40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

Code of Federal Regulations, 2010 CFR

2010-07-01

... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a...

40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

Code of Federal Regulations, 2011 CFR

2011-07-01

... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a...

40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

Code of Federal Regulations, 2013 CFR

2013-07-01

... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a...

40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

Code of Federal Regulations, 2014 CFR

2014-07-01

... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a...

40 CFR 174.506 - Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of...

Code of Federal Regulations, 2012 CFR

2012-07-01

... Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. 174.506 Section 174.506... thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry34Ab1 and Cry35Ab1 proteins in corn are exempted from the requirement of a...

Dietary Factors Associated with Plasma Thyroid Peroxidase and Thyroglobulin Antibodies.

PubMed

Matana, Antonela; Torlak, Vesela; Brdar, Dubravka; Popović, Marijana; Lozić, Bernarda; Barbalić, Maja; Perica, Vesna Boraska; Punda, Ante; Polašek, Ozren; Hayward, Caroline; Zemunik, Tatijana

2017-10-28

The knowledge about dietary habits and their influence in the development of autoimmune thyroid disease is insufficient. The aim of this study was to analyse the association of dietary factors and plasma thyroid peroxidase antibodies (TPO-Ab) and/or thyroglobulin antibodies (Tg-Ab). The study enrolled 1887 participants originating from the South Croatia. Participants with elevated plasma TPO-Ab and/or Tg-Ab were defined as cases ( n = 462) and those with TPO-Ab and/or Tg-Ab within referent values were defined as controls ( n = 1425). Dietary intake was evaluated according to a food frequency questionnaire containing 58 food items. Principal component analysis was used to group food items into dietary groups. We used logistic regression analysis to examine dietary groups associated with positive plasma TPO-Ab and/or Tg-Ab. The results indicate that the dietary group with frequent consumption of animal fats and butter is associated with positive plasma TPO-Ab and/or Tg-Ab ( p = 0.01). The dietary group with frequent consumption of vegetables as well as the dietary group with high consumption of dried fruit, nuts, and muesli are associated with negative findings of TPO-Ab and/or Tg-Ab ( p = 0.048 and p = 0.02, respectively). We showed that the anti-inflammatory dietary groups are associated with the negative findings of plasma TPO-Ab and/or Tg-Ab.

Large haematoxylin-stainable keratohyaline granules in solar keratoses: immunohistochemical comparison using anti-Ted-H-1 antibody and antiloricrin antibody.

PubMed

Takahashi, M; Horiuchi, Y; Tezuka, T

2005-11-01

Our previous study showed that large keratohyaline granules (KHG) in molluscum contagiosum that stained with haematoxylin also reacted with anti-Ted-H-1 monoclonal antibody (mAb), but not with antifilaggrin mAb or antiloricrin polyclonal antibody (pAb). This finding indicated that the Ted-H-1 antigenic protein is a haematoxylin-stainable protein in KHG. To clarify the identity of the major component protein of the large KHG in solar keratosis, another disorder in which large KHG are observed. An enzyme immunohistochemical study was performed using antifilaggrin mAb, anti-Ted-H-1 mAb and antiloricrin pAb. Immunofluorescent double staining and immunoelectron microscopic analyses were performed using anti-Ted-H-1 mAb and antiloricrin pAb. Antifilaggrin mAb, anti-Ted-H-1 mAb and antiloricrin pAb reacted with normal KHG in nonlesional skin of solar keratosis, while only anti-Ted-H-1 mAb reacted with the large KHG in the lesions of solar keratosis. Antifilaggrin mAb did not react with large KHG. Antiloricrin pAb reacted with the cell membrane of the stratum granulosum, but not with large KHG. These findings suggest that the haematoxylin-stainable protein in the large KHG would be a Ted-H-1 antigen protein which was neither filaggrin nor loricrin.

Radiolytic Synthesis of Pt-Particle/ABS Catalysts for H₂O₂ Decomposition in Contact Lens Cleaning.

PubMed

Ohkubo, Yuji; Aoki, Tomonori; Seino, Satoshi; Mori, Osamu; Ito, Issaku; Endo, Katsuyoshi; Yamamura, Kazuya

2017-08-23

A container used in contact lens cleaning requires a Pt plating weight of 1.5 mg for H₂O₂ decomposition although Pt is an expensive material. Techniques that decrease the amount of Pt are therefore needed. In this study, Pt nanoparticles instead of Pt plating film were supported on a substrate of acrylonitrile-butadiene-styrene copolymer (ABS). This was achieved by the reduction of Pt ions in an aqueous solution containing the ABS substrate using high-energy electron-beam irradiation. Pt nanoparticles supported on the ABS substrate (Pt-particle/ABS) had a size of 4-10 nm. The amount of Pt required for Pt-particle/ABS was 250 times less than that required for an ABS substrate covered with Pt plating film (Pt-film/ABS). The catalytic activity for H₂O₂ decomposition was estimated by measuring the residual H₂O₂ concentration after immersing the catalyst for 360 min. The Pt-particle/ABS catalyst had a considerably higher specific catalytic activity for H₂O₂ decomposition than the Pt-film/ABS catalyst. In addition, sterilization performance was estimated from the initial rate of H₂O₂ decomposition over 60 min. The Pt-particle/ABS catalyst demonstrated a better sterilization performance than the Pt-film/ABS catalyst. The difference between Pt-particle/ABS and Pt-film/ABS was shown to reflect the size of the O₂ bubbles formed during H₂O₂ decomposition.

