatographic assay for rapid identification of Mycobacterium tuberculosis complex in liquid cultures has been developed. The principle of the assay is binding of the Mycobacterium tuberculosis complex specific antigen to the monoclonal antibody conjugated on the test strip. The aim of this study is evaluation of the performance of immunochromatographic assay in identification of Mycobacterium tuberculosis complex in primary positive liquid cultures of BacT/Alert automated system. A total of 159 primary positive liquid cultures were tested using the immunochromatographic assay (BD MGIT TBc ID) and the conventional subculture, followed by identification using biochemical tests. Of 159 positive liquid cultures, using the conventional method, Mycobacterium tuberculos is was identified in 119 (74.8%), nontuberculous mycobacteria were found in 4 (2.5%), 14 (8.8%) cultures were contaminated and 22 (13.8%) cultures were found to be negative. Using the immunochromatographic assay, Mycobacterium tuberculosis complex was detected in 118 (74.2%) liquid cultures, and 41 (25.8%) tests were negative. Sensitivity, specificity, positive and negative predictive values of the test were 98.3%; 97.5%; 99.15%; 95.12%, respectively. The value of kappa test was 0.950, and McNemar test was 1.00. The immunochromatographic assay is a simple and rapid test which represents a suitable alternative to the conventional subculture method for the primary identification of Mycobacterium tuberculosis complex in liquid cultures of BacT/Alert automated system. PMID:22364301

Development of a one-step immunochromatographic strip test using gold nanoparticles for the rapid detection of Salmonella typhi in human serum.

PubMed

Preechakasedkit, Pattarachaya; Pinwattana, Kulwadee; Dungchai, Wijitar; Siangproh, Weena; Chaicumpa, Wanpen; Tongtawe, Pongsri; Chailapakul, Orawon

2012-01-15

An immunochromatographic strip test using gold nanoparticles was developed for the rapid detection of Salmonella typhi (S. typhi) in human serum. The strip test based on the principle of sandwich immunoassay by the specific binding of antigens from S. typhi O901 and antibody of S. typhi O901 on a nitrocellulose membrane. Antibody-gold nanoparticle conjugate was used as the label and was coated onto a glass fiber membrane, which was used as a conjugate pad. To create a test and control zone, antibody of S. typhi O901 and an anti-IgG were dotted on the nitrocellulose membrane, respectively. Positive samples were displayed as red dots at the test and control zones of the nitrocellulose membrane, while negative samples resulted in a red dot only in the control zone. The limit of detection (LOD) was found to be 1.14×10(5) cfu mL(-1), which could be visually detected by the naked eye within 15 min. This strip test provided a lower detection limit and analysis time than a dot blot immunoassay (8.88×10(6) cfu mL(-1) for LOD and 110 min for reaction time). In addition, our immunochromatographic strip test was employed to detect S. typhi in human serum effectively, with high accuracy. This strip test offers great promise for a rapid, simple and low-cost analysis of S. typhi. Copyright © 2011 Elsevier B.V. All rights reserved.

Occult Hepatitis B virus infection in previously screened, blood donors in Ile-Ife, Nigeria: implications for blood transfusion and stem cell transplantation.

PubMed

Olotu, Amadin A; Oyelese, Adesola O; Salawu, Lateef; Audu, Rosemary A; Okwuraiwe, Azuka P; Aboderin, Aaron O

2016-05-05

Hepatitis B virus (HBV) transmission through blood transfusion is reduced by screening for hepatitis B surface antigen (HBsAg). However this method cannot detect the presence of occult hepatitis B virus infection. This study sought to determine the prevalence of occult hepatitis B virus infection among blood donors in Ile-Ife, Nigeria. For the first time in Nigeria we employed an automated real-time PCR- method to investigate the prevalence of occult HBV in blood donors. Blood donors screened with HBsAg immunochromatographic rapid test kits at the blood transfusion units of two hospitals and found to be negative were recruited into the study. Questionnaires to elicit risk factors for HBV infection were administered and then 10 ml of blood was collected from each donor. Plasma samples obtained from these HBsAg negative blood donors were screened again for HBsAg using an enzyme-linked immunosorbent assay (ELISA) method, and those found negative were screened for the presence of total antibody to the HBV core antigen (anti-HBc) using ELISA method. Those positive to anti-HBc were then tested for HBV DNA, using an automated real-time PCR method. Five hundred and seven blood donors found HBsAg negative by immunochromatographic rapid test kits at both blood transfusion units, were tested for HBsAg using ELISA and 5 (1 %) were HBsAg positive. The 502 found negative were tested for anti-HBc and 354 (70.5 %) were found positive implying previous exposure to HBV and 19 (5.4 %) of the 354 anti-HBc positive had HBV DNA signifying occult HBV infection. No risk factors were found to be associated with the presence of HBV DNA among those who tested positive. Occult HBV infection exists in blood donors in Ile-Ife, Nigeria and the use of HBsAg alone for screening prospective donors will not eliminate the risk of HBV transmission in blood transfusion or stem cell transplantation.

Understanding original antigenic sin in influenza with a dynamical system.

PubMed

Pan, Keyao

2011-01-01

Original antigenic sin is the phenomenon in which prior exposure to an antigen leads to a subsequent suboptimal immune response to a related antigen. Immune memory normally allows for an improved and rapid response to antigens previously seen and is the mechanism by which vaccination works. I here develop a dynamical system model of the mechanism of original antigenic sin in influenza, clarifying and explaining the detailed spin-glass treatment of original antigenic sin. The dynamical system describes the viral load, the quantities of healthy and infected epithelial cells, the concentrations of naïve and memory antibodies, and the affinities of naïve and memory antibodies. I give explicit correspondences between the microscopic variables of the spin-glass model and those of the present dynamical system model. The dynamical system model reproduces the phenomenon of original antigenic sin and describes how a competition between different types of B cells compromises the overall effect of immune response. I illustrate the competition between the naïve and the memory antibodies as a function of the antigenic distance between the initial and subsequent antigens. The suboptimal immune response caused by original antigenic sin is observed when the host is exposed to an antigen which has intermediate antigenic distance to a second antigen previously recognized by the host's immune system.

Serological and molecular diagnostic tests for canine visceral leishmaniasis in Brazilian endemic area: one out of five seronegative dogs are infected.

PubMed

Lopes, E G; Sevá, A P; Ferreira, F; Nunes, C M; Keid, L B; Hiramoto, R M; Ferreira, H L; Oliveira, T M F S; Bigotto, M F D; Galvis-Ovallos, F; Galati, E A B; Soares, R M

2017-09-01

Euthanasia of infected dogs is one of the measures adopted in Brazil to control visceral leishmaniasis (VL) in endemic areas. To detect infected dogs, animals are screened with the rapid test DPP® Visceral Canine Leishmaniasis for detection of antibodies against K26/K39 fusion antigens of amastigotes (DPP). DPP-positives are confirmed with an immunoenzymatic assay probing soluble antigens of promastigotes (ELISA), while DPP-negatives are considered free of infection. Here, 975 dogs from an endemic region were surveyed by using DPP, ELISA and real-time PCR (qPCR) for the diagnosis of VL. When DPP-negative dogs were tested by qPCR applied in blood and lymph node aspirates, 174/887 (19·6%) were positive in at least one sample. In a second sampling using 115 cases, the DPP-negative dogs were tested by qPCR in blood, lymph node and conjunctival swab samples, and 36/79 (45·6%) were positive in at least one sample. Low-to-moderate pairwise agreement was observed between all possible pair of tests. In conclusion, the official diagnosis of VL in dogs in Brazilian endemic areas failed to accuse an expressive number of infected animals and the impact of the low accuracy of serological tests in the success of euthanasia-based measure for VL control need to be assessed.

The utility of rapid antigen detection testing for the diagnosis of streptococcal pharyngitis in low-resource settings.

PubMed

Rimoin, Anne W; Walker, Christa L Fischer; Hamza, Hala S; Elminawi, Nevine; Ghafar, Hadeer Abdel; Vince, Adriana; da Cunha, Antonia L A; Qazi, Shamim; Gardovska, Dace; Steinhoff, Mark C

2010-12-01

To evaluate the utility of rapid antigen detection testing (RADT) for the diagnosis of group A streptococcal (GAS) pharyngitis in pediatric outpatient clinics in four countries with varied socio-economic and geographic profiles. We prospectively evaluated the utility of a commercial RADT in children aged 2-12 years presenting with symptoms of pharyngitis to urban outpatient clinics in Brazil, Croatia, Egypt, and Latvia between August 2001 and December 2005. We compared the performance of the RADT to culture using diagnostic and agreement statistics, including sensitivity, specificity, and positive and negative predictive values. The Centor scores for GAS diagnosis were used to assess the potential effect of spectrum bias on RADT results. Two thousand four hundred and seventy-two children were enrolled at four sites. The prevalence of GAS by throat culture varied by country (range 24.5-39.4%) and by RADT (range 23.9-41.8%). Compared to culture, RADT sensitivity ranged from 72.4% to 91.8% and specificity ranged from 85.7% to 96.4%. The positive predictive value ranged from 67.9% to 88.6% and negative predictive value ranged from 88.1% to 95.7%. In limited-resource regions where microbiological diagnosis is not feasible or practical, RADTs should be considered an option that can be performed in a clinic and provide timely results. Copyright © 2010 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Impact of rapid antigen detection testing on antibiotic prescription in acute pharyngitis in adults. FARINGOCAT STUDY: a multicentric randomized controlled trial

PubMed Central

2010-01-01

Background Acute pharyngitis is one of the most frequent consultations to the general practitioner and in most of the cases an antibiotic is prescribed in primary care in Spain. Bacterial etiology, mainly by group A beta-hemolytic streptococcus (GABHS), accounts for 10-20% of all these infections in adults. The purpose of this study is to assess the impact of rapid antigen detection testing (RADT) to identify GABHS in acute pharyngitis on the utilization of antibiotics in primary care. Methods/design Multicentric randomized controlled trial in which antibiotic prescription between two groups of patients with acute pharyngitis will be compared. The trial will include two arms, a control and an intervention group in which RADT will be performed. The primary outcome measure will be the proportion of inappropriate antibiotic prescription in each group. Two hundred seventy-six patients are required to detect a reduction in antibiotic prescription from 85% in the control group to 75% in the intervention group with a power of 90% and a level of significance of 5%. Secondary outcome measures will be specific antibiotic treatment, antibiotic resistance rates, secondary effects, days without working, medical visits during the first month and patient satisfaction. Discussion The implementation of RADT would allow a more rational use of antibiotics and would prevent adverse effects of antibiotics, emergence of antibiotic resistance and the growth of inefficient health expenses. Trial registration ISRCTN23587778 PMID:20331895

Use of recombinant purified protein derivative (PPD) antigens as specific skin test for tuberculosis.

PubMed

Stavri, Henriette; Bucurenci, Nadia; Ulea, Irina; Costache, Adriana; Popa, Loredana; Popa, Mircea Ioan

2012-11-01

Purified protein derivative (PPD) is currently the only available skin test reagent used worldwide for the diagnosis of tuberculosis (TB). The aim of this study was to develop a Mycobacterium tuberculosis specific skin test reagent, without false positive results due to Bacillus Calmette-Guerin (BCG) vaccination using recombinant antigens. Proteins in PPD IC-65 were analyzed by tandem mass spectrometry and compared to proteins in M. tuberculosis culture filtrate; 54 proteins were found in common. Top candidates MPT64, ESAT 6, and CFP 10 were overexpressed in Escherichia coli expression strains and purified as recombinant proteins. To formulate optimal immunodiagnostic PPD cocktails, the antigens were evaluated by skin testing guinea pigs sensitized with M. tuberculosis H37Rv and BCG. For single antigens and a cocktail mixture of these antigens, best results were obtained using 3 μg/0.1 ml, equivalent to 105 TU (tuberculin units). Each animal was simultaneously tested with PPD IC-65, 2 TU/0.1 ml, as reference. Reactivity of the multi-antigen cocktail was greater than that of any single antigen. The skin test results were between 34.3 and 76.6 per cent the level of reactivity compared to that of the reference when single antigens were tested and 124 per cent the level of reactivity compared to the reference for the multi-antigen cocktail. Our results showed that this specific cocktail could represent a potential candidate for a new skin diagnostic test for TB.

Evaluation of the Clearview® Malaria pLDH Malaria Rapid Diagnostic Test in a non-endemic setting.

PubMed

Houzé, Sandrine; Hubert, Véronique; Cohen, Dorit Pessler; Rivetz, Baruch; Le Bras, Jacques

2011-09-27

Malaria Rapid Diagnostic Tests (RDTs) are widely used to diagnose malaria. The present study evaluated a new RDT, the Clearview® Malaria pLDH test targeting the pan-Plasmodium antigen lactate dehydrogenase (pLDH). The Clearview® Malaria pLDH test was evaluated on fresh samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included samples were Plasmodium falciparum (139), Plasmodium vivax (22), Plasmodium ovale (20), Plasmodium malariae (7), and 102 negative. Overall sensitivity for the detection of Plasmodium spp was 93.2%. For P. falciparum, the sensitivity was 98.6%; for P. vivax, P. ovale and P. malariae, overall sensitivities were 90.9%, 60.0% and 85.7% respectively. For P. falciparum and for P. vivax, the sensitivities increased to 100% at parasite densities above 100/μl. The specificity was 100%. The test was easily to perform and the result was stable for at least 1 hour. The Clearview® Malaria pLDH was efficient for the diagnosis of malaria. The test was very sensitive for P. falciparum and P. vivax detection. The sensitivities for P. ovale and P. malariae were better than other RDTs.

Rapid Tests and the Diagnosis of Visceral Leishmaniasis and Human Immunodeficiency Virus/Acquired Immunodeficiency Syndrome Coinfection.

PubMed

Barbosa Júnior, Walter Lins; Ramos de Araújo, Paulo Sérgio; Dias de Andrade, Luiz; Aguiar Dos Santos, Ana Maria; Lopes da Silva, Maria Almerice; Dantas-Torres, Filipe; Medeiros, Zulma

2015-11-01

After the emergence of the human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS), the number of visceral leishmaniasis (VL)-HIV/AIDS coinfections has increased worldwide. Herein, we assessed the usefulness of an rK39-based immunochromatographic test (rK39 ICT) (DiaMed-IT LEISH(®); DiaMed AG, Cressier-sur-Morat, Switzerland) and a latex agglutination test (KAtex; Kalon Biological, Guildford, United Kingdom) for urinary antigen detection to diagnose VL in 15 HIV/AIDS patients from northeastern Brazil. VL diagnosis was based on clinical findings, cytology, serology, parasite DNA, and/or urinary antigen detection. VL was confirmed in seven out of 15 HIV/AIDS patients. Only three patients were positive in bone marrow cytology, three patients were conventional polymerase chain reaction (PCR) positive, while six were real-time PCR positive. All patients were direct agglutination test (DAT) (Royal Tropical Institute, Amsterdam, The Netherlands) positive; of these, four were positive by rK39 ICT and five by KAtex. Large-scale studies are needed to validate the use of the KAtex in the national public health laboratory network in Brazil, aiming at improving the diagnosis of VL in HIV/AIDS patients in this country. © The American Society of Tropical Medicine and Hygiene.

Application of immunoassay for detection of Helicobacter pylori antigens in the dental plaque.

PubMed

Leszczyńska, K; Namiot, D B; Namiot, Z; Leszczyńska, J K; Jakoniuk, P; Kemona, A

2009-01-01

The aim of this study was to determine the viability of the commercial test currently used for detection of H. pylori antigens in the stool for detection of H. pylori antigens in dental plaque. A total of 164 dyspeptic patients entered the study; 95 H. pylori infected (positive result of at least 4 of 5 diagnostic tests: Campylobacter-like organisms test (CLO test), histology, culture, stool antigens, serology) and 69 noninfected (negative results of 4 diagnostic tests: CLO test, histology, culture, stool antigens). Dental plaque was collected from natural teeth of the patients and incubated in microaerophilic conditions for 72 hours before immunoassay. Experimental findings included that optimal dental plaque weight to perform the examination was over 2 mg and that preliminary incubation increased significantly the number of positive results (p<0.002). It was also found that H. pylori antigens in the dental plaque were positive in 81.2% of infected and only 17.7% of non-infected subjects (p<0.001), while the reproducibility of results was 95%. The immunoassay for detection of H. pylori antigens in the stool may be used, after minor adaptations (specifically pre-incubation in microaerophilic conditions) for H. pylori antigen detection in dental plaque.

Submicron polymer particles containing fluorescent semiconductor nanocrystals CdSe/ZnS for bioassays.

PubMed

Generalova, Alla N; Sizova, Svetlana V; Zdobnova, Tatiana A; Zarifullina, Margarita M; Artemyev, Michail V; Baranov, Alexander V; Oleinikov, Vladimir A; Zubov, Vitaly P; Deyev, Sergey M

2011-02-01

This study aimed to design a panel of uniform particulate biochemical reagents and to test them in specific bioassays. These reagents are polymer particles of different sizes doped with semiconductor nanocrystals and conjugated with either full-size antibodies or recombinant mini-antibodies (4D5 scFv fragment) designed by genetic engineering approaches. A panel of highly fluorescent polymer particles (150-800 nm) were formed by embedding CdSe/ZnS nanocrystals (quantum dots) into preformed polyacrolein and poly(acrolein-co-styrene) particles. Morphology, content and fluorescence characteristics of the prepared materials were studied by laser correlation spectroscopy, spectrophotometry, optical and fluorescent microscopy and fluorimetry. The obtained fluorescent particles sensitized by anti-Yersinia pestis antibodies were used for rapid agglutination glass test suitable for screening analysis of Y. pestis antigen and for microtiter particle agglutination, which, owing to its speed and simplicity, is very beneficial for diagnostic detection of Y. pestis antigen. Recombinant 4D5 scFv antibodies designed and conjugated with polymer particles containing quantum dots provide multipoint highly specific binding with cancer marker HER2/neu on the surface of SKOV-3 cell.

Changes in leucocyte migration after renal transplantation

PubMed Central

Smith, M. G. M.; Eddleston, A. L. W. F.; Dominguez, J. A.; Evans, D. B.; Bewick, M.; Williams, Roger

1969-01-01

The leucocyte migration test, an in-vitro measure of cellular immunity, has been used to follow the changes in cell-mediated hypersensitivity to kidney and histocompatibility antigens in three patients after renal transplantation. Inhibition of leucocyte migration, indicating strong sensitization to the antigens used, occurred in each patient, starting five to seven days after transplantation. Satisfactory renal function had not been established in any of the patients at this time. In one case inhibition of leucocyte migration persisted almost continuously until the 24th day and was associated with poor renal function proved histologically to be due to rejection. Treatment with increased dosage of prednisone was associated with a rapid reversion to normal of the migration index and improvement in renal function. Later, inhibition of migration occurred again, and shortly afterwards the graft ceased to function. In the other two cases the migration index became normal without alteration in immunosuppressive therapy and a satisfactory diuresis followed. It is suggested that this simple test should prove useful in the specific diagnosis of rejection and in control of immunosuppressive therapy. ImagesFig. 3Fig. 4 PMID:4899455

Long-term human serum antibody responses after immunization with whole-cell pertussis vaccine in France.

PubMed Central

Grimprel, E; Bégué, P; Anjak, I; Njamkepo, E; François, P; Guiso, N

1996-01-01

Three hundred sixty children were tested for pertussis serology 0.5 to 1.58 months after complete whole-cell pertussis vaccination. An immunoblot assay was used to detect serum antibodies to pertussis toxin, filamentous hemagglutinin, adenylate cyclase-hemolysin, and pertactin, and agglutination was used for detection of anti-agglutinogen antibodies. Antibodies against pertussis toxin, pertactin, and agglutinogens decreased rapidly after vaccination but increased secondarily, suggesting exposure to infected persons. In contrast, anti-filamentous hemagglutinin antibodies persisted and anti-adenylate cyclase-hemolysin antibodies increased continuously, suggesting either cross-reaction with non-Bordetella antigens or exposure to Bordetella isolates expressing these two antigens, including Bordetella pertussis. These data suggest that unrecognized pertussis is common in France despite massive and sustained immunization in infants and that vaccinated children become susceptible to infection more than 6 years after their last vaccination. PMID:8770511

21 CFR 660.4 - Potency test.

Code of Federal Regulations, 2012 CFR

2012-04-01

... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Antibody to Hepatitis B Surface Antigen § 660.4 Potency test. To be satisfactory for release, each filling of Antibody to Hepatitis B Surface Antigen shall be tested against the Reference Hepatitis B Surface Antigen Panel and shall be sufficiently potent...

21 CFR 660.4 - Potency test.

Code of Federal Regulations, 2011 CFR

2011-04-01

... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Antibody to Hepatitis B Surface Antigen § 660.4 Potency test. To be satisfactory for release, each filling of Antibody to Hepatitis B Surface Antigen shall be tested against the Reference Hepatitis B Surface Antigen Panel and shall be sufficiently potent...

21 CFR 660.4 - Potency test.

Code of Federal Regulations, 2014 CFR

2014-04-01

... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Antibody to Hepatitis B Surface Antigen § 660.4 Potency test. To be satisfactory for release, each filling of Antibody to Hepatitis B Surface Antigen shall be tested against the Reference Hepatitis B Surface Antigen Panel and shall be sufficiently potent...

21 CFR 660.4 - Potency test.

Code of Federal Regulations, 2010 CFR

2010-04-01

... STANDARDS FOR DIAGNOSTIC SUBSTANCES FOR LABORATORY TESTS Antibody to Hepatitis B Surface Antigen § 660.4 Potency test. To be satisfactory for release, each filling of Antibody to Hepatitis B Surface Antigen shall be tested against the Reference Hepatitis B Surface Antigen Panel and shall be sufficiently potent...

A scalable method for O-antigen purification applied to various Salmonella serovars

PubMed Central

Micoli, F.; Rondini, S.; Gavini, M.; Pisoni, I.; Lanzilao, L.; Colucci, A.M.; Giannelli, C.; Pippi, F.; Sollai, L.; Pinto, V.; Berti, F.; MacLennan, C.A.; Martin, L.B.; Saul, A.

2014-01-01

The surface lipopolysaccharide of gram-negative bacteria is both a virulence factor and a B cell antigen. Antibodies against O-antigen of lipopolysaccharide may confer protection against infection, and O-antigen conjugates have been designed against multiple pathogens. Here, we describe a simplified methodology for extraction and purification of the O-antigen core portion of Salmonella lipopolysaccharide, suitable for large-scale production. Lipopolysaccharide extraction and delipidation are performed by acetic acid hydrolysis of whole bacterial culture and can take place directly in a bioreactor, without previous isolation and inactivation of bacteria. Further O-antigen core purification consists of rapid filtration and precipitation steps, without using enzymes or hazardous chemicals. The process was successfully applied to various Salmonella enterica serovars (Paratyphi A, Typhimurium, and Enteritidis), obtaining good yields of high-quality material, suitable for conjugate vaccine preparations. PMID:23142430

Screening Yield of HIV Antigen/Antibody Combination and Pooled HIV RNA Testing for Acute HIV Infection in a High-Prevalence Population.

PubMed

Peters, Philip J; Westheimer, Emily; Cohen, Stephanie; Hightow-Weidman, Lisa B; Moss, Nicholas; Tsoi, Benjamin; Hall, Laura; Fann, Charles; Daskalakis, Demetre C; Beagle, Steve; Patel, Pragna; Radix, Asa; Foust, Evelyn; Kohn, Robert P; Marmorino, Jenni; Pandori, Mark; Fu, Jie; Samandari, Taraz; Gay, Cynthia L

2016-02-16

Although acute HIV infection contributes disproportionately to onward HIV transmission, HIV testing has not routinely included screening for acute HIV infection. To evaluate the performance of an HIV antigen/antibody (Ag/Ab) combination assay to detect acute HIV infection compared with pooled HIV RNA testing. Multisite, prospective, within-individual comparison study conducted between September 2011 and October 2013 in 7 sexually transmitted infection clinics and 5 community-based programs in New York, California, and North Carolina. Participants were 12 years or older and seeking HIV testing, without known HIV infection. All participants with a negative rapid HIV test result were screened for acute HIV infection with an HIV Ag/Ab combination assay (index test) and pooled human immunodeficiency virus 1 (HIV-1) RNA testing. HIV RNA testing was the reference standard, with positive reference standard result defined as detectable HIV-1 RNA on an individual RNA test. Number and proportion with acute HIV infections detected. Among 86,836 participants with complete test results (median age, 29 years; 75.0% men; 51.8% men who have sex with men), established HIV infection was diagnosed in 1158 participants (1.33%) and acute HIV infection was diagnosed in 168 participants (0.19%). Acute HIV infection was detected in 134 participants with HIV Ag/Ab combination testing (0.15% [95% CI, 0.13%-0.18%]; sensitivity, 79.8% [95% CI, 72.9%-85.6%]; specificity, 99.9% [95% CI, 99.9%-99.9%]; positive predictive value, 59.0% [95% CI, 52.3%-65.5%]) and in 164 participants with pooled HIV RNA testing (0.19% [95% CI, 0.16%-0.22%]; sensitivity, 97.6% [95% CI, 94.0%-99.4%]; specificity, 100% [95% CI, 100%-100%]; positive predictive value, 96.5% [95% CI, 92.5%-98.7%]; sensitivity comparison, P < .001). Overall HIV Ag/Ab combination testing detected 82% of acute HIV infections detectable by pooled HIV RNA testing. Compared with rapid HIV testing alone, HIV Ag/Ab combination testing increased the relative HIV diagnostic yield (both established and acute HIV infections) by 10.4% (95% CI, 8.8%-12.2%) and pooled HIV RNA testing increased the relative HIV diagnostic yield by 12.4% (95% CI, 10.7%-14.3%). In a high-prevalence population, HIV screening using an HIV Ag/Ab combination assay following a negative rapid test detected 82% of acute HIV infections detectable by pooled HIV RNA testing, with a positive predictive value of 59%. Further research is needed to evaluate this strategy in lower-prevalence populations and in persons using preexposure prophylaxis for HIV prevention.

Strategies for the prevention of perinatal hepatitis B transmission in a marginalized population on the Thailand-Myanmar border: a cost-effectiveness analysis.

PubMed

Devine, Angela; Harvey, Rebecca; Min, Aung Myat; Gilder, Mary Ellen T; Paw, Moo Koh; Kang, Joy; Watts, Isabella; Hanboonkunupakarn, Borimas; Nosten, François; McGready, Rose

2017-08-09

Data on the cost effectiveness of hepatitis B virus (HBV) screening and vaccination strategies for prevention of vertical transmission of HBV in resource limited settings is sparse. A decision tree model of HBV prevention strategies utilised data from a cohort of 7071 pregnant women on the Thailand-Myanmar border using a provider perspective. All options included universal HBV vaccination for newborns in three strategies: (1) universal vaccination alone; (2) universal vaccination with screening of women during antenatal visits with rapid diagnostic test (RDT) plus HBV immune globulin (HBIG) administration to newborns of HBV surface antigen positive women; and (3) universal vaccination with screening of women during antenatal visits plus HBIG administration to newborns of women testing HBV e antigen positive by confirmatory test. At the time of the study, the HBIG after confirmatory test strategy was used. The costs in United States Dollars (US$), infections averted and incremental cost effectiveness ratios (ICERs) were calculated and sensitivity analyses were conducted. A willingness to pay threshold of US$1200 was used. The universal HBV vaccination was the least costly option at US$4.33 per woman attending the clinic. The HBIG after (RDT) strategy had an ICER of US$716.78 per infection averted. The HBIG after confirmatory test strategy was not cost-effective due to extended dominance. The one-way sensitivity analysis showed that while the transmission parameters and cost of HBIG had the biggest impact on outcomes, the HBIG after confirmatory test only became a cost-effective option when a low test cost was used or a high HBIG cost was used. The probabilistic sensitivity analysis showed that HBIG after RDT had an 87% likelihood of being cost-effective as compared to vaccination only at a willingness to pay threshold of US$1200. HBIG following confirmatory test is not a cost-effective strategy for preventing vertical transmission of HBV in the Thailand-Myanmar border population. By switching to HBIG following rapid diagnostic test, perinatal infections will be reduced by nearly one third. This strategy may be applicable to similar settings for marginalized populations where the confirmatory test is not logistically possible.

Detection of hepatitis A viral antigen by radioimmunoassay

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hollinger, F.B.; Bradley, D.W.; Maynard, J.E.

1975-11-01

With coded samples, the effectiveness and specificity of a micro-SPIRA procedure for rapidly and quantitatively detecting type A hepatitis-associated antigen in large numbers of specimens from infected liver, stool, or serum has been demonstrated. Samples which were judged to be negative by IEM were found to contain significant levels of HAV antigen by this immunoradiometric technique. The detection of significant levels of HAV antigen in infected chimpanzees supports epidemiologic evidence of viremia during the acute stage of the disease. The results of this study suggest that the diagnosis of type A hepatitis by a convention serologic procedure may now bemore » at hand. (auth)« less

CD1d-restricted immunoglobulin G formation to GPI-anchored antigens mediated by NKT cells.

PubMed

Schofield, L; McConville, M J; Hansen, D; Campbell, A S; Fraser-Reid, B; Grusby, M J; Tachado, S D

1999-01-08

Immunoglobulin G (IgG) responses require major histocompatibility complex (MHC)-restricted recognition of peptide fragments by conventional CD4(+) helper T cells. Immunoglobulin G responses to glycosylphosphatidylinositol (GPI)- anchored protein antigens, however, were found to be regulated in part through CD1d-restricted recognition of the GPI moiety by thymus-dependent, interleukin-4-producing CD4(+), natural killer cell antigen 1.1 [(NK1.1)+] helper T cells. The CD1-NKT cell pathway regulated immunogobulin G responses to the GPI-anchored surface antigens of Plasmodium and Trypanosoma and may be a general mechanism for rapid, MHC-unrestricted antibody responses to diverse pathogens.

Antibodies to dendritic neuronal surface antigens in opsoclonus myoclonus ataxia syndrome

PubMed Central

Panzer, Jessica A.; Anand, Ronan; Dalmau, Josep; Lynch, David R.

2015-01-01

Opsoclonus myoclonus ataxia syndrome (OMAS) is an autoimmune disorder characterized by rapid, random, conjugate eye movements (opsoclonus), myoclonus, and ataxia. Given these symptoms, autoantibodies targeting the cerebellum or brainstem could mediate the disease or be markers of autoimmunity. In a subset of patients with OMAS, we identified such autoantibodies, which bind to non-synaptic puncta on the surface of live cultured cerebellar and brainstem neuronal dendrites. These findings implicate autoimmunity to a neuronal surface antigen in the pathophysiology of OMAS. Identification of the targeted antigen(s) could elucidate the mechanisms underlying OMAS and provide a biomarker for diagnosis and response to therapy. PMID:26298330

Heat treatment and false-positive heartworm antigen testing in ex vivo parasites and dogs naturally infected by Dirofilaria repens and Angiostrongylus vasorum.