Neurological Autoantibody Prevalence in Epilepsy of Unknown Etiology.

PubMed

Dubey, Divyanshu; Alqallaf, Abdulradha; Hays, Ryan; Freeman, Matthew; Chen, Kevin; Ding, Kan; Agostini, Mark; Vernino, Steven

2017-04-01

Autoimmune epilepsy is an underrecognized condition, and its true incidence is unknown. Identifying patients with an underlying autoimmune origin is critical because these patients' condition may remain refractory to conventional antiseizure medications but may respond to immunotherapy. To determine the prevalence of neurological autoantibodies (Abs) among adult patients with epilepsy of unknown etiology. Consecutive patients presenting to neurology services with new-onset epilepsy or established epilepsy of unknown etiology were identified. Serum samples were tested for autoimmune encephalitis Abs as well as thyroperoxidase (TPO) and glutamic acid decarboxylase 65 (GAD65) Abs. An antibody prevalence in epilepsy (APE) score based on clinical characteristics was assigned prospectively. Data were collected from June 1, 2015, to June 1, 2016. Presence of neurological Abs. A score based on clinical characteristics was assigned to estimate the probability of seropositivity prior to antibody test results. Good seizure outcome was estimated on the basis of significant reduction of seizure frequency at the first follow-up or seizure freedom. Of the 127 patients (68 males and 59 females) enrolled in the study, 15 were subsequently excluded after identification of an alternative diagnosis. Serum Abs suggesting a potential autoimmune etiology were detected in 39 (34.8%) cases. More than 1 Ab was detected in 7 patients (6.3%): 3 (2.7%) had TPO-Ab and voltage-gated potassium channel complex (VGKCc) Ab, 2 (1.8%) had GAD65-Ab and VGKCc-Ab, 1 had TPO-Ab and GAD65-Ab, and 1 had anti-Hu Ab and GAD65-Ab. Thirty-two patients (28.6%) had a single Ab marker. Among 112 patients included in the study, 15 (13.4%) had TPO-Ab, 14 (12.5%) had GAD65-Ab, 12 (10.7%) had VGKCc (4 of whom were positive for leucine-rich glioma-inactivated protein 1 [LGI1] Ab), and 4 (3.6%) had N-methyl-D-aspartate receptor (NMDAR) Ab. Even after excluding TPO-Ab and low-titer GAD65-Ab, Abs strongly suggesting an autoimmune cause of epilepsy were seen in 23 patients (20.5%). Certain clinical features, such as autonomic dysfunction, neuropsychiatric changes, viral prodrome, faciobrachial dystonic spells or facial dyskinesias, and mesial temporal sclerosis abnormality on magnetic resonance imaging, correlated with seropositivity. The APE score was a useful tool in predicting positive serologic findings. Patients who were Ab positive were more likely to have good seizure outcome than were patients with epilepsy of unknown etiology (15 of 23 [65.2%] vs 24 of 89 [27.0%]; odds ratio, 4.8; 95% CI, 1.8-12.9; P = .002). In patients who were seropositive, reduction in seizure frequency was associated with use of immunomodulatory therapy. Among adult patients with epilepsy of unknown etiology, a significant minority had detectable serum Abs suggesting an autoimmune etiology. Certain clinical features (encoded in the APE score) could be used to identify patients with the highest probability of harboring neurological Abs.

Performance evaluation of four dominant anti-hepatitis B core antigen (HBcAb) kits in Japan for preventing de novo hepatitis B virus (HBV) infection.

PubMed

Kobayashi, Eiji; Deguchi, Matsuo; Kagita, Masanori; Yoshioka, Nori; Kita, Mifumi; Asari, Seishi; Suehisa, Etsuji; Hidaka, Yoh; Iwatani, Yoshinori

2015-01-01

The determination of antibody to hepatitis B core antigen (HBcAb) has become an important means of evaluating the risk factors of de novo hepatitis B virus (HBV) infection before starting intensive immunosuppressive drug therapies. Four dominant HBcAb determination reagents used in Japan were evaluated with HBcIgM, HBsAg, HBsAb, HBeAb, and HBV DNA reagents in order to study their clinical utility. Four kinds of HBcAb reagent kits (HBcAb Total and HBcAb-IgG reagent) were evaluated with 526 clinical specimens, including 344 negative specimens, at Osaka University Hospital. The dynamic range of each kit was evaluated by testing serially diluted serum from pooled sera with high HBcAb concentration. The reagent that showed the largest dynamic range was the Lumipulse HBcAb-N (HBcAb-IgG reagent). Regarding clinical sensitivity and specificity, Centaur HBcAb (HBcAb Total reagent) gave several "doubtful negative" results and ARCHITECT HBcII (HBcAb Total reagent) had the most discrepant positive results. By comparing the cut-off-index distribution of negative specimens using a parameter of "distance from the mean to the cut-off divided by the SD", Centaur was determined to be the best (distance/SD = 12.65), with Lumipulse and Elecsys Anti-HBc (HBcAb Total reagent) in the second group (8.13 and 7.00, respectively), and ARCHITECT rated as the worst (3.25). In this evaluation, Elecsys and Lumipulse HBcAb kits showed good clinical sensitivity and specificity and were considered to be suitable for evaluating the risk factors of de novo HBV infection.