PubMed

Venco, Luigi; Manzocchi, Simone; Genchi, Marco; Kramer, Laura H

2017-11-09

Heartworm antigen testing is considered sensitive and specific. Currently available tests are reported as detecting a glycoprotein found predominantly in the reproductive tract of the female worm and can reach specificity close to 100%. Main concerns regard sensitivity in the case of light infections, the presence of immature females or cases of all-male infections. Research and development have been aimed at increasing sensitivity. Recently, heat treatment of serum prior to antigen testing has been shown to result in an increase in positive antigen test results, presumably due to disruption of natural antigen-antibody complexes. Cross-reactions in dogs with both natural and experimental infections with Angiostrongylus vasorum and Spirocerca lupi have been reported, but cross-reactions with other helminths have not been extensively studied. In order to evaluate potential cross-reactivity with other canine and feline parasites, two studies were performed. Study 1: Live adults of Dirofilaria immitis, Dirofilaria repens, Toxocara canis, Toxocara cati, Dipylidium caninum, Taenia taeniaeformis and Mesocestoides spp. larvae were washed and incubated in tubes with saline solution. All worms were alive at the time of removal from the saline. Saline solutions containing excretory/secretory antigens were then tested for heartworm with six different, commercially available antigen tests. All results were evaluated blind by three of the authors. Study 2: Sera from dogs with natural infections by A. vasorum or D. repens, living in areas free of heartworm disease, were tested with the same tests before and after heat treatment (103 °C for 10 min). Results suggest that antigens detected by currently available tests are not specific for D. immitis. They may give positive results through detection of different parasites' antigens that are normally not released into the bloodstream or released in a low amount and/or bound to antibodies. Tests may even detect antigens released by male D. immitis adult worms. D. repens appears to release more detectable antigens than the other worms studied. Cross-reaction with A. vasorum and D. repens does occur in the field and could potentially occur with other helminths. Heat treatment decreases specificity by enhancing cross-reactivity.

Rapid identification of serotypes of Mycobacterium avium-M. intracellulare complex by using infected swine sera and reference antigenic glycolipids.

PubMed

Ikawa, H; Oka, S; Murakami, H; Hayashi, A; Yano, I

1989-11-01

The species of 136 strains of acid-fast bacteria isolated from swine with mycobacteriosis were identified by numerical taxonomy and chemotaxonomy on the basis of mycolic acid subclass composition as members of the Mycobacterium avium-M. intracellulare (MAI) complex. The isolates were further classified by using both thin-layer chromatography of the antigenic glycopeptidolipids (GPL) obtained from the bacteria by the method of Tsang et al. (A. Y. Tsang, I. Drupa, M. Goldberg, J. K. McClatchy, and P. J. Brennan, Int. J. Syst. Bacteriol. 33:285-292, 1983) and the seroagglutination test devised by W. B. Schaefer (Am. Rev. Respir. Dis. 92[Suppl.]:85-93, 1965). For the reference standard, purified antigenic GPL of serotypes 4, 8, and 9 were isolated and their structures were analyzed by negative fast-atom bombardment-mass spectrometry. The fast-atom bombardment-mass spectrometric spectra of the intact GPL antigens of serotypes 4, 8, and 9 agreed with the structures reported earlier by Brennan et al. (P. J. Brennan and M. B. Goren, J. Biol. Chem. 254:4205-4211, 1979; P. J. Brennan, G. O. Aspinall, and J. E. Nam Shin, J. Biol. Chem. 256:6817-6822, 1981). With these antigenic GPL, the thin-layer chromatographic behaviors of the alkali-stable lipids of the above-described isolates were examined. These MAI complex isolates fell into the serotype 8 (85 strains), 4 (33 strains), and 9 (7 strains) and untypeable (11 strains) categories. Furthermore, an enzyme-linked immunosorbent assay (ELISA) based on type-specific glycolipid antigens and infected swine sera was used to diagnose the serological types of the MAI complex isolates. Of 14 cases typed by both the seroagglutination reaction and thin-layer chromatography, 13 showed clear agreement with the ELISA results. The results demonstrated that ELISA using infected sera was especially useful, and it can be recommended on the basis of simplicity, sensitivity, and specificity as an adjunct to the seroaggulutination test and thin-layer chromatography for identification of mycobacteria belonging to the MAI complex.

Comparison of a newly developed automated and quantitative hepatitis C virus (HCV) core antigen test with the HCV RNA assay for clinical usefulness in confirming anti-HCV results.

PubMed

Kesli, Recep; Polat, Hakki; Terzi, Yuksel; Kurtoglu, Muhammet Guzel; Uyar, Yavuz

2011-12-01

Hepatitis C virus (HCV) is a global health care problem. Diagnosis of HCV infection is mainly based on the detection of anti-HCV antibodies as a screening test with serum samples. Recombinant immunoblot assays are used as supplemental tests and for the final detection and quantification of HCV RNA in confirmatory tests. In this study, we aimed to compare the HCV core antigen test with the HCV RNA assay for confirming anti-HCV results to determine whether the HCV core antigen test may be used as an alternative confirmatory test to the HCV RNA test and to assess the diagnostic values of the total HCV core antigen test by determining the diagnostic specificity and sensitivity rates compared with the HCV RNA test. Sera from a total of 212 treatment-naive patients were analyzed for anti-HCV and HCV core antigen both with the Abbott Architect test and with the molecular HCV RNA assay consisting of a reverse transcription-PCR method as a confirmatory test. The diagnostic sensitivity, specificity, and positive and negative predictive values of the HCV core antigen assay compared to the HCV RNA test were 96.3%, 100%, 100%, and 89.7%, respectively. The levels of HCV core antigen showed a good correlation with those from the HCV RNA quantification (r = 0.907). In conclusion, the Architect HCV antigen assay is highly specific, sensitive, reliable, easy to perform, reproducible, cost-effective, and applicable as a screening, supplemental, and preconfirmatory test for anti-HCV assays used in laboratory procedures for the diagnosis of hepatitis C virus infection.

Stabilizing short-lived Schiff base derivatives of 5-aminouracils that activate mucosal-associated invariant T cells

NASA Astrophysics Data System (ADS)

Mak, Jeffrey Y. W.; Xu, Weijun; Reid, Robert C.; Corbett, Alexandra J.; Meehan, Bronwyn S.; Wang, Huimeng; Chen, Zhenjun; Rossjohn, Jamie; McCluskey, James; Liu, Ligong; Fairlie, David P.

2017-03-01

Mucosal-associated invariant T (MAIT) cells are activated by unstable antigens formed by reactions of 5-amino-6-D-ribitylaminouracil (a vitamin B2 biosynthetic intermediate) with glycolysis metabolites such as methylglyoxal. Here we show superior preparations of antigens in dimethylsulfoxide, avoiding their rapid decomposition in water (t1/2 1.5 h, 37 °C). Antigen solution structures, MAIT cell activation potencies (EC50 3-500 pM), and chemical stabilities are described. Computer analyses of antigen structures reveal stereochemical and energetic influences on MAIT cell activation, enabling design of a water stable synthetic antigen (EC50 2 nM). Like native antigens, this antigen preparation induces MR1 refolding and upregulates surface expression of human MR1, forms MR1 tetramers that detect MAIT cells in human PBMCs, and stimulates cytokine expression (IFNγ, TNF) by human MAIT cells. These antigens also induce MAIT cell accumulation in mouse lungs after administration with a co-stimulant. These chemical and immunological findings provide new insights into antigen properties and MAIT cell activation.

Dynamics of T cell, antigen-presenting cell, and pathogen interactions during recall responses in the lymph node.

PubMed

Chtanova, Tatyana; Han, Seong-Ji; Schaeffer, Marie; van Dooren, Giel G; Herzmark, Paul; Striepen, Boris; Robey, Ellen A

2009-08-21

Memory T cells circulate through lymph nodes where they are poised to respond rapidly upon re-exposure to a pathogen; however, the dynamics of memory T cell, antigen-presenting cell, and pathogen interactions during recall responses are largely unknown. We used a mouse model of infection with the intracellular protozoan parasite, Toxoplasma gondii, in conjunction with two-photon microscopy, to address this question. After challenge, memory T cells migrated more rapidly than naive T cells, relocalized toward the subcapsular sinus (SCS) near invaded macrophages, and engaged in prolonged interactions with infected cells. Parasite invasion of T cells occurred by direct transfer of the parasite from the target cell into the T cell and corresponded to an antigen-specific increase in the rate of T cell invasion. Our results provide insight into cellular interactions during recall responses and suggest a mechanism of pathogen subversion of the immune response.

Induction of Robust Immune Responses in Swine by Using a Cocktail of Adenovirus-Vectored African Swine Fever Virus Antigens.

PubMed

Lokhandwala, Shehnaz; Waghela, Suryakant D; Bray, Jocelyn; Martin, Cameron L; Sangewar, Neha; Charendoff, Chloe; Shetti, Rashmi; Ashley, Clay; Chen, Chang-Hsin; Berghman, Luc R; Mwangi, Duncan; Dominowski, Paul J; Foss, Dennis L; Rai, Sharath; Vora, Shaunak; Gabbert, Lindsay; Burrage, Thomas G; Brake, David; Neilan, John; Mwangi, Waithaka

2016-11-01

The African swine fever virus (ASFV) causes a fatal hemorrhagic disease in domestic swine, and at present no treatment or vaccine is available. Natural and gene-deleted, live attenuated strains protect against closely related virulent strains; however, they are yet to be deployed and evaluated in the field to rule out chronic persistence and a potential for reversion to virulence. Previous studies suggest that antibodies play a role in protection, but induction of cytotoxic T lymphocytes (CTLs) could be the key to complete protection. Hence, generation of an efficacious subunit vaccine depends on identification of CTL targets along with a suitable delivery method that will elicit effector CTLs capable of eliminating ASFV-infected host cells and confer long-term protection. To this end, we evaluated the safety and immunogenicity of an adenovirus-vectored ASFV (Ad-ASFV) multiantigen cocktail formulated in two different adjuvants and at two immunizing doses in swine. Immunization with the cocktail rapidly induced unprecedented ASFV antigen-specific antibody and cellular immune responses against all of the antigens. The robust antibody responses underwent rapid isotype switching within 1 week postpriming, steadily increased over a 2-month period, and underwent rapid recall upon boost. Importantly, the primed antibodies strongly recognized the parental ASFV (Georgia 2007/1) by indirect fluorescence antibody (IFA) assay and Western blotting. Significant antigen-specific gamma interferon-positive (IFN-γ + ) responses were detected postpriming and postboosting. Furthermore, this study is the first to demonstrate induction of ASFV antigen-specific CTL responses in commercial swine using Ad-ASFV multiantigens. The relevance of the induced immune responses in regard to protection needs to be evaluated in a challenge study. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Induction of Robust Immune Responses in Swine by Using a Cocktail of Adenovirus-Vectored African Swine Fever Virus Antigens

PubMed Central

Waghela, Suryakant D.; Bray, Jocelyn; Martin, Cameron L.; Sangewar, Neha; Charendoff, Chloe; Shetti, Rashmi; Ashley, Clay; Chen, Chang-Hsin; Berghman, Luc R.; Mwangi, Duncan; Dominowski, Paul J.; Foss, Dennis L.; Rai, Sharath; Vora, Shaunak; Gabbert, Lindsay; Burrage, Thomas G.; Brake, David; Neilan, John

2016-01-01

The African swine fever virus (ASFV) causes a fatal hemorrhagic disease in domestic swine, and at present no treatment or vaccine is available. Natural and gene-deleted, live attenuated strains protect against closely related virulent strains; however, they are yet to be deployed and evaluated in the field to rule out chronic persistence and a potential for reversion to virulence. Previous studies suggest that antibodies play a role in protection, but induction of cytotoxic T lymphocytes (CTLs) could be the key to complete protection. Hence, generation of an efficacious subunit vaccine depends on identification of CTL targets along with a suitable delivery method that will elicit effector CTLs capable of eliminating ASFV-infected host cells and confer long-term protection. To this end, we evaluated the safety and immunogenicity of an adenovirus-vectored ASFV (Ad-ASFV) multiantigen cocktail formulated in two different adjuvants and at two immunizing doses in swine. Immunization with the cocktail rapidly induced unprecedented ASFV antigen-specific antibody and cellular immune responses against all of the antigens. The robust antibody responses underwent rapid isotype switching within 1 week postpriming, steadily increased over a 2-month period, and underwent rapid recall upon boost. Importantly, the primed antibodies strongly recognized the parental ASFV (Georgia 2007/1) by indirect fluorescence antibody (IFA) assay and Western blotting. Significant antigen-specific gamma interferon-positive (IFN-γ+) responses were detected postpriming and postboosting. Furthermore, this study is the first to demonstrate induction of ASFV antigen-specific CTL responses in commercial swine using Ad-ASFV multiantigens. The relevance of the induced immune responses in regard to protection needs to be evaluated in a challenge study. PMID:27628166

Indirect Competitive Enzyme-Linked Immunosorbent Assay (ELISA).

PubMed

Kohl, Thomas O; Ascoli, Carl A

2017-07-05

The indirect competitive ELISA (indirect cELISA) pits plate-immobilized antigen against antigens in solution for binding to antigen-specific antibody. The antigens in solution are in the test sample and are first incubated with antigen-specific antibody. These antibody-antigen complexes are then added to microtiter plates whose wells have been coated with purified antigen. The wells are washed to remove unbound antigen-antibody complexes and free antigen. A reporter-labeled secondary antibody is then added followed by the addition of substrate. Substrate hydrolysis yields a signal that is inversely proportional to antigen concentration within the sample. This is because when antigen concentration is high in the test sample, most of the antibody is bound before adding the solution to the plate. Most of the antibody remains in solution (as complexes) and is thus washed away before the addition of the reporter-labeled secondary antibody and substrate. Thus, the higher the antigen concentration in the test sample, the weaker the resultant signal in the detection step. The indirect cELISA is often used for competitive detection and quantification of antibodies against viral diseases in biological samples. © 2017 Cold Spring Harbor Laboratory Press.

Latex agglutination test (LAT) for the diagnosis of typhoid fever.

PubMed

Sahni, Gopal Shankar

2013-06-01

The efficacy of latex agglutination test in the rapid diagnosis of typhoid fever was studied and the result compared with that of blood culture. This study included 80 children suffering from typhoid fever, among which 40 were confirmed by blood culture isolation and 40 had possible typhoid fever based on high Widal's titre (a four-fold rise in the titre of antibody to typhi "O" and "H" antigen was considered as a positive Widal's test result). Eighty children, 40 with febrile illness confirmed to be other than typhoid and 40 normal healthy children were used as negative controls. The various groups were: (i) Study group ie, group I had 40 children confirmed by culture isolation of Salmonella typhi(confirmed typhoid cases). (ii) Control groups ie, (a) group II with 40 febrile controls selected from paediatrics ward where cause other than S typhi has been established, (b) group III with 40 afebrile healthy controls that were siblings of the children admitted in paediatric ward for any reason with no history of fever and TAB vaccination in the last one year, and (c) group IV with 40 children with high Widal's titre in paired sera sample. Widal's test with paired sera with a one week interval between collections were done in all 40 patients. Latex aggtutination test which could detect 900 ng/ml of antigen as observed in checker board titration, was positive in all 40 children from group I who had positive blood culture and in 30 children from group IV who had culture negative and had high Widal's titre positive. Latex agglutination test was positive in 4 children in group II and none in group III. Using blood culture positive cases as true positive and children in groups II and III as true negative, the test had a sensitivity of 100% and specificity of 96%. Latex agglutination test was found to be significantly sensitive (100%) and specific (96%) and could detect 75% more cases in group IV (possible typhoid cases). Thus latex agglutination test can be used for rapid diagnosis of typhoid fever though it cannot replace conventional blood culture required for isolation of organism to report the antibiotic sensitivity.

Crypto-Giardia antigen rapid test versus conventional modified Ziehl-Neelsen acid fast staining method for diagnosis of cryptosporidiosis.

PubMed

Zaglool, Dina Abdulla Muhammad; Mohamed, Amr; Khodari, Yousif Abdul Wahid; Farooq, Mian Usman

2013-03-01

To evaluate the validity of Crypto-Giardia antigen rapid test (CA-RT) in comparison with the conventional modified Ziehl-Neelsen acid fast (MZN-AF) staining method for the diagnosis of cryptosporidiosis. Fifteen preserved stool samples from previously confirmed infections were used as positive controls and 40 stool samples from healthy people were used as negative control. A total of 85 stool samples were collected from suspected patients with cryptosporidiosis over 6 months during the period from January till June, 2011. The study was conducted in the department of parasitology, central laboratory, Alnoor Specialist Hospital, Makkah, Saudi Arabia. All samples were subjected to CA-RT and conventional MZN-AF staining method. Validation parameters including sensitivity (SN), specificity (SP), accuracy index (AI), positive predictive value (PPV), and negative predictive value (NPV) were evaluated for both tests. Out of 15 positive controls, CA-RT detected 13 (86.7%) while MZN-AF detected 11(73.3%) positive cases. However, CA-RT detected no positive case in 40 normal controls but MZN-AF detected 2(5%) as positive cases. Based on the results, the SN, SP, AI, PPV and NPV were high in CA-RT than MZN-AF staining method, ie., 86.7%vs. 73.3%, 100%vs. 95%, 96.4%vs. 89.1%, 100%vs. 84.6% and 95.2%vs. 90.5%, respectively. Out of a total of 85 suspected specimens, CA-RT detected 7(8.2%) but MZN-AF detected 6(7.1%) cases as positive. CA-RT immunoassay is more valid and reliable than MZN-AF staining method. Copyright © 2013 Hainan Medical College. Published by Elsevier B.V. All rights reserved.

Interpretations of Antibody Responses to Salmonella enterica Serotype Enteritidis gm Flagellin in Poultry Flocks Are Enhanced by a Kinetics-Based Enzyme-Linked Immunosorbent Assay

PubMed Central

McDonough, Patrick L.; Jacobson, Richard H.; Timoney, John F.; Mutalib, Ahmed; Kradel, David C.; Chang, Yung-fu; Shin, Sang J.; Lein, Donald H.; Trock, Susan; Wheeler, Kaye

1998-01-01

Many regulatory and diagnostic programs for the detection of Salmonella enterica serotype Enteritidis infection in commercial poultry flocks have relied on rapid Pullorum agglutination tests to screen birds because of the shared antigens of S. enterica Enteritidis and S. enterica Pullorum and Gallinarum; however, the use of the enzyme-linked immunosorbent assay (ELISA) format affords better analytical sensitivity than crude agglutination tests. In this study, we adapted our earlier conventional indirect ELISA, using gm flagellin as the antigen, to a kinetics-based, computer-controlled ELISA (KELA). The KELA was used to screen for flagellin antibody from three commercial flocks: (i) a large flock involved in a U.S. Department of Agriculture trace back from a human S. enterica Enteritidis foodborne outbreak (n = 3,209), (ii) a flock infected with the endemic S. enterica Enteritidis serotype but which also had multiple other salmonella serotypes (n = 65), and (iii) an S. enterica Pullorum-infected flock (n = 12). The first flock (S. enterica Enteritidis prevalence of 2.45% based on culture) provided a field test of the KELA and allowed the calculation of diagnostic sensitivity (D-Sn) and diagnostic specificity (D-Sp). With a cutoff of 10 (used for screening flocks [i.e., high sensitivity]), the KELA has a D-Sn of 95.2% and a D-Sp of 18.5%; with a cutoff of 140 (used in confirmatory flock testing [i.e., high specificity]), the KELA has a D-Sn of 28.0% and a D-Sp of 99.1%. We found that with a cutoff of 60 (D-Sn = 63.1%; D-Sp = 91.6%), we could eliminate reactions in the KELA caused by other non-S. enterica Enteritidis salmonellae. The KELA was also compared to two commercial rapid Pullorum tests, the Solvay (D-Sn = 94.9%; D-Sp = 55.5%) and the Vineland (D-Sn = 62.0%; D-Sp = 75.3%). PMID:9665965

Epidemic outbreaks, diagnostics, and control measures of the H5N1 highly pathogenic avian influenza in the Kingdom of Saudi Arabia, 2007-08.

PubMed

Lu, Huaguang; Ismail, Mahmoud Moussa; Khan, Owais Ahmed; Al Hammad, Yousef; Abdel Rhman, Salah Shaban; Al-Blowi, Mohamed Hamad

2010-03-01

The first outbreak of H5N1 highly pathogenic avian influenza (HPAI) in the Kingdom of Saudi Arabia (KSA) occurred in two "backyard" flocks of Houbara bustards and falcons in February 2007. Subsequent outbreaks were seen through the end of 2007 in "backyard" birds including native chickens, ostriches, turkeys, ducks, and peacocks. From November 2007 through January 2008, H5N1 HPAI outbreaks occurred in 19 commercial poultry premises, including two broiler breeder farms, one layer breeder farm, one ostrich farm, and 15 commercial layer farms, with approximately 4.75 million birds affected. Laboratory diagnosis of all H5N1-positive cases was conducted at the Central Veterinary Diagnostic Laboratory (CVDL) in Riyadh, Saudi Arabia. A combination of diagnostic tests was used to confirm the laboratory diagnosis. A rapid antigen-capture test and real-time reverse transcriptase-PCR (rtRT-PCR) assay on clinical and field specimens were conducted initially. Meanwhile, virus isolation in specific-pathogen-free embryonating chicken eggs was performed and was followed by hemagglutinin (HA) and hemagglutination inhibition tests, then rapid antigen-capture and rtRT-PCR tests on HA-positive allantoic fluid samples. In most HPAI cases, a complete laboratory diagnosis was made within 24-48 hr at the CVDL. Saudi Arabian government officials made immediate decisions to depopulate all H5N1-affected and nonaffected flocks within a 5-km radius area and applied quarantine zones to prevent the virus from spreading to other areas. Other control measures, such as closure of live bird markets and intensive surveillance tests on all poultry species within quarantine zones, were in place during the outbreaks. As a result, the HPAI outbreaks were quickly controlled, and no positive cases were detected after January 29, 2008. The KSA was declared free of HPAI on April 30, 2008, by the World Animal Health Organization.

Detection of H5N1 high-pathogenicity avian influenza virus in meat and tracheal samples from experimentally infected chickens.

PubMed

Das, Amaresh; Spackman, Erica; Thomas, Colleen; Swayne, David E; Suarez, David L

2008-03-01

The Asian H5N1 highly pathogenic avian influenza (HPAI) virus causes a systemic disease with high mortality of poultry and is potentially zoonotic. In both chickens and ducks, the virus has been demonstrated to replicate in both cardiac and skeletal muscle cells. Experimentally, H5N1 HPAI virus has been transmitted to chickens through the consumption of raw infected meat. In this study, we investigated virus replication in cardiac and skeletal muscle and in the trachea of chickens after experimental intranasal inoculation with the H5N1 HPAI virus. The virus was detected in tissues by real-time reverse transcription-polymerase chain reaction (RRT-PCR) and virus isolation, and in the trachea by RRT-PCR and a commercial avian influenza (AI) viral antigen detection test. A modified RNA extraction protocol was developed for rapid detection of the virus in tissues by RRT-PCR. The H5N1 HPAI virus was sporadically detected in meat and the tracheas of infected birds without any clinical sign of disease as early as 6 hr postinfection (PI), and was detected in all samples tested at 24 hr PI and later. No differences in sensitivity were seen between virus isolation and RRT-PCR in meat samples. The AI viral antigen detection test on tracheal swabs was a useful method for identifying infected chickens when they were sick or dead, but was less sensitive in detecting infected birds when they were preclinical. This study provides data indicating that preslaughter tracheal swab testing can identify birds infected with HPAI among the daily mortality and prevent infected flocks from being sent to processing plants. In addition, the modified RNA extraction and RRT-PCR test on meat samples provide a rapid and sensitive method of identifying HPAI virus in illegal contraband or domestic meat samples.

Interpretations of antibody responses to Salmonella enterica serotype enteritidis gm flagellin in poultry flocks are enhanced by a kinetics-based enzyme-linked immunosorbent assay.

PubMed

McDonough, P L; Jacobson, R H; Timoney, J F; Mutalib, A; Kradel, D C; Chang, Y F; Shin, S J; Lein, D H; Trock, S; Wheeler, K

1998-07-01

Many regulatory and diagnostic programs for the detection of Salmonella enterica serotype Enteritidis infection in commercial poultry flocks have relied on rapid Pullorum agglutination tests to screen birds because of the shared antigens of S. enterica Enteritidis and S. enterica Pullorum and Gallinarum; however, the use of the enzyme-linked immunosorbent assay (ELISA) format affords better analytical sensitivity than crude agglutination tests. In this study, we adapted our earlier conventional indirect ELISA, using gm flagellin as the antigen, to a kinetics-based, computer-controlled ELISA (KELA). The KELA was used to screen for flagellin antibody from three commercial flocks: (i) a large flock involved in a U.S. Department of Agriculture trace back from a human S. enterica Enteritidis foodborne outbreak (n = 3,209), (ii) a flock infected with the endemic S. enterica Enteritidis serotype but which also had multiple other Salmonella serotypes (n = 65), and (iii) an S. enterica Pullorum-infected flock (n = 12). The first flock (S. enterica Enteritidis prevalence of 2.45% based on culture) provided a field test of the KELA and allowed the calculation of diagnostic sensitivity (D-Sn) and diagnostic specificity (D-Sp). With a cutoff of 10 (used for screening flocks [i.e., high sensitivity]), the KELA has a D-Sn of 95.2% and a D-Sp of 18.5%; with a cutoff of 140 (used in confirmatory flock testing [i.e., high specificity]), the KELA has a D-Sn of 28.0% and a D-Sp of 99.1%. We found that with a cutoff of 60 (D-Sn = 63.1%; D-Sp = 91.6%), we could eliminate reactions in the KELA caused by other non-S. enterica Enteritidis salmonellae. The KELA was also compared to two commercial rapid Pullorum tests, the Solvay (D-Sn = 94.9%; D-Sp = 55.5%) and the Vineland (D-Sn = 62.0%; D-Sp = 75.3%).

New strategy to improve quality control of Montenegro skin test at the production level.

PubMed

Guedes, Deborah Carbonera; Minozzo, João Carlos; Pasquali, Aline Kuhn Sbruzzi; Faulds, Craig; Soccol, Carlos Ricardo; Thomaz-Soccol, Vanete

2017-01-01

The production of the Montenegro antigen for skin test poses difficulties regarding quality control. Here, we propose that certain animal models reproducing a similar immune response to humans may be used in the quality control of Montenegro antigen production. Fifteen Cavia porcellus (guinea pigs) were immunized with Leishmania amazonensis or Leishmania braziliensis , and, after 30 days, they were skin tested with standard Montenegro antigen. To validate C. porcellus as an animal model for skin tests, eighteen Mesocricetus auratus (hamsters) were infected with L. amazonensis or L. braziliensis , and, after 45 days, they were skin tested with standard Montenegro antigen. Cavia porcellus immunized with L. amazonensis or L. braziliensis , and hamsters infected with the same species presented induration reactions when skin tested with standard Montenegro antigen 48-72h after the test. The comparison between immunization methods and immune response from the two animal species validated C. porcellus as a good model for Montenegro skin test, and the model showed strong potential as an in vivo model in the quality control of the production of Montenegro antigen.

Detection of antibodies to tuberculosis antigens in free-ranging lions (Panthera leo) infected with Mycobacterium bovis in Kruger National Park, South Africa.

PubMed

Miller, Michele; Joubert, Jennifer; Mathebula, Nomkhosi; De Klerk-Lorist, Lin-Marie; Lyashchenko, Konstantin P; Bengis, Roy; van Helden, Paul; Hofmeyr, Markus; Olea-Popelka, Francisco; Greenwald, Rena; Esfandiari, Javan; Buss, Peter

2012-06-01

Bovine tuberculosis (TB), caused by Mycobacterium bovis, has become established in Kruger National Park, South Africa, in the cape buffalo (Syncerus caffer) population and in other species. TB in prey species has resulted in infection and morbidity in the resident lion (Panthera leo) prides. The only validated live animal test currently available for lions is the intradermal tuberculin test. Because this test requires capture twice, 72 hr apart, of free-ranging lions to read results, it is logistically difficult to administer in a large ecosystem. Therefore, development of a rapid animal-side screening assay would be ideal in providing information for wildlife managers, veterinarians, and researchers working with free-living lion prides. This study reports preliminary descriptive results from an ongoing project evaluating two serologic tests for M. bovis (ElephantTB Stat-Pak and dual path platform VetTB). Disease status was determined by postmortem culture and presence of pathologic lesions in 14 free-ranging lions. Seropositivity was found to be associated with M. bovis infection. Extended field studies are underway to validate these rapid animal-side immunoassays for antemortem screening tests for TB in lions.

Sensitivity, specificity, and confounding factors of novel serological tests used for the rapid diagnosis of bovine tuberculosis in farmed red deer (Cervus elaphus).

PubMed

Buddle, Bryce M; Wilson, Tania; Denis, Michel; Greenwald, Rena; Esfandiari, Javan; Lyashchenko, Konstantin P; Liggett, Simon; Mackintosh, Colin G

2010-04-01

In this study, novel serological tests were used to detect tuberculosis (TB) in groups of farmed red deer (Cervus elaphus) varying in disease status or possible confounding factors. Groups of deer naturally or experimentally infected with Mycobacterium bovis and animals vaccinated against paratuberculosis were studied, as were uninfected animals and animals naturally or experimentally infected with Mycobacterium avium subsp. paratuberculosis. Sera were assayed using two rapid lateral-flow tests, Chembio's CervidTB STAT-PAK and DPP VetTB tests, and results were compared to those from tuberculin skin tests. Both serological tests had a high sensitivity, but specificity was adversely affected after animals had received a vaccine against paratuberculosis and were subsequently skin tested. The specificity of the DPP VetTB test was higher than that of the CervidTB STAT-PAK test, with natural infection with M. avium subsp. paratuberculosis adversely affecting the specificity of only the CervidTB STAT-PAK test. The sera from M. avium subsp. paratuberculosis-infected deer that produced false-positive reactions in the CervidTB STAT-PAK test were retested with a multiantigen print immunoassay (MAPIA), and some of these sera were shown to react with the MPB83 antigen. Combining the results from the serological tests and the skin tests showed only a slight increase in the sensitivity of detection of M. bovis-infected animals. It is concluded that both the CervidTB STAT-PAK and DPP VetTB tests offer rapid, convenient, and easy detection of bovine tuberculosis in deer, albeit with significant interference from paratuberculosis vaccination status and subsequent skin testing. The latter finding illustrates one of the limitations of currently available vaccines against paratuberculosis.

Direct Measurement of T Cell Receptor Affinity and Sequence from Naïve Anti-Viral T Cells

PubMed Central

Zhang, Shuqi; Parker, Patricia; Ma, Keyue; He, Chenfeng; Shi, Qian; Cui, Zhonghao; Williams, Chad; Wendel, Ben S.; Meriwether, Amanda; Salazar, Mary A.; Jiang, Ning

2016-01-01

T cells recognize and kill a myriad of pathogen-infected or cancer cells using a diverse set of T cell receptors (TCR). The affinity of TCR to cognate antigen is of high interest in adoptive T cell transfer immunotherapy and antigen-specific T cell repertoire immune profiling because it is widely known to correlate with downstream T cell responses. Here, we introduce the in situ TCR affinity and sequence test (iTAST) for simultaneous measurement of TCR affinity and sequence from single primary CD8+ T cells in human blood. We demonstrate that the repertoire of primary antigen-specific T cells from pathogen inexperienced individuals has a surprisingly broad affinity range of 1000-fold composed of diverse TCR sequences. Within this range, samples from older individuals contained a reduced frequency of high affinity T cells compared to young individuals, demonstrating an age-related effect of T cell attrition that could cause holes in the repertoire. iTAST should enable the rapid selection of high affinity TCRs ex vivo for adoptive immunotherapy and measurement of T cell response for immune monitoring applications. PMID:27252176

PREVALENCE OF LYMPHATIC FILARIASIS, MALARIA AND SOIL TRANSMITTED HELMINTHIASIS IN A COMMUNITY OF BARDIYA DISTRICT, WESTERN NEPAL.

PubMed

Ranjitkar, Samir; Alifrangis, Michael; Adhikari, Madhav; Olsen, Annette; Simonsen, Paul E; Meyrowitsch, Dan Wolf

2014-11-01

Lymphatic filariasis (LF), malaria and soil transmitted helminthiasis (STH) cause major health problems in Nepal, but in spite of this very few stud- ies have been carried out on these parasitic infections in Nepal. A cross sectional survey of all three categories of parasitic infections was carried out in Deuda- kala Village of Bardiya District, western Nepal. A total of 510 individuals aged 5 years and above were examined from finger prick blood for circulating filarial antigen (CFA), malaria antigen using a rapid diagnostic test (RDT), and malaria DNA using a PCR-based assay. In addition, 317 individuals were examined for soil-transmitted helminth (STH) eggs by the Kato-Katz technique. Prevalence of LF, malaria (antigen) and STH infection was 25.1%, 0.6% and 18.3%, respectively. PCR analysis did not detect any additional malaria cases. The prevalence of LF and STH infections differ significantly among different age groups and ethnic communities. The high prevalence of LF in the community studied indicates an immediate need for implementing a mass drug administration program for its control in this particular geographical area of Nepal.

Intrinsic Bioprobes, Inc. (Tempe, AZ)

DOEpatents

Nelson, Randall W [Phoenix, AZ; Williams, Peter [Phoenix, AZ; Krone, Jennifer Reeve [Granbury, TX

2008-07-15

Rapid mass spectrometric immunoassay methods for detecting and/or quantifying antibody and antigen analytes utilizing affinity capture to isolate the analytes and internal reference species (for quantification) followed by mass spectrometric analysis of the isolated analyte/internal reference species. Quantification is obtained by normalizing and calibrating obtained mass spectrum against the mass spectrum obtained for an antibody/antigen of known concentration.

Mass spectrometric immunoassay

DOEpatents

Nelson, Randall W; Williams, Peter; Krone, Jennifer Reeve

2007-12-04

Rapid mass spectrometric immunoassay methods for detecting and/or quantifying antibody and antigen analytes utilizing affinity capture to isolate the analytes and internal reference species (for quantification) followed by mass spectrometric analysis of the isolated analyte/internal reference species. Quantification is obtained by normalizing and calibrating obtained mass spectrum against the mass spectrum obtained for an antibody/antigen of known concentration.

Mass spectrometric immunoassay

DOEpatents

Nelson, Randall W; Williams, Peter; Krone, Jennifer Reeve

2013-07-16

Rapid mass spectrometric immunoassay methods for detecting and/or quantifying antibody and antigen analytes utilizing affinity capture to isolate the analytes and internal reference species (for quantification) followed by mass spectrometric analysis of the isolated analyte/internal reference species. Quantification is obtained by normalizing and calibrating obtained mass spectrum against the mass spectrum obtained for an antibody/antigen of known concentration.

Mass spectrometric immunoassay

DOEpatents

Nelson, Randall W.; Williams, Peter; Krone, Jennifer Reeve

2005-12-13

Rapid mass spectrometric immunoassay methods for detecting and/or quantifying antibody and antigen analytes utilizing affinity capture to isolate the analytes and internal reference species (for quantification) followed by mass spectrometric analysis of the isolated analyte/internal reference species. Quantification is obtained by normalizing and calibrating obtained mass spectrum against the mass spectrum obtained for an antibody/antigen of known concentration.

Prediction, dynamics, and visualization of antigenic phenotypes of seasonal influenza viruses

PubMed Central

Neher, Richard A.; Bedford, Trevor; Daniels, Rodney S.; Shraiman, Boris I.

2016-01-01

Human seasonal influenza viruses evolve rapidly, enabling the virus population to evade immunity and reinfect previously infected individuals. Antigenic properties are largely determined by the surface glycoprotein hemagglutinin (HA), and amino acid substitutions at exposed epitope sites in HA mediate loss of recognition by antibodies. Here, we show that antigenic differences measured through serological assay data are well described by a sum of antigenic changes along the path connecting viruses in a phylogenetic tree. This mapping onto the tree allows prediction of antigenicity from HA sequence data alone. The mapping can further be used to make predictions about the makeup of the future A(H3N2) seasonal influenza virus population, and we compare predictions between models with serological and sequence data. To make timely model output readily available, we developed a web browser-based application that visualizes antigenic data on a continuously updated phylogeny. PMID:26951657

Rapid detection of semenogelin by one-step immunochromatographic assay for semen identification.

PubMed

Sato, Itaru; Kojima, Koichiro; Yamasaki, Tadashi; Yoshida, Kaoru; Yoshiike, Miki; Takano, Shoichi; Mukai, Toshiji; Iwamoto, Teruaki

2004-04-01

To identify semen in forensic samples, we developed an analytical system for one-step immunoassay that has been constructed using the concept of immunochromatography and can identify semenogelin (Sg), which originates in the seminal vesicles. The system employed monoclonal antibody (mAb) and polyclonal antibody (pAb) against recombinant Sg-II (63 kDa), which has been synthesized in insect cells using baculovirus. The two antibodies bound with the seminal plasma motility inhibitor (SPMI; 14 kDa) as a final fragment peptide of Sg. The test stick is based on the sandwich technique using the above antibodies. When serial dilutions of seminal plasma were analyzed using this test stick, the intensity of a clear immunoreactive signal peaked at 2000-fold dilution. Thereafter, the signals decreased slowly but still persisted up to 400,000-fold dilution. The Sg antigen was undetectable in saliva, urine, breast milk, serum or vaginal secretions. Also, the test stick shown did not react with animal semen samples, such as those from horses, dogs, swine and bulls. When semen samples, diluted 100,000-fold from 100 men were tested, the Sg antigenic activity was detectable in all samples. In addition, the specificity and sensitivity of the test stick for identification of semen were demonstrated by comparative forensic studies. We conclude that this immunoassay method is a useful confirmatory test for the identification of semen. The immunochromatographic system for forensic testing or research use will become available commercially soon.

Malaria Rapid Diagnostic Devices: Performance Characteristics of the ParaSight F Device Determined in a Multisite Field Study

PubMed Central

Forney, J. Russ; Magill, Alan J.; Wongsrichanalai, Chansuda; Sirichaisinthop, Jeeraphat; Bautista, Christian T.; Heppner, D. Gray; Miller, R. Scott; Ockenhouse, Christian F.; Gubanov, Alex; Shafer, Robyn; DeWitt, Caroline Cady; Quino-Ascurra, Higinio A.; Kester, Kent E.; Kain, Kevin C.; Walsh, Douglas S.; Ballou, W. Ripley; Gasser, Robert A.

2001-01-01

Microscopic detection of parasites has been the reference standard for malaria diagnosis for decades. However, difficulty in maintaining required technical skills and infrastructure has spurred the development of several nonmicroscopic malaria rapid diagnostic devices based on the detection of malaria parasite antigen in whole blood. The ParaSight F test is one such device. It detects the presence of Plasmodium falciparum-specific histidine-rich protein 2 by using an antigen-capture immunochromatographic strip format. The present study was conducted at outpatient malaria clinics in Iquitos, Peru, and Maesod, Thailand. Duplicate, blinded, expert microscopy was employed as the reference standard for evaluating device performance. Of 2,988 eligible patients, microscopy showed that 547 (18%) had P. falciparum, 658 (22%) had P. vivax, 2 (0.07%) had P. malariae, and 1,750 (59%) were negative for Plasmodium. Mixed infections (P. falciparum and P. vivax) were identified in 31 patients (1%). The overall sensitivity of ParaSight F for P. falciparum was 95%. When stratified by magnitude of parasitemia (no. of asexual parasites per microliter of whole blood), sensitivities were 83% (>0 to 500 parasites/μl), 87% (501 to 1,000/μl), 98% (1,001 to 5,000/μl), and 98% (>5,000/μl). Device specificity was 86%. PMID:11474008

T Cells with Low Avidity for a Tissue-Restricted Antigen Routinely Evade Central and Peripheral Tolerance and Cause Autoimmunity

PubMed Central

Zehn, Dietmar; Bevan, Michael J.

2009-01-01

Summary T cells causing autoimmunity must escape tolerance. We observed that CD8+ T cells with high avidity for an antigen expressed in the pancreas, kidney, and thymic medulla were efficiently removed from a polyclonal repertoire by central and peripheral tolerance mechanisms. However, both mechanisms spared low-avidity T cells from elimination. Neither the introduction of activated, self-antigen-specific CD4+ helper T cells nor a global inflammatory stimulus were sufficient to activate the low-avidity CD8+ T cells and did not break tolerance. In contrast, challenge with a recombinant bacterium expressing the self antigen primed the low-avidity T cells, and the animals rapidly developed autoimmune diabetes. We suggest that whereas thymic and peripheral tolerance mechanisms remove cells that can be primed by endogenous amounts of self antigen, they do not guard against tissue destruction by low-avidity effector T cells, which have been primed by higher amounts of self antigen or by crossreactive antigens. PMID:16879996

Estimated saving of antibiotics in pharyngitis and lower respiratory tract infections if general practitioners used rapid tests and followed guidelines.

PubMed

Llor, Carles; Moragas, Ana; Cots, Josep M; López-Valcárcel, Beatriz González

General practitioners (GP) in Spain do not have access to rapid tests and adherence to guidelines is usually suboptimal. The aim of the study is to evaluate the estimated number of antibiotics that could have been saved if GPs had appropriately used these tests and had followed the guidelines. Observational study. Primary care centres from eight Autonomous Communities in Spain. GPs who had not participated in previous studies on rational use of antibiotics. GPs registered all the cases of pharyngitis and lower respiratory tract infections (LRTI) during 15 working days in 2015, by means of a 47-item audit. Actual GPs' antibiotic prescription and estimated number of antibiotics that could have been saved according to recent guidelines. A total of 126 GPs registered 1012 episodes of pharyngitis and 1928 LRTIs. Antibiotics were given or patients were referred in 497 patients with pharyngitis (49.1%) and 963 patients with LRTI (49.9%). If GPs had appropriately used rapid antigen detection tests and C-reactive protein tests and had strictly followed current guidelines, antibiotics would have been given to 7.6% and 15.1%, respectively, with an estimated saving of 420 antibiotics in patients with sore throat (estimated saving of 84.5%; 95% CI: 81.1-87.4%) and 672 antibiotics in LRTIs (estimated saving of 69.8%,95% CI: 67.1-72.5%). GP adherence to guidelines and a correct introduction of rapid tests in clinical practice in Spain could result in a considerable saving of unnecessary prescription of antibiotics in pharyngitis and LRTIs. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

Evaluation of a newly designed sandwich enzyme linked immunosorbent assay for the detection of hydatid antigen in serum, urine and cyst fluid for diagnosis of cystic echinococcosis.

PubMed

Chaya, Dr; Parija, Subhash Chandra

2013-07-01

Cystic echinococcosis (CE) is a zoonotic disease of humans with variable clinical manifestations. Imaging and immunological methods are currently the mainstay of diagnosis of this disease. Although the immunological tests for detection of anti-echinococcal antibodies have several disadvantages, they are widely being used. Antigen is far more superior than antibody detection test as they can provide a specific parasitic diagnosis. A sandwich enzyme linked immunosorbent assay (ELISA) was designed using antibodies to 24 kDa urinary hydatid antigen for the detection of hydatid antigens in urine, serum and cyst fluid specimens. The performance of this novel test was compared with that of other hydatid antibody detection ELISA and enzyme immune transfer blot (EITB) using radiological and surgical confirmation as the gold standard. The antigen detection ELISA showed 100% sensitivity and specificity when tested with cyst fluid. On testing urine and serum, the antigen detection ELISA was found to be more specific than antibody detection ELISA. EITB was found to be the most sensitive and specific test. ELISA using polyclonal antibodies against 24 kDa urinary hydatid protein was moderately sensitive to detect hydatid antigen in serum and urine. Hence polyclonal antibodies to 24 kDa urinary hydatid antigen can be used as an alternative source of antibody to detect hydatid antigen in serum, urine and cyst fluid. In the present study, EITB was found to be highly specific test for detection of hydatid antibodiesin serum. 24 kDa protein was found to be specific and of diagnostic value in CE.

Comparison of Antibody Responses to a Potential Combination of Specific Glycolipids and Proteins for Test Sensitivity Improvement in Tuberculosis Serodiagnosis

PubMed Central

Julián, Esther; Matas, Lurdes; Alcaide, José; Luquin, Marina

2004-01-01

The humoral response to different proteinaceous antigens of Mycobacterium tuberculosis is heterogeneous among patients with active disease, and this has originated in the proposal to use a combination of several specific antigens to find an efficient serodiagnostic test for tuberculosis (TB). However, to date, comparisons of antibody responses to several antigens in the same population have been carried out without consideration of antigenic cell wall glycolipids. In the present study the presence of immunoglobulin G (IgG), IgM, and IgA antibodies to M. tuberculosis glycolipids (sulfolipid I, diacyltrehaloses, triacyltrehaloses, and cord factor) was compared with the response to four commercially available tests based on the 38-kDa protein mixed with the 16-kDa protein or lipoarabinomannan. Fifty-two serum samples from TB patients and 83 serum samples from control individuals (48 healthy individuals and 35 non-TB pneumonia patients) were studied. Three relevant results were obtained. (i) Smear-negative TB patients presented low humoral responses, but the sera which did react principally showed IgA antibodies to some glycolipidic antigens. (ii) TB patients exhibit heterogeneous humoral responses against glycolipidic antigens. (iii) Finally, test sensitivity is improved (from 23 to 62%) when IgG and IgA antibodies are detected together in tests based on different antigens (proteins and glycolipids). We conclude that it is possible to include glycolipidic antigens in a cocktail of specific antigens from M. tuberculosis to develop a serodiagnostic test. PMID:14715547

Skin testing of guinea pigs and footpad testing of mice with a new antigen for detecting delayed hypersensitivity to Cryptococcus neoformans.

PubMed

Murphy, J W; Gregory, J A; Larsh, H W

1974-02-01

This study was undertaken to evaluate the potential of a cryptococcal culture filtrate antigen, cryptococcin C184, for detecting delayed hypersensitivity in Cryptococcus neoformans-injected animals. The antigen was tested on guinea pigs which had received saline or C. neoformans and on animals sensitized to Histoplasma capsulatum, Blastomyces dermatitidis, Candida albicans, or Sporothrix schenckii. A delayed-type hypersensitivity response was elicited by cryptococcin C184 in C. neoformans-injected guinea pigs, whereas no indurations or erythemas were seen at 48 h after skin testing of saline controls or heterologously sensitized guinea pigs. Besides being specific for Cryptococcus, the antigen showed a high degree of sensitivity and was reproducible. Footpad tests were conducted with the antigen on mice which had previously received either 10(5) viable C. neoformans cells or saline. Delayed hypersensitivity was indicated in the C. neoformans-injected mice by the increase in thickness of antigen-injected footpads when compared with the saline-injected footpads. In control mice, antigen- and saline-injected footpads were comparable in thickness 24 h after injection. Mice sensitized to B. dermatitidis were footpad tested with C184, and no cross-reactivity was demonstrated.

Detection of CMV pneumonitis after lung transplantation using PCR of DNA from bronchoalveolar lavage cells.

PubMed

Bewig, B; Haacke, T C; Tiroke, A; Bastian, A; Böttcher, H; Hirt, S W; Rautenberg, P; Haverich, A

2000-01-01

Cytomegalovirus (CMV) is known as a common pathogen causing infections after lung transplantation. Rapid diagnosis of CMV infection is important for the initiation of a specific treatment. Evaluation of methods for the rapid diagnosis of CMV pneumonitis. The detection rates of CMV DNA in bronchoalveolar lavage (BAL) and bronchial brushes by polymerase chain reaction (PCR), of viral antigens (p52 and IE1) in BAL and of pp65 antigen in peripheral blood leukocytes were compared to the clinical status after lung transplantation. In 28 patients, 105 BAL, 96 blood samples and 14 brushes were analyzed. In 6 patients, a total of eight episodes of CMV pneumonitis occurred. Five of the 6 with positive CMV antigens in BAL (p52 or IE1) showed signs of CMV pneumonitis. All episodes of CMV pneumonitis were detected by the PCR of BAL cells. Fourteen samples positive for CMV pp65 antigen in blood were negative in BAL PCR. In these cases, no clinical signs of pulmonary CMV infection occurred. Overall sensitivity, specificity, and positive and negative predictive values for the detection of CMV pneumonitis by PCR of BAL cells were 100, 98.9, 88.9 and 100%, respectively. In brush samples, PCR did not provide additional information to the results of the PCR of BAL cells. PCR of DNA from BAL cells is suitable for reliable and rapid detection of CMV pneumonitis. Copyright 2000 S. Karger AG, Basel.

Carbon nanotubes as vaccine scaffolds

PubMed Central

Scheinberg, David A.; McDevitt, Michael R.; Dao, Tao; Mulvey, Justin J.; Feinberg, Evan; Alidori, Simone

2013-01-01

Carbon nanotubes display characteristics that are potentially useful in their development as scaffolds for vaccine compositions. These features include stability in vivo, lack of intrinsic immunogenicity, low toxicity, and the ability to be appended with multiple copies of antigens. In addition, the particulate nature of carbon nanotubes and their unusual properties of rapid entry into antigen-presenting cells, such as dendritic cells, make them especially useful as carriers of antigens. Early attempts demonstrating carbon nanotube-based vaccines can be used in both infectious disease settings and cancer are promising. PMID:23899863

Rapid, whole blood diagnostic test for detecting anti-hantavirus antibody in rats.

PubMed

Amada, Takako; Yoshimatsu, Kumiko; Yasuda, Shumpei P; Shimizu, Kenta; Koma, Takaaki; Hayashimoto, Nobuhito; Gamage, Chandika D; Nishio, Sanae; Takakura, Akira; Arikawa, Jiro

2013-10-01

Hantavirus is a causative agent of rodent-borne viral zoonoses, hemorrhagic fever with renal syndrome (HFRS) and hantavirus pulmonary syndrome. Seoul virus (SEOV) is a causative agent of urban and laboratory rat-associated HFRS worldwide. Surveillance of rodents has been done mainly by serological detection of hantavirus-specific antibodies by enzyme linked immunosorbent assay (ELISA) and immunofluorescent antibody assay (IFA). An immunochromatographic (ICG) test was developed with the N-terminal 103 amino acids of nucleocapsid protein of Hantaan virus expressed by Escherichia coli as an antigen to detect IgG antibody specific to hantavirus in sera from Rattus sp. animals. Antibody-detecting sensitivity of the ICG test was the same as that of ELISA and about 100-times higher than that of IFA. Overall sensitivities and specificities of the ICG test in comparison to ELISA and IFA for sera from 192 urban rats and 123 laboratory rats were 99.3% and 100%, respectively. Diluted whole blood samples without separation could be used for the ICG test. The ICG test enabled detection of antibodies to SEOV, Hantaan, Dobrava/Belgrade, and Thailand viruses, which are causative agents of HFRS throughout Eurasia. The ICG test is a rapid, simple and safe method for diagnosis of SEOV infection in rats. Copyright © 2013 Elsevier B.V. All rights reserved.

Development of an immunochromatographic test with anti-LipL32-coupled gold nanoparticles for Leptospira detection.

PubMed

Chirathaworn, Chintana; Janwitthayanan, Weena; Sereemaspun, Amornpun; Lertpocasombat, Kanchalee; Rungpanich, Utane; Ekpo, Pattama; Suwancharoen, Duangjai

2014-04-01

Detection of antibody specific to Leptospira by various immunological techniques has been used for leptospirosis diagnosis. However, the sensitivity of antibody detection during the first few days after infection is low. Molecular techniques are suggested to provide earlier diagnosis than antibody detection, but a rapid and easy to perform assay for Leptospira antigen detection would provide an additional useful tool for disease diagnosis. In this study, we coupled gold nanoparticles with antibody to LipL32, a protein commonly found in pathogenic Leptospira. This coupled gold reagent was used in the immunochromatographic strip for Leptospira detection. We demonstrated that the sensitivity of Leptospira detection by this strip was 10(3) ml(-1). There was no positive result detected when strips were tested with non-pathogenic Leptospira, Staphylococcus aureus, Streptococcus group B, Acinetobacter baumannii, Escherichia coli, Salmonella typhi, Klebsiella pneumoniae, Enterococcus faecalis or Enterococcus faecium. These data suggest that gold nanoparticles coupled with antibody to LipL32 could be used for Leptospira detection by a rapid test based on an immunochromatographic technique.

A novel dendritic cell-based direct ex vivo assay for detection and enumeration of circulating antigen-specific human T cells.

PubMed

Carrio, Roberto; Zhang, Ge; Drake, Donald R; Schanen, Brian C

2018-05-07

Although a variety of assays have been used to examine T cell responses in vitro, standardized ex vivo detection of antigen-specific CD4 + T cells from human circulatory PBMCs remains constrained by low-dimensional characterization outputs and the need for polyclonal, mitogen-induced expansion methods to generate detectable response signals. To overcome these limitations, we developed a novel methodology utilizing antigen-pulsed autologous human dendritic target cells in a rapid and sensitive assay to accurately enumerate antigen-specific CD4 + T cell precursor frequency by multiparametric flow cytometry. With this approach, we demonstrate the ability to reproducibly quantitate poly-functional T cell responses following both primary and recall antigenic stimulation. Furthermore, this approach enables more comprehensive phenotypic profiling of circulating antigen-specific CD4 + T cells, providing valuable insights into the pre-existing polarization of antigen-specific T cells in humans. Combined, this approach permits sensitive and detailed ex vivo detection of antigen-specific CD4 + T cells delivering an important tool for advancing vaccine, immune-oncology and other therapeutic studies.

[Laboratory diagnosis of genital herpes--direct immunofluorescence method].

PubMed

Majewska, Anna; Romejko-Wolniewicz, Ewa; Zareba-Szczudlik, Julia; Kilijańczyk, Marek; Gajewska, Małgorzata; Młynarczyk, Grazyna

2013-07-01

Aim of the study was to determine clinical usefulness of direct immunofluorescence method in the laboratory diagnosis of genital herpes in women. Overall 187 anogenital swabs were collected from 120 women. Using a dacron-tipped applicator 83 swabs were collected from women suspected of genital herpes and 104 from patients with no signs of genital infection. All samples were tested using cell culture (Vero cell line) and then direct immunofluorescence method (DIF) for the identification of antigens of herpes simplex viruses: HSV-1 and HSV-2. Characteristic cytopathic effect (CPE), indicative of alphaherpesvirus infection, was observed in 43.4% of cultures with clinical specimens collected from women with suspected genital herpes and in 29.8% of cultures of clinical specimens taken from patients with no clinical symptoms of genital herpes. Herpes simplex viruses were determined in 73 samples by direct immunofluorescence method after amplification of the virus in cell culture. The DIF test confirmed the diagnosis based on the microscopic CPE observation in 85%. In 15% of samples (taken from pregnant women without clinical signs of infection) we reported positive immunofluorescence in the absence of CPE. The frequency of antigen detection was statistically significantly higher in samples that were positive by culture study (chi-square test with Yates's correction, p < 0.01). This method proved to be highly sensitive (97%) in women with clinically suspected infection. High negative predictive value (99%) proves the clinical utility of the DIF in these group of patients. In asymptomatic infections, viral antigens were detected most frequently in the swabs from the cervical canal, and in cases of suspected genital herpes in swabs taken from the vestibule of the vagina and the vulva. However, there was no statistically significant difference in the frequency of detection of Herpes Simplex Virus antigens in specimens from different parts of the genital tract in both groups of women (chi-square test, p > 0.05). In our study HHV-1 was the main causative agent of genital herpes. The growing worldwide prevalence of genital herpes, challenges with the clinical diagnosis, and availability of effective antiviral therapy are the main reasons for a growing interest in rapid, proper laboratory diagnosis of infected patients. Optimal testing diagnostic algorithm depends on patient population, clinical circumstances and availability. Our results indicated that combination of laboratory tests may help to establish the diagnosis if genital herpes is suspected but there are no typical signs.

Development of a Colloidal Gold-Based Immunochromatographic Strip for Rapid Detection of Klebsiella pneumoniae Serotypes K1 and K2

PubMed Central

Siu, L. Kristopher; Tsai, Yu-Kuo; Lin, Jung-Chung; Chen, Te-Li; Fung, Chang-Phone

2016-01-01

In this study, a novel colloidal gold-based immunochromatographic strip (ICS) containing anti-Klebsiella pneumoniae capsular polysaccharide polyclonal antibodies was developed to specifically detect K. pneumoniae serotypes K1 and K2. Capsular polysaccharide K1 and K2 antigens were first used to produce polyclonal anti-K1 and anti-K2 antibodies. Reference strains with different serotypes, nontypeable K. pneumoniae strains, and other bacterial species were then used to assess the sensitivity and specificity of these test strips. The detection limit was found to be 105 CFU, and the ICSs were stable for 6 months when stored at room temperature. No false-positive or false-negative results were observed, and equivalent results were obtained compared to those of more conventional test methods, such as PCR or serum agglutination. In conclusion, the ICS developed here requires no technical expertise and allows for the specific, rapid, and simultaneous detection of K. pneumoniae serotypes K1 and K2. PMID:27707941

Ecobody technology: rapid monoclonal antibody screening method from single B cells using cell-free protein synthesis for antigen-binding fragment formation.

PubMed

Ojima-Kato, Teruyo; Nagai, Satomi; Nakano, Hideo

2017-10-25

We report a rapid and cost-effective monoclonal antibody screening method from single animal B cells using reverse transcription (RT)-PCR and Escherichia coli cell-free protein synthesis (CFPS), which allows evaluation of antibodies within 2 working days. This process is named "Ecobody technology". The method includes strategies to isolate B cells that specifically bind an antigen from the peripheral blood of immunised animals, and single-cell RT-PCR to generate DNA fragments of the V H and V L genes, followed by CFPS for production of fragments of antigen binding (Fab). In the CFPS step, we employed our techniques: 1) 'Zipbody' as a method for producing Fab, in which the association of heavy and light chains is facilitated by adhesive leucine zipper peptides fused at the C-termini of the Fab; and 2) an N-terminal SKIK peptide tag that can increase protein expression levels. Using Ecobody technology, we obtained highly-specific monoclonal antibodies for the antigens Vibrio parahaemolyticus and E. coli O26. The anti-V. parahaemolyticus Zipbody mAb was further produced in E. coli strain SHuffle T7 Express in inclusion bodies and refolded by a conventional method, resulting in significant antigen-binding activity (K D  = 469 pM) and productivity of 8.5 mg purified antibody/L-culture.

From genomes to vaccines: Leishmania as a model.

PubMed Central

Almeida, Renata; Norrish, Alan; Levick, Mark; Vetrie, David; Freeman, Tom; Vilo, Jaak; Ivens, Alasdair; Lange, Uta; Stober, Carmel; McCann, Sharon; Blackwell, Jenefer M

2002-01-01

The 35 Mb genome of Leishmania should be sequenced by late 2002. It contains approximately 8500 genes that will probably translate into more than 10 000 proteins. In the laboratory we have been piloting strategies to try to harness the power of the genome-proteome for rapid screening of new vaccine candidate. To this end, microarray analysis of 1094 unique genes identified using an EST analysis of 2091 cDNA clones from spliced leader libraries prepared from different developmental stages of Leishmania has been employed. The plan was to identify amastigote-expressed genes that could be used in high-throughput DNA-vaccine screens to identify potential new vaccine candidates. Despite the lack of transcriptional regulation that polycistronic transcription in Leishmania dictates, the data provide evidence for a high level of post-transcriptional regulation of RNA abundance during the developmental cycle of promastigotes in culture and in lesion-derived amastigotes of Leishmania major. This has provided 147 candidates from the 1094 unique genes that are specifically upregulated in amastigotes and are being used in vaccine studies. Using DNA vaccination, it was demonstrated that pooling strategies can work to identify protective vaccines, but it was found that some potentially protective antigens are masked by other disease-exacerbatory antigens in the pool. A total of 100 new vaccine candidates are currently being tested separately and in pools to extend this analysis, and to facilitate retrospective bioinformatic analysis to develop predictive algorithms for sequences that constitute potentially protective antigens. We are also working with other members of the Leishmania Genome Network to determine whether RNA expression determined by microarray analyses parallels expression at the protein level. We believe we are making good progress in developing strategies that will allow rapid translation of the sequence of Leishmania into potential interventions for disease control in humans. PMID:11839176

Detection of soluble antigens of Toxoplasma gondii by a four-layer modification of an enzyme immunoassay.

PubMed Central

Turunen, H J

1983-01-01

A sensitive four-layer modification of an enzyme immunoassay for the detection of soluble antigens of Toxoplasma gondii is described. Microtiter plates were sensitized with rabbit anti-toxoplasma immunoglobulins (6 micrograms/ml) used as the primary antibodies; guinea pig anti-toxoplasma immunoglobulins (6 micrograms/ml) were used as the secondary trapping antibodies. Horseradish peroxidase-conjugated anti-guinea pig immunoglobulins were used as the indicator antibodies. The specificity of the antigen assay was confirmed by using guinea pig immunoglobulins from preimmunization sera. The sensitivity of the antigen assay was found to be at least 10 ng of antigen protein per ml. The suitability of the method for detecting antigens of T. gondii in different specimens was studied by experimental toxoplasma infection in mice. Antigenic components of T. gondii could be detected in different tissue specimens from infected animals from the first day after infection onwards. Toxoplasma antigen in serum and urine samples from infected mice reached detectable levels on day 2 after infection followed by a linear increase in antigen concentration in succeeding samples. This method might offer a valuable aid for a rapid etiological diagnosis also in human cases of acute toxoplasmosis. PMID:6345574

Production and Evaluation of a Purified Adenovirus Group-Specific (Hexon) Antigen for Use in the Diagnostic Complement Fixation Test

PubMed Central

Dowdle, W. R.; Lambriex, M.; Hierholzer, J. C.

1971-01-01

A simple procedure for the production of large volumes of purified adenovirus group-specific complement-fixing (CF) (hexon) antigen by selective adsorption to and elution from CaHPO4 is described. Results of immunodiffusion tests, electrophoresis, electron microscopy, and tests for hemagglutination and infectivity indicate that the purified antigen consisted of a single virus component (hexon). The purified product contained little host materials. Unlike the crude virus harvest usually employed for serodiagnostic CF tests, the purified antigen demonstrated no anticomplementary activity and did not develop such activity during storage. The purified antigen was equal to or slightly more sensitive than crude virus harvests for serodiagnosis of adenovirus infections. Images PMID:4325021

Lateral Flow Rapid Test for Accurate and Early Diagnosis of Scrub Typhus: A Febrile Illness of Historically Military Importance in the Pacific Rim.

PubMed

Chao, Chien-Chung; Zhangm, Zhiwen; Weissenberger, Giulia; Chen, Hua-Wei; Ching, Wei-Mei

2017-03-01

Scrub typhus (ST) is an infection caused by Orientia tsutsugamushi. Historically, ST was ranked as the second most important arthropod-borne medical problem only behind malaria during World War II and the Vietnam War. The disease occurs mainly in Southeast Asia and has been shown to emerge and reemerge in new areas, implying the increased risk for U.S. military and civilian personnel deployed to these regions. ST can effectively be treated by doxycycline provided the diagnosis is made early, before the development of severe complications. Scrub Typhus Detect is a lateral flow rapid test based on a mixture of recombinant 56-kDa antigens with broad reactivity. The performance of this prototype product was evaluated against indirect immunofluorescence assay, the serological gold standard. Using 249 prospectively collected samples from Thailand, the sensitivity and specificity for IgM was found to be 100% and 92%, respectively, suggesting a high potential of this product for clinical use. This product will provide a user friendly, rapid, and accurate diagnosis of ST for clinicians to provide timely and accurate treatments of deployed personnel. Reprint & Copyright © 2017 Association of Military Surgeons of the U.S.