Human Antibodies that Recognize Novel Immunodominant Quaternary Epitopes on the HIV-1 Env Protein

PubMed Central

Hicar, Mark D.; Chen, Xuemin; Sulli, Chidananda; Barnes, Trevor; Goodman, Jason; Sojar, Hakimuddin; Briney, Bryan; Willis, Jordan; Chukwuma, Valentine U.; Kalams, Spyros A.; Doranz, Benjamin J.; Spearman, Paul; Crowe, James E.

2016-01-01

Numerous broadly neutralizing antibodies (Abs) target epitopes that are formed or enhanced during mature HIV envelope formation (i.e. quaternary epitopes). Generally, it is thought that Env epitopes that induce broadly neutralizing Abs are difficult to access and poorly immunogenic because of the characteristic oligomerization, conformational flexibility, sequence diversity and extensive glycosylation of Env protein. To enhance for isolation of quaternary epitope-targeting Abs (QtAbs), we previously used HIV virus-like particles (VLPs) to bind B cells from long-term non-progressor subjects to identify a panel of monoclonal Abs. When expressed as recombinant full-length Abs, a subset of these novel Abs exhibited the binding profiles of QtAbs, as they either failed to bind to monomeric Env protein or showed much higher affinity for Env trimers and VLPs. These QtAbs represented a significant proportion of the B-cell response identified with VLPs. The Ab genes of these clones were highly mutated, but they did not neutralize common HIV strains. We sought to further define the epitopes targeted by these QtAbs. Competition-binding and mapping studies revealed these Abs targeted four separate epitopes; they also failed to compete for binding by Abs to known major neutralizing epitopes. Detailed epitope mapping studies revealed that two of the four epitopes were located in the gp41 subunit of Env. These QtAbs bound pre-fusion forms of antigen and showed differential binding kinetics depending on whether oligomers were produced as recombinant gp140 trimers or as full-length Env incorporated into VLPs. Antigenic regions within gp41 present unexpectedly diverse structural epitopes, including these QtAb epitopes, which may be targeted by the naturally occurring Ab response to HIV infection. PMID:27411063

Analytical and clinical performance of thyroglobulin autoantibody assays in thyroid cancer follow-up.

PubMed

Katrangi, Waddah; Grebe, Stephan K G; Algeciras-Schimnich, Alicia

2017-10-26

While thyroglobulin autoantibodies (TgAb) can result in false low serum thyroglobulin (Tg) immunoassay (IA) measurements, they might also be indicators of disease persistence/recurrence. Hence, accurate TgAb measurement, in addition to Tg quantification, is crucial for thyroid cancer monitoring. We compared the analytical and clinical performance of four commonly used TgAb IAs. We measured Tg by mass spectrometry (Tg-MS) and by four pairs of Tg and TgAb IAs (Beckman, Roche, Siemens, Thermo) in 576 samples. Limit of quantitation (LOQ) and manufacturers' upper reference interval cut-off (URI) were used for comparisons. Clinical performance was assessed by receiving operator characteristics (ROC) curve analysis. Quantitative and qualitative agreement between TgAb-IAs was moderate with R2 of 0.20-0.70 and κ from 0.41-0.66 using LOQ and 0.47-0.71 using URI. In samples with TgAb interference, detection rates of TgAb were similar using LOQ and URI for Beckman, Siemens, and Thermo, but much lower for the Roche TgAb-IA when the URI was used. In TgAb positive cases, the ROC areas under the curve (AUC) for the TgAb-IAs were 0.59 (Beckman), 0.62 (Siemens), 0.59 (Roche), and 0.59 (Thermo), similar to ROC AUCs achieved with Tg. Combining Tg and TgAb measurements improved the ROC AUCs compared to Tg or TgAb alone. TgAb-IAs show significant qualitative and quantitative differences. For 2 of the 4 TgAb-IAs, using the LOQ improves the detection of interfering TgAbs. All assays showed suboptimal clinical performance when used as surrogate markers of disease, with modest improvements when Tg and TgAb were combined.

Performance of the Alere Determine™ HIV-1/2 Ag/Ab Combo Rapid Test with algorithm-defined acute HIV-1 infection specimens.