A novel double recognition enzyme-linked immunosorbent assay based on the nucleocapsid protein for early detection of European porcine reproductive and respiratory syndrome virus infection.

PubMed

Venteo, A; Rebollo, B; Sarraseca, J; Rodriguez, M J; Sanz, A

2012-04-01

Precise and rapid detection of porcine reproductive respiratory syndrome virus (PRRSV) infection in swine farms is critical. Improvement of control procedures, such as testing incoming gilt and surveillance of seronegative herds requires more rapid and sensitive methods. However, standard serological techniques detect mainly IgG antibodies. A double recognition enzyme-linked immunosorbent assay (DR-ELISA) was developed for detection of antibodies specific to European PRRSV. This new assay can recognize both IgM and IgG antibodies to PRSSV which might be useful for detecting in routine surveillance assays pigs that are in the very early stages of infection and missed by conventional assays detecting only IgG antibodies. DR-ELISA is based on the double recognition of antigen by antibody. In this study, the recombinant nucleocapsid protein (N) of PRRSV was used both as the coating and the enzyme-conjugated antigen. To evaluate the sensitivity of the assay at early stages of the infection, sera from 69 pigs infected with PRRSV were collected during successive days post infection (pi) and tested. While standard methods showed low sensitivity rates before day 14 pi, DR-ELISA detected 88.4% seropositive samples at day 7 showing greater sensitivity at early stages of the infection. Further studies were carried out to assess the efficiency of the new assay, and the results showed DR-ELISA to be a sensitive and accurate method for early diagnosis of EU-PRRSV infection. Copyright © 2012 Elsevier B.V. All rights reserved.

Guideline for the management of acute sore throat.

PubMed

Pelucchi, C; Grigoryan, L; Galeone, C; Esposito, S; Huovinen, P; Little, P; Verheij, T

2012-04-01

The European Society for Clinical Microbiology and Infectious Diseases established the Sore Throat Guideline Group to write an updated guideline to diagnose and treat patients with acute sore throat. In diagnosis, Centor clinical scoring system or rapid antigen test can be helpful in targeting antibiotic use. The Centor scoring system can help to identify those patients who have higher likelihood of group A streptococcal infection. In patients with high likelihood of streptococcal infections (e.g. 3-4 Centor criteria) physicians can consider the use of rapid antigen test (RAT). If RAT is performed, throat culture is not necessary after a negative RAT for the diagnosis of group A streptococci. To treat sore throat, either ibuprofen or paracetamol are recommended for relief of acute sore throat symptoms. Zinc gluconate is not recommended to be used in sore throat. There is inconsistent evidence of herbal treatments and acupuncture as treatments for sore throat. Antibiotics should not be used in patients with less severe presentation of sore throat, e.g. 0-2 Centor criteria to relieve symptoms. Modest benefits of antibiotics, which have been observed in patients with 3-4 Centor criteria, have to be weighed against side effects, the effect of antibiotics on microbiota, increased antibacterial resistance, medicalisation and costs. The prevention of suppurative complications is not a specific indication for antibiotic therapy in sore throat. If antibiotics are indicated, penicillin V, twice or three times daily for 10 days is recommended. At the present, there is no evidence enough that indicates shorter treatment length. © 2012 The Authors. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

Evaluation of a newly designed sandwich enzyme linked immunosorbent assay for the detection of hydatid antigen in serum, urine and cyst fluid for diagnosis of cystic echinococcosis

PubMed Central

Chaya, DR; Parija, Subhash Chandra

2013-01-01

Introduction: Cystic echinococcosis (CE) is a zoonotic disease of humans with variable clinical manifestations. Imaging and immunological methods are currently the mainstay of diagnosis of this disease. Although the immunological tests for detection of anti-echinococcal antibodies have several disadvantages, they are widely being used. Antigen is far more superior than antibody detection test as they can provide a specific parasitic diagnosis. Materials and Methods: A sandwich enzyme linked immunosorbent assay (ELISA) was designed using antibodies to 24 kDa urinary hydatid antigen for the detection of hydatid antigens in urine, serum and cyst fluid specimens. The performance of this novel test was compared with that of other hydatid antibody detection ELISA and enzyme immune transfer blot (EITB) using radiological and surgical confirmation as the gold standard. Results: The antigen detection ELISA showed 100% sensitivity and specificity when tested with cyst fluid. On testing urine and serum, the antigen detection ELISA was found to be more specific than antibody detection ELISA. EITB was found to be the most sensitive and specific test. Conclusions: ELISA using polyclonal antibodies against 24 kDa urinary hydatid protein was moderately sensitive to detect hydatid antigen in serum and urine. Hence polyclonal antibodies to 24 kDa urinary hydatid antigen can be used as an alternative source of antibody to detect hydatid antigen in serum, urine and cyst fluid. In the present study, EITB was found to be highly specific test for detection of hydatid antibodiesin serum. 24 kDa protein was found to be specific and of diagnostic value in CE. PMID:24470996

[Contribution of HCV core antigen testing in HCV diagnosis by test from the company Abbott Laboratories].

PubMed

Trbusek, J

2009-11-01

Detection of HCV core antigen as direct marker of hepatitis C infection clearly improves diagnosis of this disease (especially reduction of window period) and brings broad clinical utilization. The company Abbott Laboratories offers fully automated laboratory test for measurement of HCV core antigen on ARCHITECT analyzers.

Evaluation of single-round infectious, chimeric dengue type 1 virus as an antigen for dengue functional antibody assays.

PubMed

Yamanaka, Atsushi; Suzuki, Ryosuke; Konishi, Eiji

2014-07-23

Dengue fever and dengue hemorrhagic fever are endemic throughout tropical and subtropical countries. Four serotypes of dengue viruses (DENV-1 to DENV-4), each with several genotypes including various subclades, are co-distributed in most endemic areas. Infection-neutralizing and -enhancing antibodies are believed to play protective and pathogenic roles, respectively. Measurement of these functional antibodies against a variety of viral strains is thus important for evaluating coverage and safety of dengue vaccine candidates. Although transportation of live virus materials beyond national borders is increasingly limited, this difficulty may be overcome using biotechnology that enables generation of an antibody-assay antigen equivalent to authentic virus based on viral sequence information. A rapid system to produce flavivirus single-round infectious particles (SRIPs) was recently developed using a Japanese encephalitis virus (JEV) subgenomic replicon plasmid. This system allows production of chimeric SRIPs that have surface proteins of other flaviviruses. In the present study, SRIPs of DENV-1 (D1-SRIPs) were evaluated as an antigen for functional antibody assays. Inclusion of the whole mature capsid gene of JEV into the replicon plasmid provided higher D1-SRIP yields than did its exclusion in cases where a DENV-1 surface-protein-expressing plasmid was used for co-transfection of 293T cells with the replicon plasmid. In an assay to measure the balance between neutralizing and enhancing activities, dose (antibody dilution)-dependent activity curves in dengue-immune human sera or mouse monoclonal antibodies obtained using D1-SRIP antigen were equivalent to those obtained using DENV-1 antigen. Similar results were obtained using additional DENV-2 and DENV-3 systems. In a conventional Vero-cell neutralization test, a significant correlation was shown between antibody titers obtained using D1-SRIP and DENV-1 antigens. These results demonstrate the utility of D1-SRIPs as an alternative antigen to authentic DENV-1 in functional antibody assays. SRIP antigens may contribute to dengue vaccine candidate evaluation, understanding of dengue pathogenesis, and development of serodiagnostic systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

Determination of antigenicity-altering patches on the major surface protein of human influenza A/H3N2 viruses

PubMed Central

Kratsch, Christina; Klingen, Thorsten R.; Mümken, Linda; Steinbrück, Lars; McHardy, Alice C.

2016-01-01

Human influenza viruses are rapidly evolving RNA viruses that cause short-term respiratory infections with substantial morbidity and mortality in annual epidemics. Uncovering the general principles of viral coevolution with human hosts is important for pathogen surveillance and vaccine design. Protein regions are an appropriate model for the interactions between two macromolecules, but the currently used epitope definition for the major antigen of influenza viruses, namely hemagglutinin, is very broad. Here, we combined genetic, evolutionary, antigenic, and structural information to determine the most relevant regions of the hemagglutinin of human influenza A/H3N2 viruses for interaction with human immunoglobulins. We estimated the antigenic weights of amino acid changes at individual sites from hemagglutination inhibition data using antigenic tree inference followed by spatial clustering of antigenicity-altering protein sites on the protein structure. This approach determined six relevant areas (patches) for antigenic variation that had a key role in the past antigenic evolution of the viruses. Previous transitions between successive predominating antigenic types of H3N2 viruses always included amino acid changes in either the first or second antigenic patch. Interestingly, there was only partial overlap between the antigenic patches and the patches under strong positive selection. Therefore, besides alterations of antigenicity, other interactions with the host may shape the evolution of human influenza A/H3N2 viruses. PMID:27774294

Immunoglobulins drive terminal maturation of splenic dendritic cells

PubMed Central

Ziętara, Natalia; Łyszkiewicz, Marcin; Puchałka, Jacek; Pei, Gang; Gutierrez, Maximiliano Gabriel; Lienenklaus, Stefan; Hobeika, Elias; Reth, Michael; Martins dos Santos, Vitor A. P.; Krueger, Andreas; Weiss, Siegfried

2013-01-01

Nature and physiological status of antigen-presenting cells, such as dendritic cells DCs, are decisive for the immune reactions elicited. Multiple factors and cell interactions have been described that affect maturation of DCs. Here, we show that DCs arising in the absence of immunoglobulins (Ig) in vivo are impaired in cross-presentation of soluble antigen. This deficiency was due to aberrant cellular targeting of antigen to lysosomes and its rapid degradation. Function of DCs could be restored by transfer of Ig irrespective of antigen specificity and isotype. Modulation of cross-presentation by Ig was inhibited by coapplication of mannan and, thus, likely to be mediated by C-type lectin receptors. This unexpected dependency of splenic DCs on Ig to cross-present antigen provides insights into the interplay between cellular and humoral immunity and the immunomodulatory capacity of Ig. PMID:23345431

Comparison of a Clinical Prediction Rule and a LAM Antigen-Detection Assay for the Rapid Diagnosis of TBM in a High HIV Prevalence Setting

PubMed Central

Patel, Vinod B.; Singh, Ravesh; Connolly, Cathy; Kasprowicz, Victoria; Zumla, Allimudin; Ndungu, Thumbi; Dheda, Keertan

2010-01-01

Background/Objective The diagnosis of tuberculous meningitis (TBM) in resource poor TB endemic environments is challenging. The accuracy of current tools for the rapid diagnosis of TBM is suboptimal. We sought to develop a clinical-prediction rule for the diagnosis of TBM in a high HIV prevalence setting, and to compare performance outcomes to conventional diagnostic modalities and a novel lipoarabinomannan (LAM) antigen detection test (Clearview-TB®) using cerebrospinal fluid (CSF). Methods Patients with suspected TBM were classified as definite-TBM (CSF culture or PCR positive), probable-TBM and non-TBM. Results Of the 150 patients, 84% were HIV-infected (median [IQR] CD4 count = 132 [54; 241] cells/µl). There were 39, 55 and 54 patients in the definite, probable and non-TBM groups, respectively. The LAM sensitivity and specificity (95%CI) was 31% (17;48) and 94% (85;99), respectively (cut-point ≥0.18). By contrast, smear-microscopy was 100% specific but detected none of the definite-TBM cases. LAM positivity was associated with HIV co-infection and low CD4 T cell count (CD4<200 vs. >200 cells/µl; p = 0.03). The sensitivity and specificity in those with a CD4<100 cells/µl was 50% (27;73) and 95% (74;99), respectively. A clinical-prediction rule ≥6 derived from multivariate analysis had a sensitivity and specificity (95%CI) of 47% (31;64) and 98% (90;100), respectively. When LAM was combined with the clinical-prediction-rule, the sensitivity increased significantly (p<0.001) to 63% (47;68) and specificity remained high at 93% (82;98). Conclusions Despite its modest sensitivity the LAM ELISA is an accurate rapid rule-in test for TBM that has incremental value over smear-microscopy. The rule-in value of LAM can be further increased by combination with a clinical-prediction rule, thus enhancing the rapid diagnosis of TBM in HIV-infected persons with advanced immunosuppression. PMID:21203513

Canine parvovirus 2c infection in central Portugal.

PubMed

João Vieira, Maria; Silva, Eliane; Oliveira, João; Luísa Vieira, Ana; Decaro, Nicola; Desario, Costantina; Muller, Alexandra; Carvalheira, Júlio; Buonavoglia, Canio; Thompson, Gertrude

2008-07-01

Canine parvovirus (CPV) has been evolving, generating new genetic and antigenic variants throughout the world. This study was conducted to determine the types of CPV circulating in dogs in Figueira da Foz, Portugal. Thirty fecal samples, collected between 2006 and 2007 from dogs with clinical signs of CPV infection, were tested for CPV by a rapid, in-clinic, enzyme-linked immunosorbent assay (ELISA)/immunomigration test, by conventional real-time polymerase chain reaction (PCR), and by minor-groove binding TaqMan PCR. Of the 29 PCR-positive samples, 15 were identified as CPV-2b and 14 as CPV-2c. No CPV-2a was detected. The sensitivity of the ELISA test was 82.76% compared with the PCR assays. No significant associations were found between CPV type, clinical outcome, breed, vaccination status, or age.

Detection of Entamoeba histolytica by Recombinase Polymerase Amplification

PubMed Central

Nair, Gayatri; Rebolledo, Mauricio; White, A. Clinton; Crannell, Zachary; Richards-Kortum, R. Rebecca; Pinilla, A. Elizabeth; Ramírez, Juan David; López, M. Consuelo; Castellanos-Gonzalez, Alejandro

2015-01-01

Amebiasis is an important cause of diarrheal disease worldwide and has been associated with childhood malnutrition. Traditional microscopy approaches are neither sensitive nor specific for Entamoeba histolytica. Antigen assays are more specific, but many cases are missed unless tested by molecular methods. Although polymerase chain reaction (PCR) is effective, the need for sophisticated, expensive equipment, infrastructure, and trained personnel limits its usefulness, especially in the resource-limited, endemic areas. Here, we report development of a recombinase polymerase amplification (RPA) method to detect E. histolytica specifically. Using visual detection by lateral flow (LF), the test was highly sensitive and specific and could be performed without additional equipment. The availability of this inexpensive, sensitive, and field-applicable diagnostic test could facilitate rapid diagnosis and treatment of amebiasis in endemic regions. PMID:26123960

Evaluation of a rapid immunochromatographic test for the detection of low burden Dirofilaria immitis (heartworm) in dogs and cats.

PubMed

Genchi, Marco; Mangia, Carlo; Ferrari, Nicola; Loukeri, Sofia

2018-01-01

The performance of a rapid immunochromatographic test for the detection of Dirofilaria immitis antigens (Speed Diro™; BVT-Virbac, France) was assessed in 49 experimentally infected dogs and in 244 naturally infected animals; 142 dogs and 102 cats. In experimentally infected dogs, Speed Diro™ showed a sensitivity of 90.9% in dogs infected with one adult female worm and 100% in dogs infected with more than one female worm. Specificity was 100%. For naturally infected dogs, the Knott test and PetChek ® HTWM PF served as reference methods for microfilaremia and antigenemia, respectively. All microfilaemic dogs (55/142) were positive with Speed Diro™. Importantly, none of the 21 dogs infected with D. repens were positive. The results of Speed Diro™ for the detection of antigenemia were compared with two in-house tests, SNAP ® HTWM and Witness ® Dirofilaria, and all three tests were 100% specific and sensitive in comparison to PetChek ® HTWM PF. For the evaluation of feline samples, 102 cats were examined by echocardiography. Sera from 87 heartworm-infected cats were tested by Speed Diro™ and SNAP ® HTWM. The results of Speed Diro™ were equivalent to SNAP ® HTWM, with a sensitivity of 98.9% and a specificity of 100%.

Evaluation of the Clearview® malaria pLDH malaria rapid diagnostic test in a non-endemic setting

PubMed Central

2011-01-01

Background Malaria Rapid Diagnostic Tests (RDTs) are widely used to diagnose malaria. The present study evaluated a new RDT, the Clearview® Malaria pLDH test targeting the pan-Plasmodium antigen lactate dehydrogenase (pLDH). Methods The Clearview® Malaria pLDH test was evaluated on fresh samples obtained in returned international travellers using microscopy corrected by PCR as the reference method. Included samples were Plasmodium falciparum (139), Plasmodium vivax (22), Plasmodium ovale (20), Plasmodium malariae (7), and 102 negative. Results Overall sensitivity for the detection of Plasmodium spp was 93.2%. For P. falciparum, the sensitivity was 98.6%; for P. vivax, P. ovale and P. malariae, overall sensitivities were 90.9%, 60.0% and 85.7% respectively. For P. falciparum and for P. vivax, the sensitivities increased to 100% at parasite densities above 100/μl. The specificity was 100%. The test was easily to perform and the result was stable for at least 1 hour. Conclusion The Clearview® Malaria pLDH was efficient for the diagnosis of malaria. The test was very sensitive for P. falciparum and P. vivax detection. The sensitivities for P. ovale and P. malariae were better than other RDTs PMID:21951996

Recombinant antigens expressed in Pichia pastoris for the diagnosis of sleeping sickness caused by Trypanosoma brucei gambiense.

PubMed

Rogé, Stijn; Van Nieuwenhove, Liesbeth; Meul, Magali; Heykers, Annick; Brouwer de Koning, Annette; Bebronne, Nicolas; Guisez, Yves; Büscher, Philippe

2014-07-01

Screening tests for gambiense sleeping sickness, such as the CATT/T. b. gambiense and a recently developed lateral flow tests, are hitherto based on native variant surface glycoproteins (VSGs), namely LiTat 1.3 and LiTat 1.5, purified from highly virulent trypanosome strains grown in rodents. We have expressed SUMO (small ubiquitin-like modifier) fusion proteins of the immunogenic N-terminal part of these antigens in the yeast Pichia pastoris. The secreted recombinant proteins were affinity purified with yields up to 10 mg per liter cell culture. The diagnostic potential of each separate antigen and a mixture of both antigens was confirmed in ELISA on sera from 88 HAT patients and 74 endemic non-HAT controls. Replacement of native antigens in the screening tests for sleeping sickness by recombinant proteins will eliminate both the infection risk for the laboratory staff during antigen production and the need for laboratory animals. Upscaling production of recombinant antigens, e.g. in biofermentors, is straightforward thus leading to improved standardisation of antigen production and reduced production costs, which on their turn will increase the availability and affordability of the diagnostic tests needed for the elimination of gambiense HAT.

Prevalence of human immunodeficiency virus type 1 p24 antigen in U.S. blood donors--an assessment of the efficacy of testing in donor screening. The HIV-Antigen Study Group.

PubMed

Alter, H J; Epstein, J S; Swenson, S G; VanRaden, M J; Ward, J W; Kaslow, R A; Menitove, J E; Klein, H G; Sandler, S G; Sayers, M H

1990-11-08

We performed a multicenter study in 1989 to determine whether screening whole-blood donors for human immunodeficiency virus type 1 (HIV-1) p24 antigen would improve transfusion safety by identifying carriers of the virus who are seronegative for HIV-1 antibody. More than 500,000 donations were tested at 13 U.S. blood centers with test kits from two manufacturers. Units found repeatedly reactive were retested in a central laboratory; if the results were positive, they were confirmed by a neutralization assay. A subgroup of units was also tested for HIV-1 by the polymerase chain reaction. Selected donors confirmed or not confirmed as having p24 antigen were contacted for follow-up interviews to identify risk factors and undergo retesting for HIV-1 markers. Positive tests for p24 antigen were confirmed by neutralization in five donors (0.001 percent of all donations tested), all of whom were also positive for HIV-1 antibody and HIV-1 by polymerase chain reaction. Three of the antigen-positive donors had other markers of infectious disease that would have resulted in the exclusion of their blood; two had risk factors for HIV-1 that should have led to self-exclusion. Of 220 blood units with repeatedly reactive p24 antigen whose presence could not be confirmed by neutralization (0.04 percent of the donations studied), none were positive for HIV-1 antibody, HIV-1 by polymerase chain reaction (120 units tested), or virus culture (76 units tested)--attesting to the specificity of confirmatory neutralization. The finding that no donation studied was positive for p24 antigen and negative for HIV-1 antibody suggests that screening donors for p24 antigen with tests of the current level of sensitivity would not add substantially to the safety of the U.S. blood supply.

Rapid viral diagnosis of acute respiratory infections: comparison of enzyme-linked immunosorbent assay and the immunofluorescence technique for detection of viral antigens in nasopharyngeal secretions.

PubMed Central

Grandien, M; Pettersson, C A; Gardner, P S; Linde, A; Stanton, A

1985-01-01

Nasopharyngeal secretions from adults and children were obtained in Stockholm, Sweden, for routine diagnosis of influenza A virus, influenza B virus, respiratory syncytial (RS) virus, parainfluenza type 3 virus, and adenovirus infections by demonstration of viral antigens directly in the specimens. The cells in nasopharyngeal secretions were pelleted by centrifugation for preparation of cell deposits for diagnosis by the immunofluorescence technique (IF) in London, England, and in Stockholm, whereas the supernatants were used to diagnose infection by the enzyme-linked immunosorbent assay (ELISA) in Stockholm. Titrations of the various purified viruses showed that ELISA could detect viral antigens in amounts corresponding to 1 to 10 ng of virus protein per test well. In a series of 73 specimens tested for influenza A, RS, and parainfluenza type 3 viruses by IF in London and by ELISA in Stockholm, 15 of 18 RS, 14 of 15 influenza A, and 2 of 2 parainfluenza type 3 viral infections were diagnosed by ELISA as compared with IF, giving sensitivities for RS and influenza A viral diagnosis of 83 and 93%, respectively, and a specificity of 100%. In another series of specimens from 35 patients tested for influenza B virus and adenovirus, five influenza B virus and four adenovirus infections were diagnosed by both methods; one additional influenza B infection was detected only by IF and another only by ELISA. Comparisons of diagnostic results between the two methods performed in Stockholm gave nonagreement of results for 37 of 1,593 tests (2.5%) for the five viruses. The conclusion reached was that the described ELISA, although a satisfactory test, had somewhat less sensitivity than did IF for the detection of respiratory viral infections. This could possibly be explained by unnecessary dilutions of specimens at the time of collection; transportation, processing, and storage of specimens were less complicated than for IF. PMID:2997270

Novel Single-Tube Agar-Based Test System for Motility Enhancement and Immunocapture of Escherichia coli O157:H7 by H7 Flagellar Antigen-Specific Antibodies

PubMed Central

Murinda, Shelton E.; Nguyen, Lien T.; Ivey, Susan J.; Almeida, Raul A.; Oliver, Stephen P.

2002-01-01

This paper describes a novel single-tube agar-based technique for motility enhancement and immunoimmobilization of Escherichia coli O157:H7. Motility indole ornithine medium and agar (0.4%, wt/vol) media containing either nutrient broth, tryptone broth, or tryptic soy broth (TSBA) were evaluated for their abilities to enhance bacterial motility. Twenty-six E. coli strains, including 19 O157:H7 strains, 1 O157:H− strain, and 6 generic E. coli strains, were evaluated. Test bacteria were stab inoculated in the center of the agar column, and tubes were incubated at 37°C for 18 to 96 h. Nineteen to 24 of the 26 test strains (73.1 to 92.3%) were motile in the different media. TSBA medium performed best and was employed in subsequent studies of motility enhancement and H7 flagellar immunocapture. H7 flagellar antiserum (30 and 60 μl) mixed with TSBA was placed as a band (1 ml) in the middle of an agar column separating the top (3-ml) and bottom (3-ml) agar layers. The top agar layer was inoculated with the test bacterial strains. The tubes were incubated at 37°C for 12 to 18 h and for 18 to 96 h. The specificity and sensitivity of the H7 flagellar immunocapture tests were 75 and 100%, respectively. The procedure described is simple and sensitive and could be adapted easily for routine use in laboratories that do not have sophisticated equipment and resources for confirming the presence of H7 flagellar antigens. Accurate and rapid identification of H7 flagellar antigen is critical for the complete characterization of E. coli O157:H7, owing to the immense clinical, public health, and economic significance of this food-borne pathogen. PMID:12454173

Detection of gonococcal infection : pros and cons of a rapid test.

PubMed

Vickerman, Peter; Peeling, Rosanna W; Watts, Charlotte; Mabey, David

2005-01-01

WHO estimates that 62 million cases of gonorrhea occur annually worldwide. Untreated infection can cause serious long-term complications, especially in women. In addition, Neisseria gonorrheae infection can facilitate HIV transmission, and babies born to infected mothers are at risk of ocular infection, which can lead to blindness. Where diagnostic facilities are lacking, gonorrhea can be treated syndromically. However, this inevitably leads to over-treatment, especially in women in whom the syndrome of vaginal discharge may be due not to N. gonorrheae infection but to several other more prevalent conditions. Over-treatment is a major concern because of widespread N. gonorrheae antibiotic resistance. Moreover, a high proportion of gonorrhea cases are asymptomatic and so do not present for syndromic management. Such cases will only be detected by screening tests. The gold standard test for the detection of N. gonorrheae is culture, which has high sensitivity and specificity. However, it requires well trained staff and its performance is affected by specimen transport conditions. Other options include microscopy and tests that detect gonococcal antigen or nucleic acid. Nucleic acid amplification tests (NAATs) have higher sensitivity and can be used on non-invasive samples (urine). However, they can cross-react with other Neisseria species and are expensive, requiring highly trained staff and sophisticated equipment. In settings where patients are asked to return for laboratory results, some infected patients never receive treatment as they fail to return for their test results. This reduction in treatment, and the possible onward transmission of N. gonorrheae during any delay in treatment, means that a rapid test of lower sensitivity may be more effective if it results in patients being treated at the initial visit. Indeed, even with the low sensitivity of currently available rapid tests (50-70%), modeling shows that they can outperform gold standard tests in populations with high sexual activity and/or low return rates. Unfortunately, however, most of the rapid tests currently available are immunoassays that are quite expensive and involve many steps, which limit their current usefulness. In summary, the pros and cons of using a rapid test are dependent on the setting. Culture or NAATs remain the best choice in an ideal setting. However, in settings where laboratory facilities are not available, or in high-risk populations where return rates are low, rapid tests may be the most effective way of diagnosing gonorrhea. Their optimal use in these settings requires the development of simpler and cheaper rapid tests.

Field evaluation of a rapid diagnostic test to detect antibodies in human toxocariasis.

PubMed

Lim, P K C; Yamasaki, H; Mak, J W; Wong, S F; Chong, C W; Yap, I K S; Ambu, S; Kumarasamy, V

2015-08-01

Human toxocariasis which is caused mainly by the larvae of Toxocara canis and Toxocara cati, is a worldwide zoonotic disease that can be a potentially serious human infection. The enzyme-linked immunosorbent assay (ELISA) using T. canis excretory-secretory (TES) antigens harvested from T. canis larvae is currently the serological test for confirming toxocariasis. An alternative to producing large amounts of Toxocara TES and improved diagnosis for toxocariasis is through the development of highly specific recombinant antigens such as the T. canis second stage larva excretory-secretory 30 kDa protein (recTES-30). The aim of this study was to evaluate the sensitivity and specificity of a rapid diagnostic kit (RDT, named as iToxocara kit) in comparison to recTES-30 ELISA in Serendah Orang Asli village in Selangor, Malaysia. A total of 133 subjects were included in the study. The overall prevalence rates by ELISA and RDT were 29.3% and 33.1%, respectively, with more positive cases detected in males than females. However, no association was found between toxocariasis and gender or age. The percentage sensitivity, specificity, positive predictive value and negative predictive value of RDT were 85.7%, 90.1%, 80% and 93.2%, respectively. The prevalence for toxocariasis in this population using both ELISA and RDT was 27.1% (36/133) and the K-concordance test suggested good agreement of the two tests with a Cohen's kappa of 0.722, P<0.01. In addition, the followed-up Spearman rank correlation showed a moderately high correlation at R=0.704 and P<0.01. In conclusion, the RDT kit was faster and easier to use than an ELISA and is useful for the laboratory diagnosis of hospitalized cases of toxocariasis. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Reassessment of the Role of Rapid Antigen Detection Tests in Diagnosis of Invasive Group A Streptococcal Infections.

PubMed

Gazzano, Vincent; Berger, Anne; Benito, Yvonne; Freydiere, Anne-Marie; Tristan, Anne; Boisset, Sandrine; Carricajo, Anne; Poyart, Claire; Vandenesch, François; Descours, Ghislaine

2016-04-01

Rapid antigen detection tests (RADTs) for group A streptococci (GAS) are widely used for diagnosing acute pharyngitis, which has led to a considerable reduction in antibiotic prescriptions over the past decade. Beyond this intended use, their reassessment on invasive samples may be relevant in the management of life-threatening GAS infections. To this end, we evaluated the performances of three RADTs, culture, GAS PCR, and 16S rRNA gene PCR assays, and compared them with a composite gold standard (GAS-PCR assay and/or culture) for the diagnosis of severe GAS infection. A total of 192 specimens from deep-tissue (mostly normally sterile) sites enriched for 75 GAS-positive samples were enrolled in the study. The three evaluated RADTs showed sensitivities ranging from 88.0% to 94.7% versus 98.7% for GAS PCR, 84% for 16S rRNA gene PCR, and 77.3% for culture. The sensitivities of the ImmunoCardSTAT! Strep A test (Meridian Bioscience) and the NADAL Strep A strip (Nal Von Minden) were similar to that of GAS PCR (P= 0.25 and 0.03, respectively) and higher than that of culture (P= 0.001 and 0.006, respectively), whereas the SD Bioline Strep A test strip (Standard Diagnostics) showed a performance similar to that of culture (P= 0.02). The three RADTs detected 10 distinctemmtypes, including a predominance ofemm1 (33.3%),emm89 (10.6%), andemm12 (7.6%). No false-positive results were observed, leading to a specificity of 100% for all the evaluated RADTs. The GAS RADTs turned out to be sensitive, specific, and easy-to-use tools that may aid in the management of invasive GAS infections in 24/7 point-of-care laboratories by enabling early diagnosis and focused therapy. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Reassessment of the Role of Rapid Antigen Detection Tests in Diagnosis of Invasive Group A Streptococcal Infections

PubMed Central

Gazzano, Vincent; Berger, Anne; Benito, Yvonne; Freydiere, Anne-Marie; Tristan, Anne; Boisset, Sandrine; Carricajo, Anne; Poyart, Claire; Vandenesch, François

2016-01-01

Rapid antigen detection tests (RADTs) for group A streptococci (GAS) are widely used for diagnosing acute pharyngitis, which has led to a considerable reduction in antibiotic prescriptions over the past decade. Beyond this intended use, their reassessment on invasive samples may be relevant in the management of life-threatening GAS infections. To this end, we evaluated the performances of three RADTs, culture, GAS PCR, and 16S rRNA gene PCR assays, and compared them with a composite gold standard (GAS-PCR assay and/or culture) for the diagnosis of severe GAS infection. A total of 192 specimens from deep-tissue (mostly normally sterile) sites enriched for 75 GAS-positive samples were enrolled in the study. The three evaluated RADTs showed sensitivities ranging from 88.0% to 94.7% versus 98.7% for GAS PCR, 84% for 16S rRNA gene PCR, and 77.3% for culture. The sensitivities of the ImmunoCard STAT! Strep A test (Meridian Bioscience) and the NADAL Strep A strip (Nal Von Minden) were similar to that of GAS PCR (P = 0.25 and 0.03, respectively) and higher than that of culture (P = 0.001 and 0.006, respectively), whereas the SD Bioline Strep A test strip (Standard Diagnostics) showed a performance similar to that of culture (P = 0.02). The three RADTs detected 10 distinct emm types, including a predominance of emm 1 (33.3%), emm 89 (10.6%), and emm 12 (7.6%). No false-positive results were observed, leading to a specificity of 100% for all the evaluated RADTs. The GAS RADTs turned out to be sensitive, specific, and easy-to-use tools that may aid in the management of invasive GAS infections in 24/7 point-of-care laboratories by enabling early diagnosis and focused therapy. PMID:26818671

Immunochromatographic antigen testing alone is sufficient to identify asymptomatic refugees at risk of severe malaria presenting to a single health service in Victoria.