PubMed

Parker, Monica M; Bennett, S Berry; Sullivan, Timothy J; Fordan, Sally; Wesolowski, Laura G; Wroblewski, Kelly; Gaynor, Anne M

2018-05-14

The capacity of HIV Antigen/Antibody (Ag/Ab) immunoassays (IA) to detect HIV-1 p24 antigen has resulted in improved detection of HIV-1 infections in comparison to Ab-only screening assays. Since its introduction in the US, studies have shown that the Determine HIV-1/2 Ag/Ab Combo assay (Determine Ag/Ab) detects HIV infection earlier than laboratory-based IgM/IgG-sensitive IAs, but its sensitivity for HIV-1 p24 Ag detection is reduced compared to laboratory-based Ag/Ab assays. However, further evaluation is needed to assess its capacity to detect acute HIV-1 infection. To assess the performance of Determine Ag/Ab in serum from acute HIV-1 infections. Select serum specimens that screened reactive on a laboratory-based Ag/Ab IA or IgM/IgG Ab-only IA, with a negative or indeterminate supplemental antibody test and detectable HIV-1 RNA were retrospectively tested with Determine Ag/Ab. Results were compared with those of the primary screening immunoassay to evaluate concordance within this set of algorithm-defined acute infections. Of 159 algorithm-defined acute HIV-1 specimens, Determine Ag/Ab was reactive for 105 resulting in 66.0% concordance. Of 125 that were initially detected by a laboratory-based Ag/Ab IA, 81 (64.8%) were reactive by Determine Ag/Ab. A total of 34 acute specimens were initially detected by a laboratory-based IgM/IgG Ab-only IA and 24 (70.6%) of those were reactive by Determine Ag/Ab. Due to their enhanced sensitivity, laboratory-based Ag/Ab IAs continue to be preferred over the Determine Ag/Ab as the screening method used by laboratories conducting HIV diagnostic testing on serum and plasma specimens. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Oriented conjugates of monoclonal and single-domain antibodies with quantum dots for flow cytometry and immunohistochemistry diagnostic applications

NASA Astrophysics Data System (ADS)

Sukhanova, Alyona; Even-Desrumeaux, Klervi; Millot, Jean-Marc; Chames, Patrick; Baty, Daniel; Artemyev, Mikhail; Oleinikov, Vladimir; Cohen, Jacques H. M.; Nabiev, Igor

2012-03-01

Ideal diagnostic nanoprobes should not exceed 15 nm in size and should contain high-affinity homogeneously oriented capture molecules on their surface. An advanced procedure for antibody (Ab) reduction was used to cleave each Ab into two functional half-Abs, 75-kDa heavy-light chain fragments, each containing an intact antigen-binding site. Affinity purification of half-Abs followed by their linkage to quantum dots (QDs) yielded oriented QD-Ab conjugates whose functionality was considerably improved compared to those obtained using the standard protocols. Ultrasmall diagnostic nanoprobes were engineered through oriented conjugation of QDs with 13-kDa single-domain Abs (sdAbs) derived from llama IgG. sdAbs were tagged with QDs via an additional cysteine residue specifically integrated into the C-terminal region of sdAb using genetic engineering. This approach made it possible to obtain sdAb-QD nanoprobes <12 nm in diameter comprising four copies of sdAbs linked to the same QD in an oriented manner. sdAb-QD conjugates against carcinoembryonic antigen (CEA) and HER2 exhibited an extremely high specificity in flow cytometry; the quality of immunohistochemical labeling of biopsy samples was found to be superior to that of labeling according to the current "gold standard" protocols of anatomo-pathological practice. The nano-bioengineering approaches developed can be extended to oriented conjugation of Abs and sdAbs with different semiconductor, noble metal, or magnetic nanoparticles.

ABO Mistyping of cis-AB Blood Group by the Automated Microplate Technique.

PubMed

Chun, Sejong; Ryu, Mi Ra; Cha, Seung-Yeon; Seo, Ji-Young; Cho, Duck

2018-01-01

The cis -AB phenotype, although rare, is the relatively most frequent of ABO subgroups in Koreans. To prevent ABO mistyping of cis -AB samples, our hospital has applied a combination of the manual tile method with automated devices. Herein, we report cases of ABO mistyping detected by the combination testing system. Cases that showed discrepant results by automated devices and the manual tile method were evaluated. These samples were also tested by the standard tube method. The automated devices used in this study were a QWALYS-3 and Galileo NEO. Exons 6 and 7 of the ABO gene were sequenced. 13 cases that had the cis -AB allele showed results suggestive of the cis -AB subgroup by manual methods, but were interpreted as AB by either automated device. This happened in 87.5% of these cases by QWALYS-3 and 70.0% by Galileo NEO. Genotyping results showed that 12 cases were ABO*cis-AB01/ABO*O01 or ABO*cis-AB01/ABO*O02 , and one case was ABO*cis-AB01/ ABO*A102. Cis -AB samples were mistyped as AB by the automated microplate technique in some cases. We suggest that the manual tile method can be a simple supplemental test for the detection of the cis -AB phenotype, especially in countries with relatively high cis- AB prevalence.

Bridge Crossing Simulator

DTIC Science & Technology

2014-10-07

is counted as. Per the TDTC, a test bridge with longitudinal and/or lateral symmetry under non- eccentric loading can be considered as 1, 2, or 4...Level Run036 3 MLC70T (tracked) BA Run046 6 AB Run055 9 AB Run060 9 BA Run064 12 BA Run071 15 AB Run155 3 MLC96W ( wheeled ) AB...Run331 9 AB Run359 15 AB Run430 12 MLC96W ( wheeled ) BA Run434 12 AB Run447 3 BA Bank Condition: Side Slope, Even Strain Channels High

Growth suppression of colorectal cancer by plant-derived multiple mAb CO17-1A × BR55 via inhibition of ERK1/2 phosphorylation.

PubMed

Kwak, Dong Hoon; Moussavou, Ghislain; Lee, Ju Hyoung; Heo, Sung Youn; Ko, Kisung; Hwang, Kyung-A; Jekal, Seung-Joo; Choo, Young-Kug

2014-11-14

We have generated the transgenic Tabaco plants expressing multiple monoclonal antibody (mAb) CO7-1A × BR55 by cross-pollinating with mAb CO17-1A and mAb BR55. We have demonstrated the anti-cancer effect of plant-derived multiple mAb CO17-1A × BR55. We find that co-treatment of colorectal mAbs (anti-epithelial cellular adhesion molecule (EpCAM), plant-derived monoclonal antibody (mAb(P)) CO17-1A and mAb(P) CO17-1A × BR55) with RAW264.7 cells significantly inhibited the cell growth in SW620 cancer cells. In particular, multi mAb(P) CO17-1A × BR55 significantly and efficiently suppressed the growth of SW620 cancer cells compared to another mAbs. Apoptotic death-positive cells were significantly increased in the mAb(P) CO17-1A × BR55-treated. The mAb(P) CO17-1A × BR55 treatment significantly decreased the expression of B-Cell lymphoma-2 (BCl-2), but the expression of Bcl-2-associated X protein (Bax), and cleaved caspase-3 were markedly increased. In vivo, the mAb(P) CO17-1A × BR55 significantly and efficiently inhibited the growth of colon tumors compared to another mAbs. The apoptotic cell death and inhibition of pro-apoptotic proteins expression were highest by treatment with mAb(P) CO17-1A × BR55. In addition, the mAb(P) CO17-1A × BR55 significantly inhibited the extracellular signal-regulated kinase 1 and 2 (ERK1/2) phosphorylation in cancer cells and tumors. Therefore, this study results suggest that multiple mAb(P) CO17-1A × BR55 has a significant effect on apoptosis-mediated anticancer by suppression of ERK1/2 phosphorylation in colon cancer compared to another mAbs. In light of these results, further clinical investigation should be conducted on mAb(P) CO17-1A × BR55 to determine its possible chemopreventive and/or therapeutic efficacy against human colon cancer.

Memories of AB

NASA Astrophysics Data System (ADS)

Vaks, V. G.

2013-06-01

I had the good fortune to be a student of A. B. Migdal - AB, as we called him in person or in his absence - and to work in the sector he headed at the Kurchatov Institute, along with his other students and my friends, including Vitya Galitsky, Spartak Belyayev and Tolya Larkin. I was especially close with AB in the second half of the 1950s, the years most important for my formation, and AB's contribution to this formation was very great. To this day, I've often quoted AB on various occasions, as it's hard to put things better or more precisely than he did; I tell friends stories heard from AB, because these stories enhance life as AB himself enhanced it; my daughter is named Tanya after AB's wife Tatyana Lvovna, and so on. In what follows, I'll recount a few episodes in my life in which AB played an important or decisive role, and then will share some other memories of AB...

Binding characteristics of anti-atrazine monoclonal antibodies and their fragments synthesised in bacteria and plants.

PubMed

Strachan, G; Grant, S D; Learmonth, D; Longstaff, M; Porter, A J; Harris, W J

1998-09-15

Single-chain antibody fragments (scAb), specific for the herbicide atrazine, have been expressed in the bacterium Escherichia coli and in transgenic tobacco plants. The scAb could be purified as a monomer (monovalent) via a hexa-histidine tail or as a dimer (divalent) by antibody affinity chromatography. In competition ELISA, the bacterial scAb showed the same specificity for atrazine and related triazine herbicides as the parental mAb cell line, but both plant and bacterial monomeric scAbs showed increased sensitivity to free atrazine. Surface plasmon resonance (BIAcore 2000) analysis confirmed that purified scAb, derived from plant or bacteria, retained similar association rates as the mAb. However, the monomeric plant and bacterial scAbs showed a lower affinity for immobilised antigen, than the equivalent dimeric scAbs or mAb. This decrease in affinity was due to a 10 fold slower dissociation rate and is likely due to loss of the avidity contribution of dimeric molecules.

A novel cry2Ab gene from the indigenous isolate Bacillus thuringiensis subsp. kurstaki.