PubMed

Fedele, Pasquale L; Wheeler, Michael; Lemoh, Christopher; Chunilal, Sanjeev

2014-10-01

Current screening guidelines for malaria in new refugees include a combination of thick and thin film examination and immunochromatographic antigen test (ICT). However, as the prevalence of malaria in our population has decreased due to changing refugee demographics, we sought to determine if an ICT alone can reliably exclude malaria in our asymptomatic refugee population.A retrospective analysis was conducted of all investigations for malaria performed from 1 August 2011 to 31 July 2013, including thick and thin blood film examination, BinaxNOW ICT, and external morphological and polymerase chain reaction (PCR) validation where applicable.Malaria was diagnosed in 45 of 1248 (3.6%) patients investigated, all of whom were symptomatic and the majority (71.1%) returned travellers. All 599 asymptomatic refugees screened were negative. Overall, 42 of 45 malaria cases were detected by the ICT; sensitivity 93.3% (95% CI 80.7-98.3%) and negative predictive value (NPV) 99.8% (99.2-99.9%). All 21 cases of Plasmodium falciparum and 20 of 22 cases of Plasmodium vivax were detected, giving a sensitivity of 100% (80.8-100%) and 90.9% (69.4-98.4%) respectively. Too few cases of Plasmodium malariae and no cases of Plasmodium ovale or Plasmodium knowlesi were diagnosed for adequate assessment to be carried out.These data suggest that full malaria screening in all asymptomatic refugees with the combination of thick and thin blood films and rapid antigen test may not be warranted. Alternative screening approaches should be considered, including the use of ICT alone, or limiting screening of asymptomatic refugees to only those originating from countries with high incidence of malaria.

Design, fabrication, and characterization of polymeric bioMEMS for the detection of feline immunodeficiency virus (FIV)

NASA Astrophysics Data System (ADS)

Cohen, Brian; Gadre, Anand; Kaloyeros, Alain E.

2007-02-01

This project comprises the development of a novel polymeric BioMEMS device capable of rapidly detecting FIV in a minimally invasive manner. FIV severely inhibits the infected feline from mounting an immune response, and causes susceptibility to other types of diseases. Vaccines against FIV do exist, but have some strong limitations to their effectiveness; so early detection is the best method to combat the spread of the disease. Current testing methods look for antibodies to the FIV protein p24 in feline blood using established Enzyme Linked ImmunoSorbent Assay (ELISA) protocols. The focus of this research is to design and construct a device that can detect antibodies to p24 in a salivary sample by non-intrusive electrochemical means. The device is constructed upon a silicon substrate with gold microelectrodes coated with polypyrrole (PPy), an electrically conducting and biocompatible polymer. In the current phase of the research, the PPy deposition process has been optimized with regards to film thickness, uniformity and conductivity. Microfluidic channels have been fabricated using SU-8, an epoxy based polymer that enables the test sample and other solutions to pass freely through the device. The PPy will be coated with anti-FIV p24 antibodies that can capture FIV p24 antigens present in a salivary sample. Future research will involve the analysis of PPy/antibody interaction and its effect on functionality. The capture of such antigens will interfere with a reduction-oxidation (redox) reaction in a subsequently added ionic solution. This interference will change the characteristic resistance of the solution yielding a qualitative test for the presence of the viral antigens in the sample and hence determining the occurrence of infection.

Meta-analysis: accuracy of rapid tests for malaria in travelers returning from endemic areas.

PubMed

Marx, Arthur; Pewsner, Daniel; Egger, Matthias; Nüesch, Reto; Bucher, Heiner C; Genton, Blaise; Hatz, Christoph; Jüni, Peter

2005-05-17

Microscopic diagnosis of malaria is unreliable outside specialized centers. Rapid tests have become available in recent years, but their accuracy has not been assessed systematically. To determine the accuracy of rapid diagnostic tests for ruling out malaria in nonimmune travelers returning from malaria-endemic areas. The authors searched MEDLINE, EMBASE, CAB Health, and CINAHL (1988 to September 2004); hand-searched conference proceedings; checked reference lists; and contacted experts and manufacturers. Diagnostic accuracy studies in nonimmune individuals with suspected malaria were included if they compared rapid tests with expert microscopic examination or polymerase chain reaction tests. Data on study and patient characteristics and results were extracted in duplicate. The main outcome was the likelihood ratio for a negative test result (negative likelihood ratio) for Plasmodium falciparum malaria. Likelihood ratios were combined by using random-effects meta-analysis, stratified by the antigen targeted (histidine-rich protein-2 [HRP-2] or parasite lactate dehydrogenase [LDH]) and by test generation. Nomograms of post-test probabilities were constructed. The authors included 21 studies and 5747 individuals. For P. falciparum, HRP-2-based tests were more accurate than parasite LDH-based tests: Negative likelihood ratios were 0.08 and 0.13, respectively (P = 0.019 for difference). Three-band HRP-2 tests had similar negative likelihood ratios but higher positive likelihood ratios compared with 2-band tests (34.7 vs. 98.5; P = 0.003). For P. vivax, negative likelihood ratios tended to be closer to 1.0 for HRP-2-based tests than for parasite LDH-based tests (0.24 vs. 0.13; P = 0.22), but analyses were based on a few heterogeneous studies. Negative likelihood ratios for the diagnosis of P. malariae or P. ovale were close to 1.0 for both types of tests. In febrile travelers returning from sub-Saharan Africa, the typical probability of P. falciparum malaria is estimated at 1.1% (95% CI, 0.6% to 1.9%) after a negative 3-band HRP-2 test result and 97% (CI, 92% to 99%) after a positive test result. Few studies evaluated 3-band HRP-2 tests. The evidence is also limited for species other than P. falciparum because of the few available studies and their more heterogeneous results. Further studies are needed to determine whether the use of rapid diagnostic tests improves outcomes in returning travelers with suspected malaria. Rapid malaria tests may be a useful diagnostic adjunct to microscopy in centers without major expertise in tropical medicine. Initial decisions on treatment initiation and choice of antimalarial drugs can be based on travel history and post-test probabilities after rapid testing. Expert microscopy is still required for species identification and confirmation.

Diagnosis of group A streptococcal infections directly from throat secretions.

PubMed Central

Edwards, E A; Phillips, I A; Suiter, W C

1982-01-01

The diagnosis of group A streptococcal disease still relies on isolation of group A streptococcal strains on sheep blood agar followed by presumptive identification based on bacitracin sensitivity or the results of the more precise serogrouping methods such as the Lancefield precipitin test. A technique that would permit rapid identification of streptococcal infections directly from throat secretions would allow immediate appropriate antimicrobial therapy for the management of streptococcal infections to be started. We have been able to identify soluble group A antigen directly from throat secretions by using a latex agglutination test. In a clinical trial in which latex (Streptex group A) and conventional culturing techniques were used, 53 throat secretion cultures were tested: 26 were positive by both procedures, 5 were positive by culture only, 3 were positive by the latex agglutination test only, and 19 were negative by both tests. Images PMID:7042747

A simple semi-rapid HIV-1&2 confirmatory immunoassay using magnetic particles.

PubMed

Sommerfelt, Maja A; Ohlsson, Irene; Flolid, Irene; Thorstensson, Rigmor; Sørensen, Birger

2004-02-01

The Bionor HIV-1&2 Confirmatory Test is a semi-rapid simple immunoassay based on magnetic particles for the confirmation of serological status to human immunodeficiency virus (HIV). The specificity and sensitivity of this assay was evaluated by comparison with the Diagnostic Biotechnology HIV-1 Western blot (WB) 2.2 and the HIV-2/SBL-6669 WB. Bionor's confirmatory test demonstrated 98% specificity when testing sero-negative blood donors and false positive sera in screening tests compared to 81.5 and 71.6%, respectively, using the HIV-1 WB. The sensitivity of this assay for HIV-1 antibody positive sera was 97.9% compared to the WB which was 99.5%. When testing confirmed HIV-2 antibody positive samples, 2/100 scored negative using this confirmatory test similar to other HIV-2 peptide-based line immunoassays available commercially, whilst 8/100 were indeterminate reacting to HIV-2 membrane antigens only. Bionor's confirmatory test detected HIV-1 seropositivity earlier than the WB in longitudinal seroconversion panels and could discriminate between HIV-1 and -2 infection. The number of indeterminate responses was generally reduced significantly using Bionor's confirmatory test compared to the HIV-1 WB. The greater specificity, speed and ease of interpretation of Bionor's confirmatory test renders it an attractive and cost effective alternative to the WB for confirming HIV serological status worldwide.

Predicting pneumococcal community-acquired pneumonia in the emergency department: evaluation of clinical parameters.

PubMed

Huijts, S M; Boersma, W G; Grobbee, D E; Gruber, W C; Jansen, K U; Kluytmans, J A J W; Kuipers, B A F; Palmen, F; Pride, M W; Webber, C; Bonten, M J M

2014-12-01

The aim of this study was to quantify the value of clinical predictors available in the emergency department (ED) in predicting Streptococcus pneumoniae as the cause of community-acquired pneumonia (CAP). A prospective, observational, cohort study of patients with CAP presenting in the ED was performed. Pneumococcal aetiology of CAP was based on either bacteraemia, or S. pneumoniae being cultured from sputum, or urinary immunochromatographic assay positivity, or positivity of a novel serotype-specific urinary antigen detection test. Multivariate logistic regression was used to identify independent predictors and various cut-off values of probability scores were used to evaluate the usefulness of the model. Three hundred and twenty-eight (31.0%) of 1057 patients with CAP had pneumococcal CAP. Nine independent predictors for pneumococcal pneumonia were identified, but the clinical utility of this prediction model was disappointing, because of low positive predictive values or a small yield. Clinical criteria have insufficient diagnostic capacity to predict pneumococcal CAP. Rapid antigen detection tests are needed to diagnose S. pneumoniae at the time of hospital admission. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Antibodies Targeting Closely Adjacent or Minimally Overlapping Epitopes Can Displace One Another

PubMed Central

Abdiche, Yasmina Noubia; Yeung, Andy Yik; Ni, Irene; Stone, Donna; Miles, Adam; Morishige, Winse; Rossi, Andrea; Strop, Pavel

2017-01-01

Here we describe how real-time label-free biosensors can be used to identify antibodies that compete for closely adjacent or minimally overlapping epitopes on their specific antigen via a mechanism of antibody displacement. By kinetically perturbing one another’s binding towards their antigen via the formation of a transient trimolecular complex, antibodies can displace one another in a fully reversible and dose-dependent manner. Displacements can be readily identified when epitope binning assays are performed in a classical sandwich assay format whereby a solution antibody (analyte) is tested for binding to its antigen that is first captured via an immobilized antibody (ligand) because an inverted sandwiching response is observed when an analyte displaces a ligand, signifying the antigen’s unusually rapid dissociation from its ligand. In addition to classifying antibodies within a panel in terms of their ability to block or sandwich pair with one another, displacement provides a hybrid mechanism of competition. Using high-throughput epitope binning studies we demonstrate that displacements can be observed on any target, if the antibody panel contains appropriate epitope diversity. Unidirectional displacements occurring between disparate-affinity antibodies can generate apparent asymmetries in a cross-blocking experiment, confounding their interpretation. However, examining competition across a wide enough concentration range will often reveal that these displacements are reversible. Displacement provides a gentle and efficient way of eluting antigen from an otherwise high affinity binding partner which can be leveraged in designing reagents or therapeutic antibodies with unique properties. PMID:28060885

Endogenous T cell responses to antigens expressed in lung adenocarcinomas delay malignant tumor progression

PubMed Central

DuPage, Michel; Cheung, Ann; Mazumdar, Claire; Winslow, Monte M.; Bronson, Roderick; Schmidt, Leah M.; Crowley, Denise; Chen, Jianzhu; Jacks, Tyler

2010-01-01

SUMMARY Neoantigens derived from somatic mutations in tumors may provide a critical link between the adaptive immune system and cancer. Here we describe a system to introduce exogenous antigens into genetically engineered mouse lung cancers to mimic tumor neoantigens. We show that endogenous T cells respond to and infiltrate tumors, significantly delaying malignant progression. Despite continued antigen expression, T cell infiltration does not persist and tumors ultimately escape immune attack. Transplantation of cell lines derived from these lung tumors or prophylactic vaccination against the autochthonous tumors, however, results in rapid tumor eradication or selection of tumors that lose antigen expression. These results provide insight into the dynamic nature of the immune response to naturally arising tumors. PMID:21251614

Serodiagnosis of infectious mononucleosis by using recombinant Epstein-Barr virus antigens and enzyme-linked immunosorbent assay technology.

PubMed Central

Gorgievski-Hrisoho, M; Hinderer, W; Nebel-Schickel, H; Horn, J; Vornhagen, R; Sonneborn, H H; Wolf, H; Siegl, G

1990-01-01

Four recombinant, diagnostically useful Epstein-Barr virus (EBV) proteins representative of the viral capsid antigen (p150), diffuse early antigen (p54), the major DNA-binding protein (p138), and the EBV nuclear antigen (p72) (W. Hinderer, H. Nebel-Schickel, H.H. Sonneborn, M. Motz, R. Kühbeck, and H. Wolf, J. Exp. Clin. Cancer Res. 7[Suppl.]:132, 1988) were used to set up individual enzyme-linked immunosorbent assays (ELISAs) for the qualitative and quantitative detection of immunoglobulin M (IgM) and IgG antibodies. In direct comparison with results obtained by standard immunofluorescence or immunoperoxidase assays, it was then shown that the recombinant EBV ELISAs provide the means for specific and sensitive serodiagnosis of infectious mononucleosis (IM) caused by EBV. The most useful markers in sera from such patients proved to be IgM antibodies against p54, p138, and p150. Additional positive markers for recent or ongoing IM apparently were IgG antibodies against p54 and p138. In contrast, anti-p72 IgG had a high preference for sera from healthy blood donors and, therefore, can be considered indicative of past exposure to the virus. Altogether, the individual ELISAs proved to be as specific and at least as sensitive for the diagnosis of IM as the currently available standard techniques are. Moreover, our findings suggest that, by combining individual test antigens, a workable ELISA system consisting of three assays (IgM against p54, p138, and p150; IgG against p54 and p138; and IgG against p72) can be established for the standardized rapid diagnosis of acute EBV infections. PMID:2172287

Development of a single-antigen magnetic bead assay (SAMBA) for the sensitive detection of HPA-1a alloantibodies using tag-engineered recombinant soluble β3 integrin.

PubMed

Skaik, Younis; Battermann, Anja; Hiller, Oliver; Meyer, Oliver; Figueiredo, Constanca; Salama, Abdulgabar; Blasczyk, Rainer

2013-05-31

Timely and accurate testing for human platelet antigen 1a (HPA-1a) alloantibodies is vital for clinical diagnosis of neonatal alloimmune thrombocytopenia (NAIT). Current antigen-specific assays used for the detection of HPA-1 alloantibodies are technically very complex and cumbersome for most diagnostic laboratories. Hence, we designed and applied recombinant soluble (rs) β3 integrins displaying HPA-1a or HPA-1b epitopes for the development of a single-antigen magnetic bead assay (SAMBA). Soluble HPA-1a and HPA-1b were produced recombinantly in human embryonic kidney 293 (HEK293) cells and differentially tagged. The recombinant soluble proteins were then immobilized onto paramagnetic beads and used for analysis of HPA-1 alloantibodies by enzyme-linked immunosorbent assay (ELISA). HPA-1a serum samples (n=7) from NAIT patients, inert sera and sera containing non-HPA-1a antibodies were used to evaluate the sensitivity and specificity of the SAMBA. Fusion of V5-His or GS-SBP-His tags to the rsβ3 integrins resulted in high-yield expression. SAMBA was able to detect all HPA-1a and -1b alloantibodies recognized by monoclonal antibody-specific immobilization of platelet antigens assay (MAIPA). No cross-reactions between the sera were observed. Two out of seven of the HPA-1a alloantibody-containing sera demonstrated weak to moderate reactivity in MAIPA but strong signals in SAMBA. SAMBA provides a very reliable method for the detection of HPA-1 antibodies with high specificity and sensitivity. This simple and rapid assay can be adapted for use in any routine laboratory and can be potentially adapted for use on automated systems. Copyright © 2013 Elsevier B.V. All rights reserved.

The role and mechanics of dendritic cells in tumor antigen acquisition and presentation following laser immunotherapy

NASA Astrophysics Data System (ADS)

Laverty, Sean M.; Dawkins, Bryan A.; Chen, Wei R.

2018-02-01

We extend our model of the antitumor immune response initiated by laser-immunotherapy treatment to more closely examine key steps in the immune response 1) tumor antigen acquisition by antigen-presenting dendritic cells (DCs) and 2) cytotoxic T cell (CTL) priming by lymphatic DCs. Specifically we explore the formation of DC-CTL complexes that lead to CTL priming. We find that the bias in the dissociation rate of the complex influences the outcome of treatment. In particular, a bias towards priming favors a rapid activated CTL response and the clearance of tumors.

Molecular analysis of antigen-independent adhesion forces between T and B lymphocytes.

PubMed Central

Amblard, F; Auffray, C; Sekaly, R; Fischer, A

1994-01-01

The low-affinity interactions underlying antigen recognition by T-cell receptors (TCRs) are thought to involve antigen-independent adhesion mechanisms. Using a hydrodynamic approach, we found that antigen-independent adhesion occurred between human B cells and resting T cells in a transient and temperature-dependent fashion. The mean cell-cell adhesion force was 0.32 x 10(-9) N and was generated by similar contributions (0.16 x 10(-9) N) of the LFA-1- and CD2-dependent adhesion pathways. After T-cell stimulation with a phorbol ester, the force contributed by LFA-1 was drastically increased, while that of CD2 was unaffected. We propose that weak receptor-mediated adhesion initiates antigen-independent intercellular contacts required for antigen recognition by the TCR and is upregulated following TCR engagement. The method used permits adhesion forces between living cells to be resolved at the molecular level and should prove valuable for the rapid assessment of interaction forces between various types of cells and cell-sized particles. Images PMID:7909604

Pooled protein immunization for identification of cell surface antigens in Streptococcus sanguinis.

PubMed

Ge, Xiuchun; Kitten, Todd; Munro, Cindy L; Conrad, Daniel H; Xu, Ping

2010-07-26

Available bacterial genomes provide opportunities for screening vaccines by reverse vaccinology. Efficient identification of surface antigens is required to reduce time and animal cost in this technology. We developed an approach to identify surface antigens rapidly in Streptococcus sanguinis, a common infective endocarditis causative species. We applied bioinformatics for antigen prediction and pooled antigens for immunization. Forty-seven surface-exposed proteins including 28 lipoproteins and 19 cell wall-anchored proteins were chosen based on computer algorithms and comparative genomic analyses. Eight proteins among these candidates and 2 other proteins were pooled together to immunize rabbits. The antiserum reacted strongly with each protein and with S. sanguinis whole cells. Affinity chromatography was used to purify the antibodies to 9 of the antigen pool components. Competitive ELISA and FACS results indicated that these 9 proteins were exposed on S. sanguinis cell surfaces. The purified antibodies had demonstrable opsonic activity. The results indicate that immunization with pooled proteins, in combination with affinity purification, and comprehensive immunological assays may facilitate cell surface antigen identification to combat infectious diseases.

Pooled Protein Immunization for Identification of Cell Surface Antigens in Streptococcus sanguinis

PubMed Central

Ge, Xiuchun; Kitten, Todd; Munro, Cindy L.; Conrad, Daniel H.; Xu, Ping

2010-01-01

Background Available bacterial genomes provide opportunities for screening vaccines by reverse vaccinology. Efficient identification of surface antigens is required to reduce time and animal cost in this technology. We developed an approach to identify surface antigens rapidly in Streptococcus sanguinis, a common infective endocarditis causative species. Methods and Findings We applied bioinformatics for antigen prediction and pooled antigens for immunization. Forty-seven surface-exposed proteins including 28 lipoproteins and 19 cell wall-anchored proteins were chosen based on computer algorithms and comparative genomic analyses. Eight proteins among these candidates and 2 other proteins were pooled together to immunize rabbits. The antiserum reacted strongly with each protein and with S. sanguinis whole cells. Affinity chromatography was used to purify the antibodies to 9 of the antigen pool components. Competitive ELISA and FACS results indicated that these 9 proteins were exposed on S. sanguinis cell surfaces. The purified antibodies had demonstrable opsonic activity. Conclusions The results indicate that immunization with pooled proteins, in combination with affinity purification, and comprehensive immunological assays may facilitate cell surface antigen identification to combat infectious diseases. PMID:20668678

Immunization Route Dictates Cross-Priming Efficiency and Impacts the Optimal Timing of Adjuvant Delivery

PubMed Central

Bouvier, Isabelle; Jusforgues-Saklani, Hélène; Lim, Annick; Lemaître, Fabrice; Lemercier, Brigitte; Auriau, Charlotte; Nicola, Marie-Anne; Leroy, Sandrine; Law, Helen K.; Bandeira, Antonio; Moon, James J.; Bousso, Philippe; Albert, Matthew L.

2011-01-01

Delivery of cell-associated antigen represents an important strategy for vaccination. While many experimental models have been developed in order to define the critical parameters for efficient cross-priming, few have utilized quantitative methods that permit the study of the endogenous repertoire. Comparing different strategies of immunization, we report that local delivery of cell-associated antigen results in delayed T cell cross-priming due to the increased time required for antigen capture and presentation. In comparison, delivery of disseminated antigen resulted in rapid T cell priming. Surprisingly, local injection of cell-associated antigen, while slower, resulted in the differentiation of a more robust, polyfunctional, effector response. We also evaluated the combination of cell-associated antigen with poly I:C delivery and observed an immunization route-specific effect regarding the optimal timing of innate immune stimulation. These studies highlight the importance of considering the timing and persistence of antigen presentation, and suggest that intradermal injection with delayed adjuvant delivery is the optimal strategy for achieving CD8+ T cell cross-priming. PMID:22566860

Prostate-specific antigen velocity is not better than total prostate-specific antigen in predicting prostate biopsy diagnosis.

PubMed

Gorday, William; Sadrzadeh, Hossein; de Koning, Lawrence; Naugler, Christopher T

2015-12-01

1.) Identify whether prostate-specific antigen velocity improves the ability to predict prostate biopsy diagnosis. 2.) Test whether there is an increase in the predictive capability of models when Gleason 7 prostate cancers are separated into a 3+4 and a 4+3 group. Calgary Laboratory Services' Clinical Laboratory Information System was searched for prostate biopsies reported between January 1, 2009 and December 31, 2013. Total prostate-specific antigen tests were recorded for each patient from January 1, 2007 to the most recent test before their recorded prostate biopsy. The data set was divided into the following three groups for comparison; benign, all prostate cancer and Gleason 7-10. The Gleason grade 7-10 group was further divided into 4+3 and 3+4 Gleason 7 prostate cancers. Prostate-specific antigen velocity was calculated using four different methods found in the literature. Receiver operator curves were used to assess operational characteristics of the tests. 4622 men between the ages of 40-89 with a prostate biopsy were included for analysis. Combining prostate-specific antigen velocity with total prostate-specific antigen (AUC=0.570-0.712) resulted in small non-statistically significant changes to the area under the curve compared to the area under the curve of total prostate-specific antigen alone (AUC=0.572-0.699). There were marked increases in the area under curves when 3+4 and 4+3 Gleason 7 cancers were separated. Prostate-specific antigen velocity does not add predictive value for prostate biopsy diagnosis. The clinical significance of the prostate specific antigen test can be improved by separating Gleason 7 prostate cancers into a 3+4 and 4+3 group. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

[Diagnosis of streptococcal pharyngitides].

PubMed

Matas, Lurdes; Méndez, María; Rodrigo, Carlos; Ausina, Vicente

2008-11-01

Tonsillitis and pharyngitis are highly frequent infections in Spain with multiple causes, especially viruses. Etiologic diagnosis of pharyngitis is only important in Streptococcus pyogenes or group A beta-hemolytic streptococcal infections because antibiotic therapy is mandatory to prevent further complications, especially acute rheumatic fever. Diagnosis has classically been based on bacterial isolation from throat swabs. There are no standardized methods for this culture in terms of culture medium, incubation atmosphere or the identification tests to be performed. Currently, a large number of commercial reagents are available, which, through different immunological techniques, allow direct antigen detection in the clinical sample. Most of these products show good sensitivity and specificity values, with the added advantage of providing a rapid result, especially if they are performed at points of care. Many guidelines for the management of patients with pharyngitis and tonsillitis are available. Nevertheless, as antigen detection results can vary in each center according to the indications and the care taken in sample extraction, the method chosen should be evaluated before implementation.

In vitro Fab display: a cell-free system for IgG discovery

PubMed Central

Stafford, Ryan L.; Matsumoto, Marissa L.; Yin, Gang; Cai, Qi; Fung, Juan Jose; Stephenson, Heather; Gill, Avinash; You, Monica; Lin, Shwu-Hwa; Wang, Willie D.; Masikat, Mary Rose; Li, Xiaofan; Penta, Kalyani; Steiner, Alex R.; Baliga, Ramesh; Murray, Christopher J.; Thanos, Christopher D.; Hallam, Trevor J.; Sato, Aaron K.

2014-01-01

Selection technologies such as ribosome display enable the rapid discovery of novel antibody fragments entirely in vitro. It has been assumed that the open nature of the cell-free reactions used in these technologies limits selections to single-chain protein fragments. We present a simple approach for the selection of multi-chain proteins, such as antibody Fab fragments, using ribosome display. Specifically, we show that a two-chain trastuzumab (Herceptin) Fab domain can be displayed in a format which tethers either the heavy or light chain to the ribosome while retaining functional antigen binding. Then, we constructed synthetic Fab HC and LC libraries and performed test selections against carcinoembryonic antigen (CEA) and vascular endothelial growth factor (VEGF). The Fab selection output was reformatted into full-length immunoglobulin Gs (IgGs) and directly expressed at high levels in an optimized cell-free system for immediate screening, purification and characterization. Several novel IgGs were identified using this cell-free platform that bind to purified CEA, CEA positive cells and VEGF. PMID:24586053

Diagnosis of schistosomiasis japonica with interfacial co-assembly-based multi-channel electrochemical immunosensor arrays

NASA Astrophysics Data System (ADS)

Deng, Wangping; Xu, Bin; Hu, Haiyan; Li, Jianyong; Hu, Wei; Song, Shiping; Feng, Zheng; Fan, Chunhai

2013-05-01

Schistosomiasis control remains to be an important and challenging task in the world. However, lack of quick, simple, sensitive and specific sero-diagnostic test is still a hurdle in the control practice. The commonly employed enzyme-linked immuno-sorbent assay (ELISA) relies on the native soluble egg antigen (SEA) that is limited in supply. Here we developed an electrochemical immunosensor array (ECISA) assay with an interfacial co-assembly strategy. A recombinant Schistosoma japonicum (Sj) calcium-binding protein (SjE16) was used as a principal antigen, while the SEA as a minor, co-assembling agent, with a ratio of 8:1 (SjE16: SEA, Sj16EA), which was co-immobilized on a disposable 16-channel screen-printed carbon electrode array. A portable electrochemical detector was employed to detect antibodies in serum samples. The sensitivity of ECISA reached 100% with minimal cross-reactions. Therefore, we have demonstrated that this rapid, sensitive and specific ECISA technique has the potential to perform large-scale on-site screening of Sj infection.

The evaluation of three diagnostic tests for the detection of equine influenza nucleoprotein in nasal swabs.

PubMed

Galvin, Pamela; Gildea, Sarah; Nelly, Maura; Quinlivan, Michelle; Arkins, Sean; Walsh, Cathal; Cullinane, Ann

2014-05-01

Equine influenza (EI) is a highly contagious respiratory disease of horses. The aim of this study was to evaluate two rapid antigen detection kits (Directigen or DFA, and Espline) and a commercial ELISA for the detection of EI nucleoprotein in nasal swabs. Nasal swab samples from naturally and experimentally infected horses were used to compare the sensitivity and specificity of these assays to virus isolation (VI) and real-time RT-PCR. If real-time RT-PCR was considered as the gold standard, the sensitivity of the other tests in field samples was 68% (DFA), 35% (ELISA), 29% (Espline), and 9% (VI). These tests had 100% specificity when compared to real-time RT-PCR. A receiver operating characteristic (ROC) curve indicated that decreasing the cutoff of the ELISA would increase sensitivity with some loss of specificity. In samples from experimentally infected horses, the sensitivity of the tests compared with real-time RT-PCR was 69% (VI), 27% (DFA), 6% (Espline), and 2% (ELISA). The specificity was 100% for Espline and ELISA and 95% for VI and DFA. This study illustrated that DFA is the most sensitive antigen detection test evaluated for the diagnosis of EI and that it can detect virus in some subclinical infected and vaccinated horses. The results suggest that DFA is a useful adjunct to laboratory tests and may be effective as a screening test in a quarantine station or similar facility where horses are monitored daily. © 2014 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

Long-term antibody response and immunologic memory in children immunized with hepatitis B vaccine at birth.

PubMed

Saffar, M J; Rezai, M S

2004-12-01

Four hundred and fifty three healthy children immunized with a course of hepatitis B vaccine beginning at birth were tested at 10-11 years of age for persistence of anti-hepatitis B-S antigen antibody (anti-HBs); and responses of children without protective antibody to different doses of hepatitis B vaccine booster were evaluated. Although nearly 42% of them were not seroprotected, but most of boosted subjects (87.3%) retained robust immunologic memory and rapidly retained a protective anti-HBs antibody titer of at least 10 IU/L after booster vaccination.

A link between perianal strep and pediatric autoimmune neuropsychiatric disorder associated with streptococcal infection (PANDAS).

PubMed

Toufexis, Megan; Deoleo, Caroline; Elia, Josephine; Murphy, Tanya K

2014-04-01

Perianal streptococcal dermatitis is an infection caused by group A streptococcus (GAS). Children with a pediatric autoimmune neuropsychiatric disorder associated with streptococcal infections (PANDAS) phenotype may have tics or obsessive compulsive symptoms secondary to a systemic immune activation by GAS infecting perianal areas. In this retrospective case series, the authors describe three children with symptoms consistent with PANDAS and a confirmed perianal streptococcal dermatitis as the likely infectious trigger. Concomitant perianal dermatitis and new-onset obsessive-compulsive symptoms and/or tics are strong indications for perianal culture and rapid antigen detection test in young children.

Development of a novel lateral flow assay for detection of African swine fever in blood.