PubMed

Sevim, Ali; Eryüzlü, Emine; Demirbağ, Zihni; Demir, Ismail

2012-01-01

A novel cry2Ab gene was cloned and sequenced from the indigenous isolate of Bacillus thuringiensis subsp. kurstaki. This gene was designated as cry2Ab25 and its sequence revealed an open reading frame of 1,902 bp encoding a 633 aa protein with calculated molecular mass of 70 kDa and pI value of 8.98. The amino acid sequence of the Cry2Ab25 protein was compared with previously known Cry2Ab toxins, and the phylogenetic relationships among them were determined. The deduced amino acid sequence of the Cry2Ab25 protein showed 99% homology to the known Cry2Ab proteins, except for Cry2Ab10 and Cry2Ab12 with 97% homology, and a variation in one amino acid residue in comparison with all known Cry2Ab proteins. The cry2Ab25 gene was expressed in Escherichia coli BL21(DE3) cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) revealed that the Cry2Ab25 protein is about 70 kDa. The toxin expressed in BL21(DE3) exhibited high toxicity against Malacosoma neustria and Rhagoletis cerasi with 73% and 75% mortality after 5 days of treatment, respectively.

Subpopulations of Ig-secreting cells induced by peroxidase immunization: discrimination according to antibody storage and secretion.

PubMed Central

Bernard, J; Jeannesson, P; Thiernesse, N; Zagury, D; Ternynck, T; Avrameas, S

1979-01-01

Mice were injected in their hind footpads with peroxidase (PO) emulsified in Freund's complete adjuvant. The development of cells secreting anti-peroxidase antibody (Ab) and cells secreting immunoglobulins (Ig) were detected in the draining popliteal lymph nodes in the subsequent 35 days, using local haemolysis plaque assay with sheep red cell blood cells coated with either PO or anti-mouse Ig antibody. Plaque-forming cells (PFC) were isolated from the centre of plaques by micromanipulation and after appropriate treatment, were examined by electron microscopy for their intracellular Ab content and in corporation of [3H]-thymidine. Four subpopulations of Ig secreting cells were distinguished: (1) cells secreting Ig without Ab function and not containing intracellular Ab detectable between days 5 and 20; (2) cells secreting Ig without Ab function but containing Ab appearing on day 6 and present throughout the immune response; (3) cells secreting Ab and containing Ab; (4) cells secreting Ab, but without detectable intracellular Ab. These last subpopulations appeared on day 7 and were found in all subsequent assays. The analysis of the kinetics of these subpopulations suggest that cells secreting Ig without Ab function might be precursors of Ab secreting cells. Images Figure 2a Figure 2b Figure 4 PMID:374259

Combination of two anti-CD5 monoclonal antibodies synergistically induces complement-dependent cytotoxicity of chronic lymphocytic leukaemia cells.

PubMed

Klitgaard, Josephine L; Koefoed, Klaus; Geisler, Christian; Gadeberg, Ole V; Frank, David A; Petersen, Jørgen; Jurlander, Jesper; Pedersen, Mikkel W

2013-10-01

The treatment of chronic lymphocytic leukaemia (CLL) has been improved by introduction of monoclonal antibodies (mAbs) that exert their effect through secondary effector mechanisms. CLL cells are characterized by expression of CD5 and CD23 along with CD19 and CD20, hence anti-CD5 Abs that engage secondary effector functions represent an attractive opportunity for CLL treatment. Here, a repertoire of mAbs against human CD5 was generated and tested for ability to induce complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC) both as single mAbs and combinations of two mAbs against non-overlapping epitopes on human CD5. The results demonstrated that combinations of two mAbs significantly increased the level of CDC compared to the single mAbs, while no enhancement of ADCC was seen with anti-CD5 mAb combinations. High levels of CDC and ADCC correlated with low levels of Ab-induced CD5 internalization and degradation. Importantly, an anti-CD5 mAb combination enhanced CDC of CLL cells when combined with the anti-CD20 mAbs rituximab and ofatumumab as well as with the anti-CD52 mAb alemtuzumab. These results suggest that an anti-CD5 mAb combination inducing CDC and ADCC may be effective alone, in combination with mAbs against other targets or combined with chemotherapy for CLL and other CD5-expressing haematological or lymphoid malignancies. © 2013 John Wiley & Sons Ltd.

A variety of human monoclonal antibodies against epidermal growth factor receptor isolated from a phage antibody library.

PubMed

Kurosawa, Gene; Kondo, Mariko; Kurosawa, Yoshikazu

2016-11-04

When the technology for constructing human antibody (Ab) libraries using a phage-display system was developed, many researchers in Ab-related fields anticipated that it would be widely applied to the development of pharmaceutical drugs against various diseases, including cancers. However, successful examples of such applications are very limited. Moreover, researchers who utilize phage-display technology now show divergent ways of thinking about phage Ab libraries. For example, there is debate about what should be the source of V H and V L genes for the construction of libraries to cover the whole repertoire of Abs present in the human body. In the immune system, the introduction of mutations into V genes followed by selection based on binding activity, termed Ab maturation, is required for the production of Abs exhibiting high affinity to the antigen (Ag). Therefore, introduction of mutations and selection are required for isolation of Abs with high affinity after isolation of clones from phage Ab libraries. We constructed a large human Ab library termed AIMS, developed a screening method termed ICOS, and succeeded in isolating many human monoclonal Abs (mAbs) that specifically and strongly bind to various tumor-associated Ags. Eight anti-EGFR mAbs were included, which we characterized. These mAbs showed various different activities against EGFR-expressing cancer cells. In this paper, we describe these data and discuss the possibility and necessity that the mAbs isolated from the AIMS library might be developed as therapeutic drugs against cancers without introduction of mutations. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Efficacy of Genetically Modified Bt Toxins Alone and in Combinations Against Pink Bollworm Resistant to Cry1Ac and Cry2Ab