PubMed

Sastre, P; Gallardo, C; Monedero, A; Ruiz, T; Arias, M; Sanz, A; Rueda, P

2016-09-15

African swine fever (ASF) is a viral infectious disease of domestic and wild suids of all breeds and ages, causing a wide range of hemorrhagic syndromes and frequently characterized by high mortality. The disease is endemic in Sub-Saharan Africa and Sardinia. Since 2007, it has also been present in different countries of Eastern Europe, where control measures have not been effective so far. The continued spread poses a serious threat to the swine industry worldwide. In the absence of vaccine, early detection of infected animals is of paramount importance for control of the outbreak, to prevent the transmission of the virus to healthy animals and subsequent spreading of the disease. Current laboratory diagnosis is mainly based on virological methods (antigen and genome detection) and serodiagnosis. In the present work, a Lateral Flow Assay (LFA) for antigen detection has been developed and evaluated. The test is based on the use of a MAb against VP72 protein of ASFV, the major viral capsid protein and highly immunogenic. First experiments using VP72 viral and recombinant protein or inactivated culture virus showed promising results with a sensitivity similar to that of a commercially available Antigen-ELISA. Moreover, these strips were tested with blood from experimentally infected pigs and field animals and the results compared with those of PCR and Antigen-ELISA. For the experimentally infected samples, there was an excellent correlation between the LFA and the ELISA, while the PCR always showed to be more sensitive (38 % positive samples by PCR versus 27 % by LFA). The LFA was demonstrated to be positive for animals with circulating virus levels exceeding 10(4) HAU. With the field samples, once again, the PCR detected more positives than either the Antigen-ELISA or LFA, although here the number of positive samples scored by the LFA exceeded the values obtained with the Antigen-ELISA, showing 60 % positivity vs 48 % for the ELISA. For the two groups of sera, the specificity was close to 100 % indicating that hardly any false positive samples were found. The newly developed LFA allows rapid and reliable detection of ASFV, at field and laboratory level, providing a new useful tool for control programs and in situations where laboratory support and skilled personnel are limited.

Cloning and analysis of the gene for a major surface antigen of Mycoplasma gallisepticum.

PubMed

Spencer, Denise L; Kurth, Kathy Toohey; Menon, Sreekumar A; VanDyk, Tina; Minion, F Chris

2002-01-01

Myplasma gallisepticum infects a wide variety of gallineaceous birds including chickens, turkeys, and pheasants. Infection occurs both horizontally and vertically. Thus, control of the spread of M. gallisepticum to noninfected flocks is difficult. Continual monitoring is necessary to identify infected flocks even under the most stringent infectious control practices. Monitoring, however, is usually performed by measuring hemagglutination activity (HA) in serum, an insensitive and variable test. Variability in the HA test arises differences in agglutination antigen, changes in antigenic profiles of the M. gallisepticum strain, and variability in reading the agglutination reaction. Enzyme-linked immunosorbent assays (ELISAs) are the preferred method of testing because of the ease in obtaining sera and the sensitivity and reproducibility of the assays, but the ELISA suffers from a lack of standardization in the test antigen. The ELISA test will be more easily accepted once the test antigen has been standardized. To this end, we have identified, cloned, and characterized the gene for an antigen that has potential as a species-specific antigen for M. gallisepticum The gene codes for a 75-kD protein, P75, that is recognized during natural infections. Recombinant P75 is not recognized in immunoblots by convalescent sera produced in chickens infected with Mycoplasma synoviae, Mycoplasma gallinarum, and Mycoplasma gallinaceum or in turkeys infected with Mycoplasma meleagridis.

[Detection of antigen structures in blood cells in various prepared plasma transfusions].

PubMed

Barz, D

1994-01-01

We investigated the content of antigen-bearing cells and cell fragments in Fresh Frozen Plasma (FFP) from blood centers, in Octaplas (virus-inactivated fresh plasma produced with the solvent/detergent technique by the Octapharma Company) and in MB-plasma (virus-inactivated fresh plasma after photodynamic treatment with methylen blue coming from the German Red Cross in Springe, Lower Saxony). With the aid of an immunoassay (MAIPA-test) these plasmas were tested regarding Rhesus-D-antigen, HLA-class-I- and HLA-class-II-antigens, platelet specific antigens HPA-1a/HPA-1b and granulocyte specific antigens NA1/NA2. In Octaplas (n = 10) we did not find cells or cell fragments and no antigen-bearing blood cell structures. In FFP (n = 28) there were platelet specific antigens in 27 cases (96.4%) and HLA-class-I-antigens in 4 cases (14.3%). In MB-plasma (n = 14) we found platelet specific antigens in all cases, HLA-class-I-antigens in 4 cases (18.6%), HLA-class-II-antigens and granulocyte specific antigens in 1 case (7.1%) and Rhesus-D-antigen in 3 cases (21.4%). Plasma derived from whole blood showed lower levels of cells and antigens than plasma which was produced with the aid of the cell separator.

Evaluation of Gastrothylax crumenifer antigenic preparation in serodiagnosis of paramphistomiasis in sheep.

PubMed

Ahmad, Tariq; Reshi, Mohammad Latif; Cheshti, M Z; Tanveer, Syed; Shah, Zaffar Amin; Fomada, Bashir Ahmad; Raina, O K

2014-04-01

An evaluation of Gastrothylax crumenifer crude antigen preparation viz., Somatic Antigen (SAg), Excretory Secretory Antigen (ESAg) and Egg Antigen (EAg) in serodiagnosis of disease was undertaken. Test sera samples were obtained from 30 Paramphistomiasis Positive and 30 Gastrothylax free sheep slaughtered at Hazratbal Kashmir. The referral antigenic preparation were evaluated against Paramphistomiasis positive sera, via., control negative sera, using double immunodiffusion test (DID), (IEP) Immunoelectrophoretic assay and ELISA. The performance of referral antigens, as assessed from percent sensitivity and specificity, revealed an increasing trend from DID (Double immunodiffusion-An immunological technique used in the detection, identification and quantification of antibodies and antigens) to IEP (immunoelectrophoresis-A general name for a number of biochemical methods for separation and characterization of proteins based on electrophoresis and reaction with antibodies), followed by ELISA, detecting higher number of sheep positive for paramphistomiasis. In ELISA the ESAg and SAg were evaluated as most reactive antigens with no significant difference and EAg was the least antigenic. In IEP, EAg had the higher sensitivity (60%) and analogous specificity of SAg and ESAg. The formation of the preceptin lines in the proximity to EAg containing wells (cathode end) in IEP was suggestive of higher molecular weight of G. crumenifer specific protein molecules with slower rate of migration. Purification and characterization of G. crumenifer and identification of specific antigenic molecules, particularly in EAg has been suggested for qualitative improvement of diagnostic value of the antigens in the tests used here in.

[Express diagnostics of bovine leucosis by immune sensor based on surface plasmon resonance].

PubMed

Pyrohova, L V; Starodub, M F; Artiukh, V P; Nahaieva, L I; Dobrosol, H I

2002-01-01

An immune sensor based on the surface plasmon resonance (SPR) was developed for express diagnostics of bovine leucosis. The sensor was used for detection of the level of antibodies against bovine leukaemia virus (BLV) in the blood serum. The industrially manufactured BLV antigen for screening test in the agar gel immunodiffusion (AGID) required the additional purification in order to be used in immune sensor analysis. It was shown that immune sensor analysis was more sensitive, rapid and simple in comparison with the traditional AGID test. It was stated that the developed immune sensor was capable to be used for performance of bovine leucosis screening at the farms and the minimal dilution of the serum should be 1:500.

Antigen-activated dendritic cells ameliorate influenza A infections

PubMed Central

Boonnak, Kobporn; Vogel, Leatrice; Orandle, Marlene; Zimmerman, Daniel; Talor, Eyal; Subbarao, Kanta

2013-01-01

Influenza A viruses cause significant morbidity and mortality worldwide. There is a need for alternative or adjunct therapies, as resistance to currently used antiviral drugs is emerging rapidly. We tested ligand epitope antigen presentation system (LEAPS) technology as a new immune-based treatment for influenza virus infection in a mouse model. Influenza-J-LEAPS peptides were synthesized by conjugating the binding ligand derived from the β2-microglobulin chain of the human MHC class I molecule (J-LEAPS) with 15 to 30 amino acid–long peptides derived from influenza virus NP, M, or HA proteins. DCs were stimulated with influenza-J-LEAPS peptides (influenza-J-LEAPS) and injected intravenously into infected mice. Antigen-specific LEAPS-stimulated DCs were effective in reducing influenza virus replication in the lungs and enhancing survival of infected animals. Additionally, they augmented influenza-specific T cell responses in the lungs and reduced the severity of disease by limiting excessive cytokine responses, which are known to contribute to morbidity and mortality following influenza virus infection. Our data demonstrate that influenza-J-LEAPS–pulsed DCs reduce virus replication in the lungs, enhance survival, and modulate the protective immune responses that eliminate the virus while preventing excessive cytokines that could injure the host. This approach shows promise as an adjunct to antiviral treatment of influenza virus infections. PMID:23934125

Toxoplasma gondii Recombinant Antigens as Tools for Serodiagnosis of Human Toxoplasmosis: Current Status of Studies

PubMed Central

2013-01-01

Toxoplasma gondii is a parasitic protozoan which is the cause of toxoplasmosis. Although human toxoplasmosis in healthy adults is usually asymptomatic, serious disease can occur in the case of congenital infections and immunocompromised individuals. Furthermore, despite the exact recognition of its etiology, it still presents a diagnostic problem. Diagnosis of toxoplasmosis is mainly based on the results of serological tests detecting anti-T. gondii-specific antibodies in the patient's serum sample. The specificities and sensitivities of serology tests depend mostly on the diagnostic antigen(s) used. Most of the commercial serological kits currently available are based on Toxoplasma lysate antigens (TLAs). In recent years, many studies showed that recombinant antigenic proteins of T. gondii may be an alternative source of antigens which are very useful for the serodiagnosis of toxoplasmosis. This article presents a review of current studies on the application and usefulness of different T. gondii recombinant antigens in serological tests for the diagnosis of human toxoplasmosis. PMID:23784855

Performance of a lateral flow immunochromatography test for the rapid diagnosis of active tuberculosis in a large multicentre study in areas with different clinical settings and tuberculosis exposure levels.

PubMed

Manga, Selene; Perales, Rocio; Reaño, Maria; D'Ambrosio, Lia; Migliori, Giovanni Battista; Amicosante, Massimo

2016-11-01

Tuberculosis (TB) continues to cause an outsized burden of morbidity and mortality worldwide, still missing efficient and largely accessible diagnostic tools determining an appropriate control of the disease. Serological tests have the potentially to impact TB diagnosis, in particular in extreme clinical settings. The diagnostic performances of the TB-XT HEMA EXPRESS (HEMA-EXPRESS) immunochromatographic rapid test for active TB diagnosis, based on use of multiple Mycobacterium tuberculosis (MTB) specific antigens, have been evaluated in a large study multicentre TB case-finding study, in populations with different exposure level to TB. A total of 1,386 subjects were enrolled in the six participating centres in Peru: 290 active-TB and 1,096 unaffected subjects. The TB prevalence (overall 20.5%) varied between 4.0% and 41.1% in the different study groups. Overall, the HEMA-EXPRESS test had 30.6% sensitivity (range 3.9-77.9%) and 84.6% specificity (range 51.6-97.3%). A significant inverse correlation between test accuracy (overall 73.5%, range 40.4-96.4%) and TB prevalence in the various study populations was observed (Pearson's r=-0.7985; P=0.05). HEMA-EXPRESS, is rapid and relatively inexpensive test suitable for routine use in TB diagnosis. In low TB prevalence conditions, test performance appears in line with WHO Target Product Profile for TB diagnostics. Performances appear suboptimal in high TB prevalence settings. Appropriate set-up in operative clinical settings has to be considered for novel serological tests for TB diagnosis, particularly for formats suitable for point-of-care use.

Development of a serotype colloidal gold strip using monoclonal antibody for rapid detection type Asia1 foot-and-mouth disease

PubMed Central

2011-01-01

Background In this study, we developed a rapid, one step colloid gold strip (CGS) capable of specifically detecting type Asia1 foot-and-mouth disease virus (FMDV). We have produced two monoclonal antibodies (mAb) to type Asia1 FMD (named 1B8 and 5E2). On the test strip, the purified 1B8 labelled with the colloidal gold was used as the detector, and the purified 5E2 and goat anti-mouse antibodies were wrapped onto nitrocellulose (NC) membranes as the test and the control line, respectively. The rapid colloidal gold stereotype diagnostic strip was housed in a plastic case. Results In specificity and sensitivity assay, there was no cross-reaction of the antigen with the other type of FMD and SVDV. The detection sensitivity was found to be as high as 10-5 dilution of Asia1/JSL/05 (1 × 107.2TCID50/50 μL). There was excellent agreement between the results obtained by CGS and reverse indirect hemagglutination assay (RIHA), and the agreement can reach to 98.75%. Conclusion We developed colloidal gold strips that have good qualities and does not require specialized equipment or technicians. This method provided a feasible, convenient, rapid, and effective for detecting type Asia1 FMDV in the fields. PMID:21880157

Development of a serotype colloidal gold strip using monoclonal antibody for rapid detection type Asia1 foot-and-mouth disease.

PubMed

Lin, Tong; Shao, Jun-jun; Du, Jun-zheng; Cong, Guo-zheng; Gao, Shan-dian; Chang, Huiyun

2011-09-01

In this study, we developed a rapid, one step colloid gold strip (CGS) capable of specifically detecting type Asia1 foot-and-mouth disease virus (FMDV). We have produced two monoclonal antibodies (mAb) to type Asia1 FMD (named 1B8 and 5E2). On the test strip, the purified 1B8 labelled with the colloidal gold was used as the detector, and the purified 5E2 and goat anti-mouse antibodies were wrapped onto nitrocellulose (NC) membranes as the test and the control line, respectively. The rapid colloidal gold stereotype diagnostic strip was housed in a plastic case. In specificity and sensitivity assay, there was no cross-reaction of the antigen with the other type of FMD and SVDV. The detection sensitivity was found to be as high as 10(-5) dilution of Asia1/JSL/05 (1 × 10(7.2)TCID(50)/50 μL). There was excellent agreement between the results obtained by CGS and reverse indirect hemagglutination assay (RIHA), and the agreement can reach to 98.75%. We developed colloidal gold strips that have good qualities and does not require specialized equipment or technicians. This method provided a feasible, convenient, rapid, and effective for detecting type Asia1 FMDV in the fields.

Application of a SERS-based lateral flow immunoassay strip for the rapid and sensitive detection of staphylococcal enterotoxin B

NASA Astrophysics Data System (ADS)

Hwang, Joonki; Lee, Sangyeop; Choo, Jaebum

2016-06-01

A novel surface-enhanced Raman scattering (SERS)-based lateral flow immunoassay (LFA) biosensor was developed to resolve problems associated with conventional LFA strips (e.g., limits in quantitative analysis and low sensitivity). In our SERS-based biosensor, Raman reporter-labeled hollow gold nanospheres (HGNs) were used as SERS detection probes instead of gold nanoparticles. With the proposed SERS-based LFA strip, the presence of a target antigen can be identified through a colour change in the test zone. Furthermore, highly sensitive quantitative evaluation is possible by measuring SERS signals from the test zone. To verify the feasibility of the SERS-based LFA strip platform, an immunoassay of staphylococcal enterotoxin B (SEB) was performed as a model reaction. The limit of detection (LOD) for SEB, as determined with the SERS-based LFA strip, was estimated to be 0.001 ng mL-1. This value is approximately three orders of magnitude more sensitive than that achieved with the corresponding ELISA-based method. The proposed SERS-based LFA strip sensor shows significant potential for the rapid and sensitive detection of target markers in a simplified manner.A novel surface-enhanced Raman scattering (SERS)-based lateral flow immunoassay (LFA) biosensor was developed to resolve problems associated with conventional LFA strips (e.g., limits in quantitative analysis and low sensitivity). In our SERS-based biosensor, Raman reporter-labeled hollow gold nanospheres (HGNs) were used as SERS detection probes instead of gold nanoparticles. With the proposed SERS-based LFA strip, the presence of a target antigen can be identified through a colour change in the test zone. Furthermore, highly sensitive quantitative evaluation is possible by measuring SERS signals from the test zone. To verify the feasibility of the SERS-based LFA strip platform, an immunoassay of staphylococcal enterotoxin B (SEB) was performed as a model reaction. The limit of detection (LOD) for SEB, as determined with the SERS-based LFA strip, was estimated to be 0.001 ng mL-1. This value is approximately three orders of magnitude more sensitive than that achieved with the corresponding ELISA-based method. The proposed SERS-based LFA strip sensor shows significant potential for the rapid and sensitive detection of target markers in a simplified manner. Electronic supplementary information (ESI) available. See DOI: 10.1039/c5nr07243c

Helminth antigens counteract a rapid high-fat diet-induced decrease in adipose tissue eosinophils.

PubMed

van den Berg, Susan M; van Dam, Andrea D; Kusters, Pascal J H; Beckers, Linda; den Toom, Myrthe; van der Velden, Saskia; Van den Bossche, Jan; van Die, Irma; Boon, Mariëtte R; Rensen, Patrick C N; Lutgens, Esther; de Winther, Menno P J

2017-10-01

Brown adipose tissue (BAT) activation and white adipose tissue (WAT) beiging can increase energy expenditure and have the potential to reduce obesity and associated diseases. The immune system is a potential target in mediating brown and beige adipocyte activation. Type 2 and anti-inflammatory immune cells contribute to metabolic homeostasis within lean WAT, with a prominent role for eosinophils and interleukin (IL)-4-induced anti-inflammatory macrophages. We determined eosinophil numbers in epididymal WAT (EpAT), subcutaneous WAT (ScAT) and BAT after 1 day, 3 days or 1 week of high-fat diet (HFD) feeding in C57Bl/6 mice. One day of HFD resulted in a rapid drop in eosinophil numbers in EpAT and BAT, and after 3 days, in ScAT. In an attempt to restore this HFD-induced drop in adipose tissue eosinophils, we treated 1-week HFD-fed mice with helminth antigens from Schistosoma mansoni or Trichuris suis and evaluated whether the well-known protective metabolic effects of helminth antigens involves BAT activation or beiging. Indeed, antigens of both helminth species induced high numbers of eosinophils in EpAT, but failed to induce beiging. In ScAT, Schistosoma mansoni antigens induced mild eosinophilia, which was accompanied by slightly more beiging. No effects were observed in BAT. To study type 2 responses on brown adipocytes directly, T37i cells were stimulated with IL-4. This increased Ucp1 expression and strongly induced the production of eosinophil chemoattractant CCL11 (+26-fold), revealing that brown adipocytes themselves can attract eosinophils. Our findings indicate that helminth antigen-induced eosinophilia fails to induce profound beiging of white adipocytes. © 2017 Society for Endocrinology.

Adjuvant-carrying synthetic vaccine particles augment the immune response to encapsulated antigen and exhibit strong local immune activation without inducing systemic cytokine release

PubMed Central

Ilyinskii, Petr O.; Roy, Christopher J.; O’Neil, Conlin P.; Browning, Erica A.; Pittet, Lynnelle A.; Altreuter, David H.; Alexis, Frank; Tonti, Elena; Shi, Jinjun; Basto, Pamela A.; Iannacone, Matteo; Radovic-Moreno, Aleksandar F.; Langer, Robert S.; Farokhzad, Omid C.; von Andrian, Ulrich H.; Johnston, Lloyd P.M.; Kishimoto, Takashi Kei

2014-01-01

Augmentation of immunogenicity can be achieved by particulate delivery of an antigen and by its co-administration with an adjuvant. However, many adjuvants initiate strong systemic inflammatory reactions in vivo, leading to potential adverse events and safety concerns. We have developed a synthetic vaccine particle (SVP) technology that enables co-encapsulation of antigen with potent adjuvants. We demonstrate that co-delivery of an antigen with a TLR7/8 or TLR9 agonist in synthetic polymer nanoparticles results in a strong augmentation of humoral and cellular immune responses with minimal systemic production of inflammatory cytokines. In contrast, antigen encapsulated into nanoparticles and admixed with free TLR7/8 agonist leads to lower immunogenicity and rapid induction of high levels of inflammatory cytokines in the serum (e.g., TNF-α and IL-6 levels are 50- to 200-fold higher upon injection of free resiquimod (R848) than of nanoparticle-encapsulated R848). Conversely, local immune stimulation as evidenced by cellular infiltration of draining lymph nodes and by intranodal cytokine production was more pronounced and persisted longer when SVP-encapsulated TLR agonists were used. The strong local immune activation achieved using a modular self-assembling nanoparticle platform markedly enhanced immunogenicity and was equally effective whether antigen and adjuvant were co-encapsulated in a single nanoparticle formulation or co-delivered in two separate nanoparticles. Moreover, particle encapsulation enabled the utilization of CpG oligonucleotides with the natural phosphodiester backbone, which are otherwise rapidly hydrolyzed by nucleases in vivo. The use of SVP may enable clinical use of potent TLR agonists as vaccine adjuvants for indications where cellular immunity or robust humoral responses are required. PMID:24593999

T Cell Receptor Engineering and Analysis Using the Yeast Display Platform

PubMed Central

Smith, Sheena N.; Harris, Daniel T.; Kranz, David M.

2017-01-01

The αβ heterodimeric T cell receptor (TCR) recognizes peptide antigens that are transported to the cell surface as a complex with a protein encoded by the major histocompatibility complex (MHC). T cells thus evolved a strategy to sense these intracellular antigens, and to respond either by eliminating the antigen-presenting cell (e.g. a virus-infected cell) or by secreting factors that recruit the immune system to the site of the antigen. The central role of the TCR in the binding of antigens as peptide-MHC (pepMHC) ligands has now been studied thoroughly. Interestingly, despite their exquisite sensitivity (e.g. T cell activation by as few as 1 to 3 pepMHC complexes on a single target cell), TCRs are known to have relatively low affinities for pepMHC, with KD values in the micromolar range. There has been interest in engineering the affinity of TCRs in order to use this class of molecules in ways similar to now done with antibodies. By doing so, it would be possible to harness the potential of TCRs as therapeutics against a much wider array of antigens that include essentially all intracellular targets. To engineer TCRs, and to analyze their binding features more rapidly, we have used a yeast display system as a platform. Expression and engineering of a single-chain form of the TCR, analogous to scFv fragments from antibodies, allow the TCR to be affinity matured with a variety of possible pepMHC ligands. In addition, the yeast display platform allows one to rapidly generate TCR variants with diverse binding affinities and to analyze specificity and affinity without the need for purification of soluble forms of the TCRs. The present chapter describes the methods for engineering and analyzing single-chain TCRs using yeast display. PMID:26060072

Evaluation of a rapid and inexpensive dipstick assay for the diagnosis of Plasmodium falciparum malaria.

PubMed Central

Mills, C. D.; Burgess, D. C.; Taylor, H. J.; Kain, K. C.

1999-01-01

Rapid, accurate and affordable methods are needed for the diagnosis of malaria. Reported here is an evaluation of a new immunochromatographic strip, the PATH Falciparum Malaria IC Strip, which is impregnated with an immobilized IgM monoclonal antibody that binds to the HRP-II antigen of Plasmodium falciparum. In contrast to other commercially available kits marketed for the rapid diagnosis of falciparum malaria, this kit should be affordable in the malaria-endemic world. Using microscopy and polymerase chain reaction (PCR)-based methods as reference standards, we compared two versions of the PATH test for the detection of P. falciparum infection in 200 febrile travellers. As determined by PCR and microscopy, 148 travellers had malaria, 50 of whom (33.8%) were infected with P. falciparum. Compared with PCR, the two versions of the PATH test had initial sensitivities of 90% and 88% and specificities of 97% and 96%, respectively, for the detection of falciparum malaria. When discrepant samples were retested blindly with a modified procedure (increased sample volume and longer washing step) the sensitivity and specificity of both kits improved to 96% and 99%, respectively. The two remaining false negatives occurred in samples with < 100 parasites per microliter of blood. The accuracy, simplicity and predicted low cost may make this test a useful diagnostic tool in malaria-endemic areas. PMID:10444878

Interlaboratory discrepancy of antigenuria results in 2 patients with AIDS and histoplasmosis.

PubMed

McKinsey, David S; McKinsey, Joel P; Northcutt, Noelle; Sarria, Juan C

2009-01-01

Histoplasma polysaccharide antigen testing is used routinely to diagnose histoplasmosis. At least 3 antigen tests are commercially available. Controversy exists about the relative accuracy of these tests. We report 2 patients with AIDS and culture-confirmed Histoplasma capsulatum meningitis from whom discrepant Histoplasma polysaccharide antigen results were obtained from different laboratories and discuss the potential clinical implications of these results.

Effect of pronase on high-incidence blood group antigens and the prevalence of antibodies to pronase-treated erythrocytes.

PubMed

Reid, M E; Greeen, C A; Hoffer, J; Øyen, R

1996-01-01

Pronase is a useful and relatively nonspecific protease that cleaves many red blood cell (RBC) membrane proteins that carry blood group antigens. Unexpected findings in tests using pronase-treated RBCs during the investigation of a patient's blood sample led us to test which high-incidence blood group antigens were sensitive and which were resistant to pronase treatment, and to determine the prevalence of antipronase in the serum of blood donors. Our results show that antigens in the Cromer and Lutheran blood group systems and the JMH antigen were sensitive to pronase treatment of RBCs. Antigens in the Dombrock blood group system and Sc1 were either sensitive to or markedly weakened by pronase treatment of RBCs. The following high-incidence antigens were resistant to treatment of RBCs with pronase: AnWj, Ata, Coa, Co3, Dib, EnaFR, Era, Fy3, Jk3, Jra, k, Kpb, Jsb, K14, Lan, Oka, Rh17, U, Vel, and Wrb. Over half of the serum samples from normal blood donors contained antibodies to pronase-treated RBCs. When testing human serum against pronase-treated RBCs, it is essential either to use an autocontrol or to perform the testing with an eluate.

Production and characterization of monoclonal antibodies specific to pangasius catfish, basa, and tra.

PubMed

Gajewski, K G; Chen, Y-T; Hsieh, Y-H P

2009-04-01

Four IgG (subclass IgG1) class monoclonal antibodies (MAbs) strongly reactive to Asian farm-raised Pangasius catfish, tra (Pangasius hypophthalmus) and basa (Pangasius bocourti), have been developed. These MAbs were raised by immunizing an animal with thermal-stable crude sarcoplasmic protein extract of cooked tra. The MAbs were selected by screening hybridoma clones against more than 70 common fish and meat protein extracts. Two MAbs, T7E10 and T1G11, were found to be specific to the Asian Pangasius catfish, tra, and basa, with no cross-reactions with any of the common fish and meat species or with the food additive proteins (bovine serum albumin, soy proteins, milk proteins, egg proteins, and gelatin) tested. MAb T7E10 recognized 2 antigenic proteins (molecular weight approximately 36 and 75 kDa) in raw and cooked tra and basa extracts, while T1G11 bound to several proteins (molecular weight between 13 and 18 kDa) in tra and basa extracts. Two other MAbs, F7B8 and F1G11, recognized a common protein (36 KDa) and cross-reacted with all the fish extracts tested and with several mammalian species. These MAbs can be employed individually or in combination in various formats of immunoassays for rapid identification of Pangasius catfish, either raw or cooked. They can also be used to study the biological, biochemical, and physiological aspects of thermal-stable antigenic proteins. This is the first study identifying these thermal-stable antigenic proteins present in Pangasius catfish as species-specific biomarkers.

Detection of Entamoeba histolytica by Recombinase Polymerase Amplification.

PubMed

Nair, Gayatri; Rebolledo, Mauricio; White, A Clinton; Crannell, Zachary; Richards-Kortum, R Rebecca; Pinilla, A Elizabeth; Ramírez, Juan David; López, M Consuelo; Castellanos-Gonzalez, Alejandro

2015-09-01

Amebiasis is an important cause of diarrheal disease worldwide and has been associated with childhood malnutrition. Traditional microscopy approaches are neither sensitive nor specific for Entamoeba histolytica. Antigen assays are more specific, but many cases are missed unless tested by molecular methods. Although polymerase chain reaction (PCR) is effective, the need for sophisticated, expensive equipment, infrastructure, and trained personnel limits its usefulness, especially in the resource-limited, endemic areas. Here, we report development of a recombinase polymerase amplification (RPA) method to detect E. histolytica specifically. Using visual detection by lateral flow (LF), the test was highly sensitive and specific and could be performed without additional equipment. The availability of this inexpensive, sensitive, and field-applicable diagnostic test could facilitate rapid diagnosis and treatment of amebiasis in endemic regions. © The American Society of Tropical Medicine and Hygiene.

Probing the Energetics of Antigen-Antibody Recognition by Titration Microcalorimetry

PubMed

Jelesarov; Leder; Bosshard

1996-06-01

Our understanding of the energetics that govern antigen-antibody recognition lags behind the increasingly rapid accumulation of structural information on antigen-antibody complexes. Thanks to the development of highly sensitive microcalorimeters, the thermodynamic parameters of antigen-antibody interactions can now be measured with precision and using only nanomole quantities of protein. The method of choice is isothermal titration calorimetry, in which a solution of the antibody (or antigen) is titrated with small aliquots of the antigen (or antibody) and the heat change accompanying the formation of the antigen-antibody complex is measured with a sensitivity as high as 0.1 μcal s-1. The free energy of binding (DeltaG), the binding enthalpy (DeltaH), and the binding entropy (DeltaS) are usually obtained from a single experiment, and no spectroscopic or radioactive label must be introduced into the antigen or antibody. The often large and negative change in heat capacity (DeltaCp) accompanying the formation of an antigen-antibody complex is obtained from DeltaH measured at different temperatures. The basic theory and the principle of the measurements are reviewed and illustrated by examples. The thermodynamic parameters relate to the dynamic physical forces that govern the association of the freely moving antigen and antibody into a well-structured and unique complex. This information complements the static picture of the antigen-antibody complex that results from X-ray diffraction analysis. Attempts to correlate dynamic and static aspects are discussed briefly.

Canine parvovirus type 2c identified from an outbreak of severe gastroenteritis in a litter in Sweden

PubMed Central

2013-01-01

A litter of recently-vaccinated puppies in Sweden experienced signs of severe haemorrhagic gastroenteritis. Canine parvovirus (CPV) was suspected as the cause of this outbreak on the basis of the clinical signs and the presence of parvoviral antigen in the faeces from one of the affected pups - confirmed using a commercial in-clinic faecal antigen ELISA test kit. A concern was raised about whether the vaccine (which contained a live, attenuated strain of CPV) could have caused the disease and so further faecal samples from the affected pups were submitted for laboratory virus isolation and identification. On cell culture, two out of four faecal samples were found to be virus-positive. This was confirmed as being canine parvovirus by immuno-staining with CPV specific monoclonal antibody. The virus was then tested using a series of PCR probes designed to confirm the identity of CPV and to distinguish the unique vaccine strain from field virus. This confirmed that the virus was indeed CPV but that it was not vaccine strain. The virus was then typed by sequencing the 426 amino acid region of the capsid gene which revealed this to be a type 2c virus. Since its emergence in the late 1970s, canine parvovirus 2 (CPV2) has spread worldwide and is recognised as an important canine pathogen in all countries. The original CPV2 rapidly evolved into two antigenic variants, CPV2a and CPV2b, which progressively replaced the original CPV2. More recently a new antigenic variant, CPV2c, has appeared. To date this variant has been identified in many countries worldwide but there have been no reports yet of its presence in any Scandinavian countries. This case report therefore represents the first published evidence of the involvement of CPV2c in a severe outbreak of typical haemorrhagic gastroenteritis in a susceptible litter of pups in Scandinavia. PMID:24016358

Canine parvovirus type 2c identified from an outbreak of severe gastroenteritis in a litter in Sweden.

PubMed

Sutton, David; Vinberg, Carina; Gustafsson, Agneta; Pearce, Jacqueline; Greenwood, Neil

2013-09-10

A litter of recently-vaccinated puppies in Sweden experienced signs of severe haemorrhagic gastroenteritis. Canine parvovirus (CPV) was suspected as the cause of this outbreak on the basis of the clinical signs and the presence of parvoviral antigen in the faeces from one of the affected pups - confirmed using a commercial in-clinic faecal antigen ELISA test kit. A concern was raised about whether the vaccine (which contained a live, attenuated strain of CPV) could have caused the disease and so further faecal samples from the affected pups were submitted for laboratory virus isolation and identification.On cell culture, two out of four faecal samples were found to be virus-positive. This was confirmed as being canine parvovirus by immuno-staining with CPV specific monoclonal antibody. The virus was then tested using a series of PCR probes designed to confirm the identity of CPV and to distinguish the unique vaccine strain from field virus. This confirmed that the virus was indeed CPV but that it was not vaccine strain. The virus was then typed by sequencing the 426 amino acid region of the capsid gene which revealed this to be a type 2c virus.Since its emergence in the late 1970s, canine parvovirus 2 (CPV2) has spread worldwide and is recognised as an important canine pathogen in all countries. The original CPV2 rapidly evolved into two antigenic variants, CPV2a and CPV2b, which progressively replaced the original CPV2. More recently a new antigenic variant, CPV2c, has appeared. To date this variant has been identified in many countries worldwide but there have been no reports yet of its presence in any Scandinavian countries. This case report therefore represents the first published evidence of the involvement of CPV2c in a severe outbreak of typical haemorrhagic gastroenteritis in a susceptible litter of pups in Scandinavia.

Soluble Antigen Fluorescent-Antibody Technique

PubMed Central

Toussaint, Andre J.; Anderson, Robert I.

1965-01-01

An indirect fluorescent-antibody (FA) procedure employing soluble antigen fixed onto an artificial matrix was developed, and a mechanical means for reading of test results was devised. The method employs two small cellulose acetate paper discs for each test. One disc contains soluble antigen diluted in 1% bovine serum albumin (BSA); the other contains only 1% BSA and serves as a control. After testing by the indirect FA procedure, the results of the tests are read on a fluorometer fitted with a paper chromatogram door. The instrument is set at zero with the control disc as a blank, and the specific fluorescence of the antigen disc is determined. Findings obtained with homologous and heterologous antisera indicated that the method yields excellent results. The soluble antigen fluorescent-antibody technique has definite advantages over the conventional indirect FA procedures. (i) The investigator may objectively select the antigen to be employed. (ii) It is possible to obtain objective mechanical reading of test results rather than the highly subjective readings required by conventional methods. (iii) The system compensates for any nonspecific fluorescence contributed either by the serum (e.g., drugs) or by free fluorescein in the conjugated antiserum. Images Fig. 1 PMID:14339261

Antibody responses in New World camelids with tuberculosis caused by Mycobacterium microti.

PubMed

Lyashchenko, K P; Greenwald, R; Esfandiari, J; Meylan, M; Burri, I Hengrave; Zanolari, P

2007-12-15

Antibody responses in New World camelids (NWC) infected with Mycobacterium microti were studied by two serological methods, multiantigen print immunoassay (MAPIA) and lateral-flow-based rapid test (RT). Serum samples were collected during 2004-2006 from 87 animals including 1 alpaca and 7 llamas with confirmed or suspected M. microti infection, 33 potentially exposed but clinically healthy animals from known infected herds, and 46 control NWC from herds where infection had not been previously diagnosed. The serological assays correctly identified infection status in 97% (MAPIA) or 87% (RT) cases. In three llamas with confirmed M. microti infection and one llama with gross pathology suggestive of disease, for which multiple serum samples collected over time were available, the antibody-based tests showed positive results 1-2 years prior to the onset of clinical signs or being found dead. In MAPIA, MPB83 protein was identified to be an immunodominant serological target antigen recognized in NWC infected with M. microti. With the limited number of animals tested in this study, the serological assays demonstrated the potential for convenient, rapid, and accurate diagnosis of M. microti infection in live llamas and alpacas.

Development of a rapid diagnostic test for pertussis: direct detection of pertussis toxin in respiratory secretions.

PubMed Central

Friedman, R L; Paulaitis, S; McMillan, J W

1989-01-01

Monoclonal antibodies (MAb) were produced against the specific Bordetella pertussis antigen pertussis toxin (PT). In preliminary studies, one MAb (IB12) was selected and used in an enzyme-linked dot blot immunoassay to evaluate the ability of the method to detect known amounts of PT in control experiments and to test its potential for direct detection of PT in nasopharyngeal secretion (NP) specimens from patients with confirmed cases of whooping cough. The dot blot assay was able to detect PT at levels as low as 10 ng per dot in either buffer or control NP specimens. The assay demonstrated specificity, reacting only with dot blots of whole B. pertussis and not Bordetella bronchiseptica, Bordetella parapertussis, or other bacterial strains. In preliminary studies, NP aspirate, swab, and wash specimens were compared. The specimen of choice was found to be the NP aspirate, for which 100% positive results were found in the assay. These initial studies suggest that the dot blot immunoassay in which a MAb is used for direct detection of PT in NP specimens may be useful as a rapid diagnostic test for pertussis. Images PMID:2808670

Multi-centre evaluation of the Determine HIV Combo assay when used for point of care testing in a high risk clinic-based population.

PubMed

Conway, Damian P; Holt, Martin; McNulty, Anna; Couldwell, Deborah L; Smith, Don E; Davies, Stephen C; Cunningham, Philip; Keen, Phillip; Guy, Rebecca

2014-01-01

Determine HIV Combo (DHC) is the first point of care assay designed to increase sensitivity in early infection by detecting both HIV antibody and antigen. We conducted a large multi-centre evaluation of DHC performance in Sydney sexual health clinics. We compared DHC performance (overall, by test component and in early infection) with conventional laboratory HIV serology (fourth generation screening immunoassay, supplementary HIV antibody, p24 antigen and Western blot tests) when testing gay and bisexual men attending four clinic sites. Early infection was defined as either acute or recent HIV infection acquired within the last six months. Of 3,190 evaluation specimens, 39 were confirmed as HIV-positive (12 with early infection) and 3,133 were HIV-negative by reference testing. DHC sensitivity was 87.2% overall and 94.4% and 0% for the antibody and antigen components, respectively. Sensitivity in early infection was 66.7% (all DHC antibody reactive) and the DHC antigen component detected none of nine HIV p24 antigen positive specimens. Median HIV RNA was higher in false negative than true positive cases (238,025 vs. 37,591 copies/ml; p = 0.022). Specificity overall was 99.4% with the antigen component contributing to 33% of false positives. The DHC antibody component detected two thirds of those with early infection, while the DHC antigen component did not enhance performance during point of care HIV testing in a high risk clinic-based population.

Survey of Spanish general practitioners' attitudes toward management of sore throat: an internet-based questionnaire study.

PubMed

Llor, Carl; Vilaseca, Isabel; Lehrer-Coriat, Eduardo; Boleda, Xavier; Cañada, José L; Moragas, Ana; Cots, Josep M

2017-02-14

The management of sore throat varies widely in Europe. The objective of this study was to gain insight into clinicians' perceptions on the current management of sore throat in Spain. Cross-sectional, internet-based questionnaire study answered from July to September 2013. General practitioners (GPs) affiliated with the two largest scientific societies of primary care were invited to participate in the study. Questions were asked about physician knowledge, the use of current national guidelines for sore throat management, and management in two clinical scenarios, depicting a young adult with sore throat and: 1. cough, coriza with or without fever, and 2. fever without cough and coriza. The questionnaire was completed by 1476 GPs (5%) and 12.7% declared using rapid antigen detection tests. Antibiotics were considered by 18.8% of the GPs in the first scenario and by 32% in the second scenario (p < 0.001). The antibiotics most commonly mentioned by GPs were amoxicillin and amoxicillin + clavulanate (52.7 and 31.2%, respectively) whereas penicillin V was only prescribed in 11.9% of the cases. The drugs most commonly considered in both scenarios were analgesics and anti-inflammatory drugs. Antitussives, decongestants and expectorants were more commonly prescribed in cases of suspected viral infection (p < 0.001). GPs have misconceptions as to the indications for using rapid antigen detection tests and prescribing drugs in the management of sore throat. These results suggest that guidelines are seldom followed since one in five GPs declared giving antibiotics for patients with a suspected viral infection and the use of second-choice antibiotics seems considerable.

Development of improved enzyme-based and lateral flow immunoassays for rapid and accurate serodiagnosis of canine brucellosis.

PubMed

Cortina, María E; Novak, Analía; Melli, Luciano J; Elena, Sebastián; Corbera, Natalia; Romero, Juan E; Nicola, Ana M; Ugalde, Juan E; Comerci, Diego J; Ciocchini, Andrés E

2017-09-01

Brucellosis is a widespread zoonotic disease caused by Brucella spp. Brucella canis is the etiological agent of canine brucellosis, a disease that can lead to sterility in bitches and dogs causing important economic losses in breeding kennels. Early and accurate diagnosis of canine brucellosis is central to control the disease and lower the risk of transmission to humans. Here, we develop and validate enzyme and lateral flow immunoassays for improved serodiagnosis of canine brucellosis using as antigen the B. canis rough lipopolysaccharide (rLPS). The method used to obtain the rLPS allowed us to produce more homogeneous batches of the antigen that facilitated the standardization of the assays. To validate the assays, 284 serum samples obtained from naturally infected dogs and healthy animals were analyzed. For the B. canis-iELISA and B. canis-LFIA the diagnostic sensitivity was of 98.6%, and the specificity 99.5% and 100%, respectively. We propose the implementation of the B. canis-LFIA as a screening test in combination with the highly accurate laboratory g-iELISA. The B. canis-LFIA is a rapid, accurate and easy to use test, characteristics that make it ideal for the serological surveillance of canine brucellosis in the field or veterinary laboratories. Finally, a blind study including 1040 serum samples obtained from urban dogs showed a prevalence higher than 5% highlighting the need of new diagnostic tools for a more effective control of the disease in dogs and therefore to reduce the risk of transmission of this zoonotic pathogen to humans. Copyright © 2017 Elsevier B.V. All rights reserved.

A Novel Heat Shock Protein 70-based Vaccine Prepared from DC-Tumor Fusion Cells.

PubMed

Weng, Desheng; Calderwood, Stuart K; Gong, Jianlin

2018-01-01

We have developed an enhanced molecular chaperone-based vaccine through rapid isolation of Hsp70 peptide complexes after the fusion of tumor and dendritic cells (Hsp70.PC-F). In this approach, the tumor antigens are introduced into the antigen processing machinery of dendritic cells through the cell fusion process and thus we can obtain antigenic tumor peptides or their intermediates that have been processed by dendritic cells. Our results show that Hsp70.PC-F has increased immunogenicity compared to preparations from tumor cells alone and therefore constitutes an improved formulation of chaperone protein-based tumor vaccine.

Multiplex serology for impact evaluation of bed net distribution on burden of lymphatic filariasis and four species of human malaria in northern Mozambique

PubMed Central

Candrinho, Baltazar; Chambe, Geraldo; Muchanga, João; Muguande, Olinda; Matsinhe, Graça; Mathe, Guidion; Rogier, Eric; Doyle, Timothy; Zulliger, Rose; Colborn, James; Saifodine, Abu; Lammie, Patrick; Priest, Jeffrey W.

2018-01-01

Background Universal coverage with long-lasting insecticidal nets (LLINs) is a primary control strategy against Plasmodium falciparum malaria. However, its impact on the three other main species of human malaria and lymphatic filariasis (LF), which share the same vectors in many co-endemic areas, is not as well characterized. The recent development of multiplex antibody detection provides the opportunity for simultaneous evaluation of the impact of control measures on the burden of multiple diseases. Methodology/Principal findings Two cross-sectional household surveys at baseline and one year after a LLIN distribution campaign were implemented in Mecubúri and Nacala-a-Velha Districts in Nampula Province, Mozambique. Both districts were known to be endemic for LF; both received mass drug administration (MDA) with antifilarial drugs during the evaluation period. Access to and use of LLINs was recorded, and household members were tested with P. falciparum rapid diagnostic tests (RDTs). Dried blood spots were collected and analyzed for presence of antibodies to three P. falciparum antigens, P. vivax MSP-119, P. ovale MSP-119, P. malariae MSP-119, and three LF antigens. Seroconversion rates were calculated and the association between LLIN use and post-campaign seropositivity was estimated using multivariate regression. The campaign covered 68% (95% CI: 58–77) of the population in Nacala-a-Velha and 46% (37–56) in Mecubúri. There was no statistically significant change in P. falciparum RDT positivity between the two surveys. Population seropositivity at baseline ranged from 31–81% for the P. falciparum antigens, 3–4% for P. vivax MSP-119, 41–43% for P. ovale MSP-119, 46–56% for P. malariae MSP-119, and 37–76% for the LF antigens. The seroconversion rate to the LF Bm33 antigen decreased significantly in both districts. The seroconversion rate to P. malariae MSP-119 and the LF Wb123 and Bm14 antigens each decreased significantly in one of the two districts. Community LLIN use was associated with a decreased risk of P. falciparum RDT positivity, P. falciparum LSA-1 seropositivity, and P. malariae MSP-119 seropositivity, but not LF antigen seropositivity. Conclusions/Significance The study area noted significant declines in LF seropositivity, but these were not associated with LLIN use. The MDA could have masked any impact of the LLINs on population LF seropositivity. The LLIN campaign did not reach adequately high coverage to decrease P. falciparum RDT positivity, the most common measure of P. falciparum burden. However, the significant decreases in the seroconversion rate to the P. malariae antigen, coupled with an association between community LLIN use and individual-level decreases in seropositivity to P. falciparum and P. malariae antigens show evidence of impact of the LLIN campaign and highlight the utility of using multiantigenic serological approaches for measuring intervention impact. PMID:29444078

Multiplex serology for impact evaluation of bed net distribution on burden of lymphatic filariasis and four species of human malaria in northern Mozambique.

PubMed

Plucinski, Mateusz M; Candrinho, Baltazar; Chambe, Geraldo; Muchanga, João; Muguande, Olinda; Matsinhe, Graça; Mathe, Guidion; Rogier, Eric; Doyle, Timothy; Zulliger, Rose; Colborn, James; Saifodine, Abu; Lammie, Patrick; Priest, Jeffrey W

2018-02-01

Universal coverage with long-lasting insecticidal nets (LLINs) is a primary control strategy against Plasmodium falciparum malaria. However, its impact on the three other main species of human malaria and lymphatic filariasis (LF), which share the same vectors in many co-endemic areas, is not as well characterized. The recent development of multiplex antibody detection provides the opportunity for simultaneous evaluation of the impact of control measures on the burden of multiple diseases. Two cross-sectional household surveys at baseline and one year after a LLIN distribution campaign were implemented in Mecubúri and Nacala-a-Velha Districts in Nampula Province, Mozambique. Both districts were known to be endemic for LF; both received mass drug administration (MDA) with antifilarial drugs during the evaluation period. Access to and use of LLINs was recorded, and household members were tested with P. falciparum rapid diagnostic tests (RDTs). Dried blood spots were collected and analyzed for presence of antibodies to three P. falciparum antigens, P. vivax MSP-119, P. ovale MSP-119, P. malariae MSP-119, and three LF antigens. Seroconversion rates were calculated and the association between LLIN use and post-campaign seropositivity was estimated using multivariate regression. The campaign covered 68% (95% CI: 58-77) of the population in Nacala-a-Velha and 46% (37-56) in Mecubúri. There was no statistically significant change in P. falciparum RDT positivity between the two surveys. Population seropositivity at baseline ranged from 31-81% for the P. falciparum antigens, 3-4% for P. vivax MSP-119, 41-43% for P. ovale MSP-119, 46-56% for P. malariae MSP-119, and 37-76% for the LF antigens. The seroconversion rate to the LF Bm33 antigen decreased significantly in both districts. The seroconversion rate to P. malariae MSP-119 and the LF Wb123 and Bm14 antigens each decreased significantly in one of the two districts. Community LLIN use was associated with a decreased risk of P. falciparum RDT positivity, P. falciparum LSA-1 seropositivity, and P. malariae MSP-119 seropositivity, but not LF antigen seropositivity. The study area noted significant declines in LF seropositivity, but these were not associated with LLIN use. The MDA could have masked any impact of the LLINs on population LF seropositivity. The LLIN campaign did not reach adequately high coverage to decrease P. falciparum RDT positivity, the most common measure of P. falciparum burden. However, the significant decreases in the seroconversion rate to the P. malariae antigen, coupled with an association between community LLIN use and individual-level decreases in seropositivity to P. falciparum and P. malariae antigens show evidence of impact of the LLIN campaign and highlight the utility of using multiantigenic serological approaches for measuring intervention impact.

Histoplasma Urinary Antigen Testing Obviates the Need for Coincident Serum Antigen Testing.

PubMed

Libert, Diane; Procop, Gary W; Ansari, Mohammad Q

2018-03-07

Serum and urine antigen (SAg, UAg) detection are common tests for Histoplasma capsulatum. UAg detection is more widely used and reportedly has a higher sensitivity. We investigated whether SAg detection contributes meaningfully to the initial evaluation of patients with suspected histoplasmosis. We reviewed 20,285 UAg and 1,426 SAg tests ordered from 1997 to 2016 and analyzed paired UAg and SAg tests completed on the same patient within 1 week. We determined the positivity rate for each test. Of 601 paired specimens, 542 were concurrent negatives and 48 were concurrent positives (98% agreement). Medical records were available for eight of 11 pairs with discrepant results. UAg was falsely positive in six instances, truly positive once, and falsely negative once. These findings support using a single antigen detection test, rather than both UAg and SAg, as an initial screen for suspected histoplasmosis. This aligns with the current practice of most physicians.

Standardization of a latex agglutination test for coproantigen detection of Fasciola sp. in bovine cattle stool.

PubMed

Orejarena Ávila, Lina Paola; Inguilan Benavides, Erika Marcela; Padilla Sanabria, Leonardo; Recalde-Reyes, Delia Piedad; Rodríguez-Salazar, Carlos Andrés; Castaño-Osorio, Jhon Carlos

2018-03-01

Fasciolosis is a zoonotic parasitic disease, which affects humans and animals; diagnosed through noncommercial immunoassay tests that cannot be used on the field. Thereby, establishing the optimal conditions to develop a latex agglutination technique with IgG and IgM antibodies directed against excretion/secretion antigens of Fasciola sp. is a priority. Latex particles were sensitized with IgG and IgM antibodies directed against excretion/secretion antigens of Fasciola sp. The specificity of the antibodies was determined against antigens of different helminths and protozoa; the sensitivity and specificity of the test was evaluated against a previously standardized direct ELISA. The coupling rates of the IgG and IgM antibodies were 85.77 and 100%, respectively. The minimum detectable concentration of Fasciola sp. excretion/secretion antigens, diluted in a phosphate-buffered saline, was 1.589 mg/mL(IgG) and 0.158 mg/mL(IgM) and for the antigens incorporated in the bovine cattle stool it was 3.178 mg/mL(IgG) and 1.589 mg/mL(IgM). The test showed crossed reaction against Giardia sp., and Cryptosporidium sp. antigens. Agreement of the IgG and IgM latex test against the ELISA test was of 78.78 and 96.96%, respectively; the specificity found was of 100% for both tests and sensitivity was 78.79% (IgG) and 96.97% (IgM). This work standardized the latex agglutination technique to detect Fasciola sp. antigens in bovine cattle stool.

Detection of peste des petits ruminants virus antigen using immunofiltration and antigen-competition ELISA methods.

PubMed

Raj, G Dhinakar; Rajanathan, T M C; Kumar, C Senthil; Ramathilagam, G; Hiremath, Geetha; Shaila, M S

2008-06-22

Peste des petits ruminants (PPR) is one of the most economically important diseases affecting sheep and goats in India. An immunofiltration-based test has been developed using either mono-specific serum/monoclonal antibodies (mAb) prepared against a recombinant truncated nucleocapsid protein of rinderpest virus (RPV) cross-reactive with PPR virus. This method consists of coating ocular swab eluate from suspected animals onto a nitrocellulose membrane housed in a plastic module, which is allowed to react with suitable dilutions of a mAb or a mono-specific polyclonal antibody. The antigen-antibody complex formed on the membrane is then detected by protein A-colloidal gold conjugate, which forms a pink colour. In the immunofiltration test, concordant results were obtained using either PPRV mAb or mono-specific serum. Another test, an antigen-competition ELISA which relies on the competition between plate-coated recombinant truncated 'N' protein of RPV and the PPRV 'N' protein present in ocular swab eluates (sample) for binding to the mono-specific antibody against N protein of RPV (in liquid phase) was developed. The cut-off value for this test was established using reverse transcription polymerase chain reaction (RT-PCR) positive and negative oculo-nasal swab samples. Linear correlation between percent inhibition (PI) values in antigen-competition ELISA and virus infectivity titres was 0.992. Comparison of the immunofiltration test with the antigen-competition ELISA yielded a sensitivity of 80% and specificity of 100%. These two tests can serve as a screening (immunofiltration) and confirmatory (antigen-competition ELISA) test, respectively, in the diagnosis of PPR in sheep or goats.

African Americans' Perceptions of Prostate-Specific Antigen Prostate Cancer Screening

ERIC Educational Resources Information Center

Hunter, Jaimie C.; Vines, Anissa I.; Carlisle, Veronica

2015-01-01

Background: In 2012, the U.S. Preventive Services Task Force released a hotly debated recommendation against prostate-specific antigen testing for all men. The present research examines African Americans' beliefs about their susceptibility to prostate cancer (PCa) and the effectiveness of prostate-specific antigen testing in the context of the…

A High-Performance Multiplex Immunoassay for Serodiagnosis of Flavivirus-Associated Neurological Diseases in Horses

PubMed Central

Beck, Cécile; Desprès, Philippe; Paulous, Sylvie; Vanhomwegen, Jessica; Lowenski, Steeve; Nowotny, Norbert; Durand, Benoit; Garnier, Annabelle; Blaise-Boisseau, Sandra; Guitton, Edouard; Yamanaka, Takashi; Zientara, Stéphan; Lecollinet, Sylvie

2015-01-01

West Nile virus (WNV), Japanese encephalitis virus (JEV), and tick-borne encephalitis virus (TBEV) are flaviviruses responsible for severe neuroinvasive infections in humans and horses. The confirmation of flavivirus infections is mostly based on rapid serological tests such as enzyme-linked immunosorbent assays (ELISAs). These tests suffer from poor specificity, mainly due to antigenic cross-reactivity among flavivirus members. Robust diagnosis therefore needs to be validated through virus neutralisation tests (VNTs) which are time-consuming and require BSL3 facilities. The flavivirus envelope (E) glycoprotein ectodomain is composed of three domains (D) named DI, DII, and DIII, with EDIII containing virus-specific epitopes. In order to improve the serological differentiation of flavivirus infections, the recombinant soluble ectodomain of WNV E (WNV.sE) and EDIIIs (rEDIIIs) of WNV, JEV, and TBEV were synthesised using the Drosophila S2 expression system. Purified antigens were covalently bonded to fluorescent beads. The microspheres coupled to WNV.sE or rEDIIIs were assayed with about 300 equine immune sera from natural and experimental flavivirus infections and 172 nonimmune equine sera as negative controls. rEDIII-coupled microspheres captured specific antibodies against WNV, TBEV, or JEV in positive horse sera. This innovative multiplex immunoassay is a powerful alternative to ELISAs and VNTs for veterinary diagnosis of flavivirus-related diseases. PMID:26457301

Report of data on children with non-typhi Salmonella gastroenteritis in a three-year period.

PubMed

Küçük, Öznur; Biçer, Suat; Ugraş, Meltem; Çöl, Defne; Giray, Tuba; Çiler Erdag, Gülay; Gürol, Yeşim; Yilmaz, Gülden; Yalvaç, Zerrin; Vitrinel, Ayça; Kaspar, Çigdem

2016-09-01

The purpose of this study was to evaluate the clinical and laboratory data of children with acute gastroenteritis caused by non-typhoid Salmonella spp. infections. Clinical (demographic data, symptoms and findings) and laboratory data (stool microscopy, rapid antigen tests, culture, multiplex polymerase chain reaction and blood test results) of children with acute gastroenteritis caused by non-typhoid Salmonella spp. between January 2010 and October 2012 were evaluated. Differences between the groups for categorical variables were estimated with a chi-square or Fisher exact test; for continuous variables with two independent samples a t test was used. P values < 0.05 were considered statistically significant. Sixty-seven children, 39 (58.2%) males and 28 (41.8%) females aged between 1 - 16 years (mean ± SD: 4.64 ± 2.91), were diagnosed with acute bacterial gastroenteritis caused by non-typhoid Salmonella spp. The main serotypes are Salmonella enteritidis (85%) and Salmonella typhimurium (7.5%). The presenting symptoms were diarrhoea (95.5%), fever (61.1%), vomiting (34.3%), abdominal pain (32.8%), loss of appetite (7.4%) and malaise (7.4%). Fever and dehydration (moderate and/or severe) were detected in 11 (16.4%) patients. The mean leukocyte count was 10.930/μL [95% confidence interval (CI), SD: ± 5.710/μL], neutrophil count was 7.880/μL (95% CI, SD: ± 4.960/μL), CRP was 64.16 mg/L (95% CI, SD: ± 76.24 mg/L), and erythrocyte sedimentation rate was 34.72 mm/hour (95% CI, SD: ± 13.64 mm/h). Stool microscopy was positive for leukocytes in 18 patients (26.8%). The definitive diagnosis was made with positive stool culture (n = 65) and/or PCR test (n = 4). Viral antigen positivity was detected in 10 patients (14.9%), evaluated as viral co-infection and false positive results. Antibiotic therapy and hospitalization were required in 26 (38.8%) and 23 (34.3%) patients, respectively. Salmonella carriage was detected in one patient (1.5%). Bloody diarrhoea, leukocytes in stool with an increased erythrocyte sedimentation rate and a CRP level without overt leukocytosis may indicate Salmonella infection. Viral antigens may cause false positive results in fast antigen tests in cases where clinical and laboratory findings indicate bacterial aetiology. Stool culture is a reference method in diagnosis whereas some agents may be detected via molecular techniques (polymerase chain reaction) in spite of negative culture. Multiplex polymerase chain reaction may be used to detect Salmonella spp. and may reveal false positivity for viruses as well as the detection of other bacteria.

Prostate Cancer Screening: Should You Get a PSA Test?

MedlinePlus

... Mayo Clinic Staff Cancer screening tests — including the prostate-specific antigen (PSA) test to look for signs of prostate ... of harm to the person undergoing the testing. Prostate-specific antigen (PSA) is a protein produced by both cancerous ( ...

A live attenuated BCG vaccine overexpressing multistage antigens Ag85B and HspX provides superior protection against Mycobacterium tuberculosis infection.

PubMed

Yuan, Xuefeng; Teng, Xindong; Jing, Yukai; Ma, Jilei; Tian, Maopeng; Yu, Qi; Zhou, Lei; Wang, Ruibo; Wang, Weihua; Li, Li; Fan, Xionglin

2015-12-01

Tuberculosis (TB) remains one of the most menacing infectious diseases, although attenuated Mycobacterium bovis Bacillus Calmette-Guerin (BCG) vaccine has been widely used to protect children against primary TB. There are increasing evidences that rapid growing and dormant Mycobacterium tuberculosis (M. tuberculosis) coexist in vivo after infection. However, BCG vaccine only elicits cell-mediated immune responses to secretory antigens expressed by rapid growing pathogen. BCG vaccine is thus unable to thwart the reactivation of latent tuberculosis infection (LTBI), and its protection wanes over age after neonatal immunization. In order to extend its ability for a durable protection, a novel recombinant BCG (rBCG) strain, named rBCG::XB, was constructed by overexpressing immunodominant multistage antigens of Ag85B and HspX, which are expressed by both rapid replicating and dormant M. tuberculosis. Long-term protective effect and immunogenicity of rBCG::XB were compared with the parental BCG in vaccinated C57BL/6 mice. Our results demonstrated that rBCG::XB provided the stronger and long-lasting protection against M. tuberculosis H37Rv intranasal infection than BCG. The rBCG::XB not only elicited the more durable multistage antigen-specific CD4(+)Th1-biased immune responses and specific polyfunctional CD4(+)T cells but also augmented the CD8(+) CTL effects against Ag85B in vivo. In particular, higher levels of CD4(+) TEM and CD8(+) TCM cells, dominated by IL2(+) CD4(+) and CD8(+) TCM cells, were obtained in the spleen of rBCG::XB vaccinated mice. Therefore, our findings indicate that rBCG::XB is a promising candidate to improve the efficacy of BCG.

Antigenic Drift of the Pandemic 2009 A(H1N1) Influenza Virus in a Ferret Model

PubMed Central

Guarnaccia, Teagan; Carolan, Louise A.; Maurer-Stroh, Sebastian; Lee, Raphael T. C.; Job, Emma; Reading, Patrick C.; Petrie, Stephen; McCaw, James M.; McVernon, Jodie; Hurt, Aeron C.; Kelso, Anne; Mosse, Jennifer; Barr, Ian G.; Laurie, Karen L.

2013-01-01

Surveillance data indicate that most circulating A(H1N1)pdm09 influenza viruses have remained antigenically similar since they emerged in humans in 2009. However, antigenic drift is likely to occur in the future in response to increasing population immunity induced by infection or vaccination. In this study, sequential passaging of A(H1N1)pdm09 virus by contact transmission through two independent series of suboptimally vaccinated ferrets resulted in selection of variant viruses with an amino acid substitution (N156K, H1 numbering without signal peptide; N159K, H3 numbering without signal peptide; N173K, H1 numbering from first methionine) in a known antigenic site of the viral HA. The N156K HA variant replicated and transmitted efficiently between naïve ferrets and outgrew wildtype virus in vivo in ferrets in the presence and absence of immune pressure. In vitro, in a range of cell culture systems, the N156K variant rapidly adapted, acquiring additional mutations in the viral HA that also potentially affected antigenic properties. The N156K escape mutant was antigenically distinct from wildtype virus as shown by binding of HA-specific antibodies. Glycan binding assays demonstrated the N156K escape mutant had altered receptor binding preferences compared to wildtype virus, which was supported by computational modeling predictions. The N156K substitution, and culture adaptations, have been detected in human A(H1N1)pdm09 viruses with N156K preferentially reported in sequences from original clinical samples rather than cultured isolates. This study demonstrates the ability of the A(H1N1)pdm09 virus to undergo rapid antigenic change to evade a low level vaccine response, while remaining fit in a ferret transmission model of immunization and infection. Furthermore, the potential changes in receptor binding properties that accompany antigenic changes highlight the importance of routine characterization of clinical samples in human A(H1N1)pdm09 influenza surveillance. PMID:23671418

Evaluation of latex agglutination test (KAtex) for early diagnosis of kala-azar.

PubMed

Ahsan, M M; Islam, M N; Mollah, A H; Hoque, M A; Hossain, M A; Begum, Z; Islam, M T

2010-07-01

Kala-azar is one of the major public health problem in Bangladesh. But the diagnosis of the problem often is difficult, unusual and time consuming, a simple, noninvasive, easy to perform, reliable and rapid diagnostic test has been a long-felt need of the clinicians. Therefore, the present study was conducted to see the sensitivity and specificity of Latex Agglutination test (KAtex) to detect leishmanial antigen from urine of kala-azar cases. The study was carried out in the department of Paediatrics, Mymensingh Medical College and Hospital, Bangladesh during July to December, 2008. A total of 100 urine samples were collected of which 50 were confirmed kala-azar cases and 50 were age and sex matched controls. Out of 50 kala-azar cases 47 showed positive result of KAtex. The test was also positive in 01 out of 30 healthy controls. None of the febrile controls was positive by KAtex. The sensitivity, specificity, positive predictive value and negative predictive value of the test using presence of LD bodies in splenic and/or bone marrow aspirate as gold standard were 94%, 98%, 97.91% and 94.23% respectively. KAtex is simple, noninvasive, easy to perform, rapid and reliable test for diagnosing kala-azar in endemic area and useful for small, less equipped laboratories as well as for the laboratories with better facilities.