PubMed Central

Tabashnik, Bruce E.; Fabrick, Jeffrey A.; Unnithan, Gopalan C.; Yelich, Alex J.; Masson, Luke; Zhang, Jie; Bravo, Alejandra; Soberón, Mario

2013-01-01

Evolution of resistance in pests threatens the long-term efficacy of insecticidal proteins from Bacillus thuringiensis (Bt) used in sprays and transgenic crops. Previous work showed that genetically modified Bt toxins Cry1AbMod and Cry1AcMod effectively countered resistance to native Bt toxins Cry1Ab and Cry1Ac in some pests, including pink bollworm (Pectinophora gossypiella). Here we report that Cry1AbMod and Cry1AcMod were also effective against a laboratory-selected strain of pink bollworm resistant to Cry2Ab as well as to Cry1Ab and Cry1Ac. Resistance ratios based on the concentration of toxin killing 50% of larvae for the resistant strain relative to a susceptible strain were 210 for Cry2Ab, 270 for Cry1Ab, and 310 for Cry1Ac, but only 1.6 for Cry1AbMod and 2.1 for Cry1AcMod. To evaluate the interactions among toxins, we tested combinations of Cry1AbMod, Cry1Ac, and Cry2Ab. For both the resistant and susceptible strains, the net results across all concentrations tested showed slight but significant synergism between Cry1AbMod and Cry2Ab, whereas the other combinations of toxins did not show consistent synergism or antagonism. The results suggest that the modified toxins might be useful for controlling populations of pink bollworm resistant to Cry1Ac, Cry2Ab, or both. PMID:24244692

Identification of an IgG CDR sequence contributing to co-purification of the host cell protease cathepsin D.

PubMed

Bee, Jared S; Machiesky, LeeAnn M; Peng, Li; Jusino, Kristin C; Dickson, Matthew; Gill, Jeffrey; Johnson, Douglas; Lin, Hung-Yu; Miller, Kenneth; Heidbrink Thompson, Jenny; Remmele, Richard L

2017-01-01

Recombinant therapeutic monoclonal antibodies (mAbs) must be purified from host cell proteins (HCPs), DNA, and other impurities present in Chinese hamster ovary (CHO) cell culture media. HCPs can potentially result in adverse clinical responses in patients and, in specific cases, have caused degradation of the final mAb product. As reported previously, residual traces of cathepsin D caused particle formation in the final product of mAb-1. The current work was focused on identification of a primary sequence in mAb-1 responsible for the binding and consequent co-purification of trace levels of CHO cathepsin D. Surface plasmon resonance (SPR) was used to detect binding between immobilized CHO cathepsin D and a panel of mAbs. Out of 13 mAbs tested, only mAb-1 and mAb-6 bound to cathepsin D. An LYY motif in the HC CDR2 was common, yet unique, to only these two mAbs. Mutation of LYY to AAA eliminated binding of mAb-1 to cathepsin D providing confirmation that this sequence motif was involved in the binding to CHO cathepsin D. Interestingly, the binding between mAb-1 and cathepsin D was weaker than that of mAb-6, which may be related to the fact that two aspartic acid residues near the LYY motif in mAb-1 are replaced with neutral serine residues in mAb-6. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:140-145, 2017. © 2016 American Institute of Chemical Engineers.

Anti-cancer Activity of Novel TM4SF5-Targeting Antibodies through TM4SF5 Neutralization and Immune Cell-Mediated Cytotoxicity

PubMed Central

Ahn, Hye-Mi; Ryu, Jihye; Song, Jin Myeong; Lee, Yunhee; Kim, Hye-Jin; Ko, Dongjoon; Choi, Inpyo; Kim, Sang Jick; Lee, Jung Weon; Kim, Semi