Multiple Diphtheria Antigen-Antibody Systems Investigated by Passive Haemagglutination Techniques and Other Methods

PubMed Central

Fulthorpe, A. J.

1962-01-01

A fair degree of correlation has been found between the in vivo antitoxin content of sera from horses immunized with crude Corynebacterium diphtheriae culture filtrates and the direct agglutinin titre of the sera when tested with sheep cells sensitized with diphtheria toxoid. Haemagglutination inhibition tests at the LA level of test with the same sera showed some rather large discrepancies from the in vivo and further tests with special agglutinin inhibiting toxins suggested that specific antitoxin free from other accessory antibodies might be non-agglutinating, and therefore not titratable by haemagglutination inhibition. The phosphate-stable, pepsin-stable and trypsin-stable antigens isolated from culture filtrates of C. diphtheriae were found to contain extremely small quantities of specific toxoid, and cross titration of each of the three antigen preparations showed that there was very little contamination by other antigens within the group. Absorption of diphtheria antiserum with red cells sensitized with each of the three accessory antigens individually, showed that the antibodies were highly specific and distinct. Absorption of diphtheria antiserum with a mixture of red cells sensitized with the three different antigens removed all demonstrable accessory antibodies, and the absorbed serum would no longer agglutinate cells sensitized with complete diphtheria toxoid. The absorbed serum, however, retained a large proportion of its neutralizing capacity for diphtheria toxin, when titrated in vivo. Titration of each of the accessory antibodies in a number of horse sera by haemagglutination inhibition demonstrated a correlation between the values for the accessory antibodies to the phosphate-stable and pepsin-stable antigens, but no correlation with the values for the antibody to the trypsin-stable antigen, when compared with results of the flocculation test. The relative proportions of diphtheria toxin and of the phosphate-stable and pepsin-stable antigens remained constant at all stages of purification of a filtrate including several recrystallization procedures. However, the concentration of the trypsin-stable antigen declined steadily during the process. The relative proportions of toxin, phosphate-stable and pepsin-stable antigens in several crude culture filtrates were constant under reasonable conditions of storage and very varied where deterioration had occurred. These three antigens also maintained a constant ratio in growing culture, the trypsin-stable antigen did not. PMID:13895880

Anti-CeHV1 antibodies of two cynomolgus macaques cross-react with HSV2 but not HSV1 antigens in ELISA.

PubMed

Coutrot, Edwin; Blancher-Sardou, Marie; Blancher, Antoine

2008-02-01

The aim of the study was to compare the cross-reactivity of macaque anti-CeHV1 antibodies with type 1 and type 2 human herpes simplex viruses (HSV1 and HSV2). We studied the serum of 344 animals which had been tested either positive (n = 39) or negative (n = 305) for the presence of CeHV1 antibodies by expert laboratories. Macaque serums were studied by means of two ELISA: one based on HSV1 antigen-coated wells, the other on polystyrene beads coated with HSV1 and HSV2 antigens in approximately equal proportions. In the serum of two animals originating from Vietnam, we found anti-CeHV1 antibodies cross-reacting with HSV2 but not with HSV1 antigens. For the serum with the highest titer, inhibition by soluble antigens confirmed the high affinity of the antibodies for HSV2 antigens. Tests using HSV1 and HSV2 in a combined way are better suited to macaque screening than tests using only HSV1 antigens.

Evaluation of Ion Torrent sequencing technology for rapid clinical human leucocyte antigen typing.

PubMed

Guerra, Sandra G; Chong, Winnie; Brown, Colin J; Navarrete, Cristina V

2018-06-05

The development of techniques to define the human leucocyte antigen (HLA) region has proven to be challenging due to its high level of polymorphism. Within a clinical laboratory, a technique for high-resolution HLA typing, which is rapid and cost effective is essential. NGS has provided a rapid, high-resolution HLA typing solution, which has reduced the number of HLA ambiguities seen with other typing methods. In this study, the One Lambda NXType NGS kit was tested on the Ion Torrent PGM platform. A total of 362 registry donors from four ethnic populations (Europeans, South Asians, Africans and Chinese) were NGS HLA typed across 9-loci (HLA-A, -B, -C, -DRB1,-DRB345 -DQB1 and -DPB1). Concordance rates of 91%-98% were obtained (for HLA-A, -B, -C, -DRB1, -DQB1 and -DPB1) when compared to historical PCR-SSO HLA types, and the identification of uncommon alleles such as A*24:07:01 and C*04:82 were observed. A turnaround time of four days was achieved for typing 44 samples. However, some limitations were observed; primer locations did not allow all ambiguities to be resolved for HLA Class II where Exon I and IV amplification are needed (HLA-DRB1*04:07:01/04:92, HLA-DRB1*09:01:02/*09:21 and HLA-DRB1*12:01:01/*12:10). This study has demonstrated high-resolution typing by NGS can be achieved in an acceptable turnaround time for a clinical laboratory; however, the Ion Torrent workflow has some technical limitations that should be addressed. © 2018 John Wiley & Sons Ltd.

Serology of Paracoccidioidomycosis Due to Paracoccidioides lutzii

PubMed Central

Gegembauer, Gregory; Araujo, Leticia Mendes; Pereira, Edy Firmina; Rodrigues, Anderson Messias; Paniago, Anamaria Mello Miranda; Hahn, Rosane Christine; de Camargo, Zoilo Pires

2014-01-01

Paracoccidioides lutzii is a new agent of paracoccidioidomycosis (PCM) and has its epicenter localized to the Central-West region of Brazil. Serological diagnosis of PCM caused by P. lutzii has not been established. This study aimed to develop new antigenic preparations from P. lutzii and to apply them in serological techniques to improve the diagnosis of PCM due to P. lutzii. Paracoccidioides lutzii exoantigens, cell free antigen (CFA), and a TCA-precipitated antigen were evaluated in immunodiffusion (ID) tests using a total of 89 patient sera from the Central-West region of Brazil. Seventy-two sera were defined as reactive for P. brasiliensis using traditional antigens (AgPbB339 and gp43). Non-reactive sera for traditional antigens (n = 17) were tested with different P. lutzii preparations and P. lutzii CFA showed 100% reactivity. ELISA was found to be a very useful test to titer anti-P. lutzii antibodies using P. lutzii-CFA preparations. Sera from patients with PCM due to P. lutzii presented with higher antibody titers than PCM due to P. brasiliensis and heterologous sera. In western blot, sera from patients with PCM due to P. lutzii were able to recognize antigenic molecules from the P. lutzii-CFA antigen, but sera from patients with PCM due to P. brasiliensis could not recognize any P. lutzii molecules. Due to the facility of preparing P. lutzii CFA antigens we recommend its use in immunodiffusion tests for the diagnosis of PCM due to P. lutzii. ELISA and western blot can be used as complementary tests. PMID:25032829

Comparison of Various Methods in the Diagnosis of Entamoeba histolytica in Stool and Serum Specimens

PubMed Central

Uslu, Hakan; Aktas, Osman; Uyanik, Muhammet Hamidullah

2016-01-01

Objective: Entamoeba histolytica is indistinguishable from Entamoeba dispar in direct microscopic examination. A definitive diagnosis of E. histolytica is important in terms of the treatment of the patient and to avoid unnecessary costs. This study’s aim is to determine the prevalence of E. histolytica and to make a comparison of the different diagnostic tests in the patients specimens defined as E. histolytica/E. dispar infection. Materials and Methods: Faecal and serum specimens of 90 patients defined as E. histolytica/E. dispar with microscopy (wet mount examination with 0.85% saline and Lugol’s iodine) were examined. Stool samples were examined by trichrome staining for trophozoites and cysts and by immunoassay methods for specific adhesin antigens (Wampole ® E. histolytica II antigen testing) and for specific serine-rich 30 kD membrane protein (Serazym® E. histolytica antigen testing). Anti-E. histolytica antibodies were investigated using a latex slide test and indirect hemagglutination methods in serum specimens. Results: Presence of E. histolytica was not confirmed in 31.1% cases with trichrome staining, 62.2% of the Wampole antigen test, 64.4%, of the Serazym antigen test, 73.3% of the indirect hemagglutination test and 75.6%. of the latex agglutination. Considering the common results from Wampole and Serazym antigen testing as a reference standard, the specificity/sensitivity is 100/53.85% for trichrome staining, 75.00/98.11% for the latex agglutination test and 78.57/96.77% for the indirect hemagglutination test. Conclusion: It has been shown that investigation of E. histolytica in stools by direct wet-smear microscopy alone can cause significant false positive results. To obtain a reliable diagnosis for E. histolytica and to avoid unnecessary treatment for this parasite, at least one more specific assay, particularly an antigen testing and microscopy, is required. PMID:27551176

Engineered artificial antigen presenting cells facilitate direct and efficient expansion of tumor infiltrating lymphocytes

PubMed Central

2011-01-01

Background Development of a standardized platform for the rapid expansion of tumor-infiltrating lymphocytes (TILs) with anti-tumor function from patients with limited TIL numbers or tumor tissues challenges their clinical application. Methods To facilitate adoptive immunotherapy, we applied genetically-engineered K562 cell-based artificial antigen presenting cells (aAPCs) for the direct and rapid expansion of TILs isolated from primary cancer specimens. Results TILs outgrown in IL-2 undergo rapid, CD28-independent expansion in response to aAPC stimulation that requires provision of exogenous IL-2 cytokine support. aAPCs induce numerical expansion of TILs that is statistically similar to an established rapid expansion method at a 100-fold lower feeder cell to TIL ratio, and greater than those achievable using anti-CD3/CD28 activation beads or extended IL-2 culture. aAPC-expanded TILs undergo numerical expansion of tumor antigen-specific cells, remain amenable to secondary aAPC-based expansion, and have low CD4/CD8 ratios and FOXP3+ CD4+ cell frequencies. TILs can also be expanded directly from fresh enzyme-digested tumor specimens when pulsed with aAPCs. These "young" TILs are tumor-reactive, positively skewed in CD8+ lymphocyte composition, CD28 and CD27 expression, and contain fewer FOXP3+ T cells compared to parallel IL-2 cultures. Conclusion Genetically-enhanced aAPCs represent a standardized, "off-the-shelf" platform for the direct ex vivo expansion of TILs of suitable number, phenotype and function for use in adoptive immunotherapy. PMID:21827675

A Single Subset of Dendritic Cells Controls the Cytokine Bias of Natural Killer T Cell Responses to Diverse Glycolipid Antigens

PubMed Central

Arora, Pooja; Baena, Andres; Yu, Karl O.A.; Saini, Neeraj K.; Kharkwal, Shalu S.; Goldberg, Michael F.; Kunnath-Velayudhan, Shajo; Carreño, Leandro J.; Venkataswamy, Manjunatha M.; Kim, John; Lazar-Molnar, Eszter; Lauvau, Gregoire; Chang, Young-tae; Liu, Zheng; Bittman, Robert; Al-Shamkhani, Aymen; Cox, Liam R.; Jervis, Peter J.; Veerapen, Natacha; Besra, Gurdyal S.; Porcelli, Steven A.

2014-01-01

Summary Many hematopoietic cell types express CD1d and are capable of presenting glycolipid antigens to invariant natural killer T cells (iNKT cells). However, the question of which cells are the principal presenters of glycolipid antigens in vivo remains controversial, and it has been suggested that this might vary depending on the structure of a particular glycolipid antigen. Here we have shown that a single type of cell, the CD8α+ DEC-205+ dendritic cell, was mainly responsible for capturing and presenting a variety of different glycolipid antigens, including multiple forms of α-galactosylceramide that stimulate widely divergent cytokine responses. After glycolipid presentation, these dendritic cells rapidly altered their expression of various costimulatory and coinhibitory molecules in a manner that was dependent on the structure of the antigen. These findings show flexibility in the outcome of two-way communication between CD8α+ dendritic cells and iNKT cells, providing a mechanism for biasing toward either proinflammatory or anti-inflammatory responses. PMID:24412610

Oral delivery of plant-derived HIV-1 p24 antigen in low doses shows a superior priming effect in mice compared to high doses.

PubMed

Lindh, Ingrid; Bråve, Andreas; Hallengärd, David; Hadad, Ronza; Kalbina, Irina; Strid, Åke; Andersson, Sören

2014-04-25

During early infection with human immunodeficiency virus type 1 (HIV-1), there is a rapid depletion of CD4(+) T-cells in the gut-associated lymphoid tissue (GALT) in the gastrointestinal tract. Therefore, immediate protection at these surfaces is of high priority for the development of an HIV-1 vaccine. Thus, transgenic plants expressing HIV-1 antigens, which are exposed to immune competent cells in the GALT during oral administration, can be interesting as potential vaccine candidates. In the present study, we used two HIV-1 p24 antigen-expressing transgenic plant systems, Arabidopsis thaliana and Daucus carota, in oral immunization experiments. Both transgenic plant systems showed a priming effect in mice and induced humoral immune responses, which could be detected as anti-p24-specific IgG in sera after an intramuscular p24 protein boost. Dose-dependent antigen analyses using transgenic A. thaliana indicated that low p24 antigen doses were superior to high p24 antigen doses. Copyright © 2014. Published by Elsevier Ltd.

Analysis of antigen-specific B-cell memory directly ex vivo.

PubMed

McHeyzer-Williams, Louise J; McHeyzer-Williams, Michael G

2004-01-01

Helper T-cell-regulated B-cell memory develops in response to initial antigen priming as a cellular product of the germinal center (GC) reaction. On antigen recall, memory response precursors expand rapidly with exaggerated differentiation into plasma cells to produce the high-titer, high-affinity antibody(Ab) that typifies the memory B-cell response in vivo. We have devised a high-resolution flow cytometric strategy to quantify the emergence and maintenance of antigen-specific memory B cells directly ex vivo. Extended cell surface phenotype establishes a level of cellular diversity not previously appreciated for the memory B-cell compartment. Using an "exclusion transfer" strategy, we ascertain the capacity of two distinct memory B-cell populations to transfer antigen-specific memory into naive adoptive hosts. Finally, we sequence expressed messenger ribonucleic acid (mRNA) from single cells within the population to estimate the level of somatic hypermutation as the best molecular indicator of B-cell memory. In this chapter, we describe the methods used in each of these four sections that serve to provide high-resolution quantification of antigen-specific B-cell memory responses directly ex vivo.

Antenatal noninvasive DNA testing: clinical experience and impact.

PubMed

Ferres, Millie A; Hui, Lisa; Bianchi, Diana W

2014-08-01

Nearly two decades ago, the discovery of circulating cell-free fetal DNA in maternal blood created a paradigm shift in prenatal testing. Recent advances in DNA sequencing technology have facilitated the rapid translation of DNA-based testing into clinical antenatal care. In this review, we summarize the technical approaches and current clinical applications of noninvasive testing using cell-free DNA in maternal plasma. We discuss the impact of these tests on clinical care, outline proposed integration models, and suggest future directions for the field. The use of cell-free DNA in maternal blood for the detection of fetal rhesus D antigen status, fetal sex, and common whole chromosomal aneuploidies is now well established, although testing for aneuploidy is still considered screening and not diagnostic. Further advances in technology and bioinformatics may see future clinical applications extend to the noninvasive detection of fetal subchromosomal aneuploidy, single gene disorders, and the entire fetal genome. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Human Parainfluenza Virus-3 can be Targeted by Rapidly ex vivo Expanded T-Lymphocytes

PubMed Central

McLaughlin, Lauren P.; Lang, Haili; Williams, Elizabeth; Wright, Kaylor E.; Powell, Allison; Cruz, Conrad R; Colberg-Poley, Anamaris M.; Barese, Cecilia; Hanley, Patrick J.; Bollard, Catherine M.; Keller, Michael D.

2016-01-01

Background Human Parainfluenza virus-3 (HPIV) is a common cause of respiratory infection in immunocompromised patients, and presently has no effective therapies. Virus-specific T-cell therapy has been successful for the treatment or prevention of viral infections in immunocompromised patients, but requires determination of T-cell antigens on targeted viruses. Methods HPIV3-specific T cells were expanded from peripheral blood of healthy donors using a rapid generation protocol targeting four HPIV3 proteins. Immunophenotyping was performed by flow cytometry. Viral specificity was determined by IFN-γ ELISpot, intracellular cytokine staining, and cytokine measurements from culture supernatants by Luminex assay. Cytotoxic activity was tested by 51Cr release and CD107a mobilization assays. Virus-specific T-cells targeting 6 viruses were then produced by rapid protocol, and the phenotype of HPIV3-specific T-cells was determined by immunomagnetic sorting for IFN-γ producing cells. Results HPIV3-specific T cells were expanded from 13 healthy donors. HPIV3-specific T-cells showed a CD4+ predominance (mean CD4:CD8 ratio 2.89), and demonstrated specificity for multiple HPIV3 antigens. The expanded T-cells were polyfunctional based on cytokine production, but only had a minor cytotoxic component. T cells targeting six viruses in a single product similarly showed HPIV3 specificity, with a predominant effector memory phenotype (CD3+/CD45RA-/CCR7-) in responder cells. Discussion HPIV3-specific T cells can be produced using a rapid ex vivo protocol from healthy donors and are predominantly CD4+ T-cells with Th1 activity. HPIV3 epitopes can also be successfully targeted alongside multiple other viral epitopes in production of 6-virus T-cells, without loss of HPIV3 specificity. These products may be clinically beneficial to combat HPIV3 infections by adoptive T-cell therapy in immune compromised patients. PMID:27692559

Development of a Colloidal Gold-Based Immunochromatographic Strip for Rapid Detection of Klebsiella pneumoniae Serotypes K1 and K2.

PubMed

Siu, L Kristopher; Tsai, Yu-Kuo; Lin, Jung-Chung; Chen, Te-Li; Fung, Chang-Phone; Chang, Feng-Yee

2016-12-01

In this study, a novel colloidal gold-based immunochromatographic strip (ICS) containing anti-Klebsiella pneumoniae capsular polysaccharide polyclonal antibodies was developed to specifically detect K. pneumoniae serotypes K1 and K2. Capsular polysaccharide K1 and K2 antigens were first used to produce polyclonal anti-K1 and anti-K2 antibodies. Reference strains with different serotypes, nontypeable K. pneumoniae strains, and other bacterial species were then used to assess the sensitivity and specificity of these test strips. The detection limit was found to be 10 5 CFU, and the ICSs were stable for 6 months when stored at room temperature. No false-positive or false-negative results were observed, and equivalent results were obtained compared to those of more conventional test methods, such as PCR or serum agglutination. In conclusion, the ICS developed here requires no technical expertise and allows for the specific, rapid, and simultaneous detection of K. pneumoniae serotypes K1 and K2. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Development of flow-through and dip-stick immunoassays for screening of sulfonamide residues.

PubMed

Zhang, Hongyan; Zhang, Yan; Wang, Shuo

2008-08-20

Two formats of membrane-based competitive enzyme immunoassays (flow-through and dip-stick) have been developed for the screening of sulfonamide residues in pig muscle and milk. Membrane was coated with anti-sulfonamide antibody and a sulfonamide hapten D2-horseradish peroxidase (HRP) conjugant was used as the labeled antigen for competitive assay of sulfonamides. Visual detection limits of the flow-through or dip-stick assay were 1-5 microg L(-1) or 1-10 microg L(-1) in buffer for seven sulfonamides, respectively. Assay validation was performed using samples spiked with single sulfonamide, spiked samples were tested using the developed strip assays and results were compared with those obtained by a validated high-performance liquid chromatograph (HPLC) method. Results showed that the two strip assays were correlated well with HPLC, respectively. With assay times of 5 min (flow-through) and 15 min (dip-stick), these rapid tests could offer simple, rapid and cost-effective on-site screening tools to detect sulfonamides in pig muscle (flow-through or dip-stick) or milk (only dip-stick).

Development of an indirect competitive enzyme-linked immunosorbent assay applied to the Botrytis cinerea quantification in tissues of postharvest fruits

PubMed Central

2011-01-01

Background Botrytis cinerea is a phytopathogenic fungus responsible for the disease known as gray mold, which causes substantial losses of fruits at postharvest. This fungus is present often as latent infection and an apparently healthy fruit can deteriorate suddenly due to the development of this infection. For this reason, rapid and sensitive methods are necessary for its detection and quantification. This article describes the development of an indirect competitive enzyme-linked immunosorbent assay (ELISA) for quantification of B. cinerea in apple (Red Delicious), table grape (pink Moscatel), and pear (William's) tissues. Results The method was based in the competition for the binding site of monoclonal antibodies between B. cinerea antigens present in fruit tissues and B. cinerea purified antigens immobilized by a crosslinking agent onto the surface of the microtiter plates. The method was validated considering parameters such as selectivity, linearity, precision, accuracy and sensibility. The calculated detection limit was 0.97 μg mL-1 B. cinerea antigens. The immobilized antigen was perfectly stable for at least 4 months assuring the reproducibility of the assay. The fungus was detected and quantified in any of the fruits tested when the rot was not visible yet. Results were compared with a DNA quantification method and these studies showed good correlation. Conclusions The developed method allowed detects the presence of B. cinerea in asymptomatic fruits and provides the advantages of low cost, easy operation, and short analysis time determination for its possible application in the phytosanitary programs of the fruit industry worldwide. PMID:21970317

Evaluation of whole blood IFNgamma test using PPD and recombinant antigen challenge for diagnosis of pulmonary and extra-pulmonary tuberculosis.

PubMed

Kalantri, Yatiraj; Hemvani, Nanda; Chitnis, D S

2009-06-01

Quantiferon TB gold (QFT-G) with recombinant antigen cocktail is well evaluated for diagnosis of pulmonary tuberculosis (PTB). However, diagnosis of extra-pulmonary tuberculosis (EPTB) is more difficult due to limitations of conventional techniques. This study compares recombinant antigens based QFT-G and low cost PPD based interferon test for the diagnosis of PTB and EPTB. IFNgamma release, with recombinant antigens and PPD, was assayed by ELISA from 140 cases of EPTB, 100 cases of PTB along with acid fast bacillus (AFB) detection, AFB culture on LJ and MGIT BACTEC. Sensitivity and specificity for QFT-G recombinant antigens was 84.29% and 96%, while for PPD based interferon was 70% and 84% for EPTB group. The sensitivity was far superior to AFB smear and culture for both the antigens. Nine samples were identified as non-tubercular mycobacteria (NTM) in the EPTB group and all were negative for QFT-G, but six of them were positive for PPD based test. Results of the study show that QFT-G using recombinant antigen is sensitive and specific for both PTB and EPTB diagnosis. The PPD based test is economic and offers comparable performance for PTB and EPTB diagnosis and also useful for diagnosis of NTM.

Influence of soya antigen levels in milk replacers on the disruption of intestinal motility patterns in calves sensitive to soya.

PubMed

Lalles, J P; Benkredda, D; Toullec, R

1995-09-01

An experiment was designed to determine the soya antigen levels in milk replacers above which gastrointestinal disorders appeared in preruminant calves previously sensitized to antigenic soya by feeding a soya-based milk replacer for 3 months. These calves were then equipped with wire electrodes on the duodenum and the mid-jejunum. The sensitization was visualized using direct skin testing, plasma anti-soya antibody determination and intestinal myoelectric activity recording. After sensitization, the calves were occasionally fed liquid test meals containing various proportions of antigenic soya and whey. The soya-fed calves displayed larger 24 h skin reactions to beta-conglycinin and higher plasma anti-soya antibody titres than the controls maintained on a skim-milk based milk replacer. Disturbances of the myoelectric activity patterns were recorded on the duodenum and mid-jejunum after feeding antigenic soya, but not non-antigenic soya or milk protein, in the soya-sensitized calves. When the level of antigens was varied, disorders in the jejunal motility patterns appeared when antigenic soya provided one-third or more of the dietary protein in the test meals, although some abnormalities were evident at lower incorporation rates. The major change was a reduction in the mean duration of the jejunal migrating motor complexes which was essentially accounted for by a decrease in the mean duration of the phase I (quiescence). This level of antigens corresponded approximately to 14 and 12 mg of immunoreactive glycinin and beta-conglycinin respectively, per gram of protein intake, i.e. 80 and 70 mg/kg0.75 per test meal.

Topical CpG Adjuvantation of a Protein-Based Vaccine Induces Protective Immunity to Listeria monocytogenes

PubMed Central

Cheng, Wing Ki; Wee, Kathleen; Kollmann, Tobias R.

2014-01-01

Robust CD8+ T cell responses are essential for immune protection against intracellular pathogens. Using parenteral administration of ovalbumin (OVA) protein as a model antigen, the effect of the Toll-like receptor 9 (TLR9) agonist, CpG oligodeoxynucleotide (ODN) 1826, as an adjuvant delivered either topically, subcutaneously, or intramuscularly on antigen-specific CD8+ T cell responses in a mouse model was evaluated. Topical CpG adjuvant increased the frequency of OVA-specific CD8+ T cells in the peripheral blood and in the spleen. The more effective strategy to administer topical CpG adjuvant to enhance CD8+ T cell responses was single-dose administration at the time of antigen injection with a prime-boost regimen. Topical CpG adjuvant conferred both rapid and long-lasting protection against systemic challenge with recombinant Listeria monocytogenes expressing the cytotoxic T lymphocyte (CTL) epitope of OVA257–264 (strain Lm-OVA) in a TLR9-dependent manner. Topical CpG adjuvant induced a higher proportion of CD8+ effector memory T cells than parenteral administration of the adjuvant. Although traditional vaccination strategies involve coformulation of antigen and adjuvant, split administration using topical adjuvant is effective and has advantages of safety and flexibility. Split administration of topical CpG ODN 1826 with parenteral protein antigen is superior to other administration strategies in enhancing both acute and memory protective CD8+ T cell immune responses to subcutaneous protein vaccines. This vaccination strategy induces rapid and persistent protective immune responses against the intracellular organism L. monocytogenes. PMID:24391136

CD8+ T-cell responses rapidly select for antigen-negative tumor cells in the prostate.

PubMed

Bak, S Peter; Barnkob, Mike Stein; Wittrup, K Dane; Chen, Jianzhu

2013-12-01

Stimulation of patients' immune systems for the treatment of solid tumors is an emerging therapeutic paradigm. The use of enriched autologous T cells for adoptive cell therapy or vaccination with antigen-loaded dendritic cells have shown clinical efficacy in melanoma and prostate cancer, respectively. However, the long-term effects of immune responses on selection and outgrowth of antigen-negative tumor cells in specific tumor types must be determined to understand and achieve long-term therapeutic effects. In this study, we have investigated the expression of a tumor-specific antigen in situ after treatment with tumor-specific CD8(+) T cells in an autochthonous mouse model of prostate cancer. After T-cell treatment, aggregates of dead antigen-positive tumor cells were concentrated in the lumen of the prostate gland and were eventually eliminated from the prostate tissue. Despite the elimination of antigen-positive tumor cells, prostate tumor continued to grow in T-cell-treated mice. Interestingly, the remaining tumor cells were antigen negative and downregulated MHC class I expression. These results show that CD8(+) T cells are effective in eliminating antigen-bearing prostate tumor cells but they also can select for the outgrowth of antigen-negative tumor cells. These findings provide insights into the requirements for an effective cancer immunotherapy within the prostate that not only induces potent immune responses but also avoids selection and outgrowth of antigen-negative tumor cells. ©2013 AACR.

Test characteristics of the SD FK80 Plasmodium falciparum/Plasmodium vivax malaria rapid diagnostic test in a non-endemic setting

PubMed Central

2009-01-01

Background The SD FK80 P.f/P.v Malaria Antigen Rapid Test (Standard Diagnostics, Korea) (FK80) is a three-band malaria rapid diagnostic test detecting Plasmodium falciparum histidine-rich protein-2 (HRP-2) and Plasmodium vivax-specific lactate dehydrogenase (Pv-pLDH). The present study assessed its performance in a non-endemic setting. Methods Stored blood samples (n = 416) from international travellers suspected of malaria were used, with microscopy corrected by PCR as the reference method. Samples infected by Plasmodium falciparum (n = 178), Plasmodium vivax (n = 99), Plasmodium ovale (n = 75) and Plasmodium malariae (n = 24) were included, as well as 40 malaria negative samples. Results Overall sensitivities for the diagnosis of P. falciparum and P. vivax were 91.6% (95% confidence interval (CI): 86.2% - 95.0%) and 75.8% (65.9% - 83.6%). For P. falciparum, sensitivity at parasite densities ≥ 100/μl was 94.6% (88.8% - 97.6%); for P. vivax, sensitivity at parasite densities ≥ 500/μl was 86.8% (75.4% - 93.4%). Four P. falciparum samples showed a Pv-pLDH line, three of them had parasite densities exceeding 50.000/μl. Two P. vivax samples, one P. ovale and one P. malariae sample showed a HRP-2 line. For the HRP-2 and Pv-pLDH lines, respectively 81.4% (136/167) and 55.8% (43/77) of the true positive results were read as medium or strong line intensities. The FK80 showed good reproducibility and reliability for test results and line intensities (kappa values for both exceeding 0.80). Conclusion The FK80 test performed satisfactorily in diagnosing P. falciparum and P. vivax infections in a non-endemic setting. PMID:19930609

Adoptive T-cell immunotherapy of cancer using chimeric antigen receptor-grafted T cells.

PubMed

Davies, David Marc; Maher, John

2010-06-01

Harnessing the power of the immune system to target cancer has long been a goal of tumor immunologists. One avenue under investigation is the modification of T cells to express a chimeric antigen receptor (CAR). Expression of such a receptor enables T-cell specificity to be redirected against a chosen tumor antigen. Substantial research in this field has been carried out, incorporating a wide variety of malignancies and tumor-associated antigens. Ongoing investigations will ensure this area continues to expand at a rapid pace. This review will explain the evolution of CAR technology over the last two decades in addition to detailing the associated benefits and disadvantages. The outcome of recent phase I clinical trials and the impact that these have had upon the direction of future research in this field will also be addressed.

Routine use of point-of-care tests: usefulness and application in clinical microbiology.

PubMed

Clerc, O; Greub, G

2010-08-01

Point-of-care (POC) tests offer potentially substantial benefits for the management of infectious diseases, mainly by shortening the time to result and by making the test available at the bedside or at remote care centres. Commercial POC tests are already widely available for the diagnosis of bacterial and viral infections and for parasitic diseases, including malaria. Infectious diseases specialists and clinical microbiologists should be aware of the indications and limitations of each rapid test, so that they can use them appropriately and correctly interpret their results. The clinical applications and performance of the most relevant and commonly used POC tests are reviewed. Some of these tests exhibit insufficient sensitivity, and should therefore be coupled to confirmatory tests when the results are negative (e.g. Streptococcus pyogenes rapid antigen detection test), whereas the results of others need to be confirmed when positive (e.g. malaria). New molecular-based tests exhibit better sensitivity and specificity than former immunochromatographic assays (e.g. Streptococcus agalactiae detection). In the coming years, further evolution of POC tests may lead to new diagnostic approaches, such as panel testing, targeting not just a single pathogen, but all possible agents suspected in a specific clinical setting. To reach this goal, the development of serology-based and/or molecular-based microarrays/multiplexed tests will be needed. The availability of modern technology and new microfluidic devices will provide clinical microbiologists with the opportunity to be back at the bedside, proposing a large variety of POC tests that will allow quicker diagnosis and improved patient care.