2017-01-01

The transmembrane four L6 family member 5 (TM4SF5) protein is a novel molecular target for the prevention and treatment of hepatocellular carcinoma. TM4SF5 is highly expressed in liver, colon, esophageal, and pancreatic cancers and is implicated in tumor progression. Here, we screened monoclonal antibodies that specifically bound to the extracellular loop 2 (EC2) of TM4SF5 from a phage-displayed murine antibody (single-chain variable fragment; scFv) library. We constructed and characterized chimeric antibodies, Ab27 and Ab79, of scFv fused with Fc domain of human IgG1. The affinity (KD) of Ab27 and Ab79 for soluble EC2 was approximately 9.2 nM and 16.9 nM, respectively, as determined by surface plasmon resonance analysis. Ab27 and Ab79 efficiently bound to native TM4SF5 on the cell surface were internalized into the cancer cells, leading to a decrease in cell surface TM4SF5. Ab27 and Ab79 inhibited the proliferation and invasion of TM4SF5-positive liver and colon cancer cells and reduced FAK and c-Src phosphorylation. Ab27 and Ab79 also enhanced anoikis sensitivity and reduced survivin. Ab27 mediated antibody-dependent cell-mediated cytotoxicity in vitro. Ab27 and Ab79 efficiently inhibited tumor growth in a liver cancer xenograft model. These results strongly support the further development of Ab27 as a novel anti-cancer agent in the clinic. PMID:28255353

Outcome of limbic encephalitis with VGKC-complex antibodies: relation to antigenic specificity.

PubMed

Malter, M P; Frisch, C; Schoene-Bake, J C; Helmstaedter, C; Wandinger, K P; Stoecker, W; Urbach, H; Surges, R; Elger, C E; Vincent, A V; Bien, C G

2014-09-01

In limbic encephalitis (LE) with antibodies (Abs) to the voltage-gated potassium channel complex (VGKC), the Abs are mainly directed to the VGKC-complex proteins, leucine-rich, glioma inactivated 1 protein (LGI1) or contactin-associated protein-like 2 (CASPR-2) or neither. Here, we relate the outcomes of VGKC-LE patients to the presence of Abs to LGI1, CASPR-2 or neither antigen (LGI1/CASPR-2-Ab(-)). Clinical, neuropsychology and MRI data were obtained from patient records for all LE patients from the Bonn Epilepsy Centre positive for VGKC-Abs by radioimmunoprecipitation assay between 2002 and 2011. Eighteen VGKC-LE patients were identified: nine patients (50 %) had LGI1-Abs, three (16 %) had CASPR-2-Abs; and six (33 %) were negative for both LGI1- and CASPR-2-Abs. At first assessment, the groups did not differ clinically or radiologically, but faciobrachial dystonic seizures were only observed in two LGI1-Ab(+) patients. All patients received monthly intravenous methylprednisolone (MP) pulses. At the most recent follow up (median 26 months), thirteen (72 %) were seizure-free, and seizure-freedom rates did not differ between the Ab groups. Hippocampal atrophy had developed in 7/9 LGI1-Ab(+) patients, but in none of the CASPR-2-Ab(+) or LGI/CASPR-2-Ab(-) patients (p = 0.003). While all subgroups improved, memory scores only normalized in six patients (33 %) and LGI1-Ab(+) patients were left with significantly poorer memory than the other two subgroups. Most VGKC-LE patients become seizure-free with pulsed monthly MP, but memory outcome is less favourable. Hippocampal atrophy and poor memory recovery is common in patients with LGI1-Abs and suggests permanent functional damage. More intense immunotherapies could improve outcomes in LGI1-Ab(+)-LE.

Effect of monoclonal antibodies (MoAb) to class I and class II HLA antigens on lectin- and MoAb OKT3-induced lymphocyte proliferation.

PubMed

Akiyama, Y; Zicht, R; Ferrone, S; Bonnard, G D; Herberman, R B

1985-04-01

We have examined the effect of several monoclonal antibodies (MoAb) to monomorphic determinants of class II HLA antigens, and MoAb to monomorphic determinants of class I HLA antigens and to beta-2-microglobulin (beta 2-mu) on lectin- and MoAb OKT3-induced proliferation of human peripheral blood mononuclear cells (PBMNC) and cultured T cells (CTC). Some, but not all, anti-class II HLA MoAb inhibited the proliferative response of PBMNC to MoAb OKT3 and pokeweed mitogen (PWM). The degree of inhibitory effect varied considerably. This effect was not limited to anti-class II HLA MoAb since anti-class I HLA MoAb and anti-beta 2-mu MoAb also inhibited MoAb OKT3- or PWM-induced proliferative responses. In contrast, the response of PBMNC to phytohemagglutinin (PHA) and concanavalin A (Con A) was not blocked by any anti-class II HLA MoAb. However, some anti-class II HLA MoAb also inhibited the proliferative response of CTC plus allogeneic peripheral blood adherent accessory cells (AC) to PHA or Con A as well as to MoAb OKT3 or PWM. This may be attributable to the substantially greater class II HLA antigen expression by CTC than by fresh lymphocytes. Pretreatment of either CTC or AC with anti-class II HLA MoAb inhibited OKT3-induced proliferation. In contrast, pretreatment of CTC, but not AC, with anti-class I HLA MoAb inhibited the proliferative response of CTC to OKT3. Pretreatment of CTC with anti-class I HLA MoAb inhibited PHA-, Con A and PWM-induced proliferation, to a greater degree than the anti-class II HLA MoAb. It appears as if lymphocyte activation by different mitogens exhibits variable requirements for the presence of cells expressing major histocompatibility determinants. Binding of Ab to membrane markers may interfere with lymphocyte-AC cooperation, perhaps by inhibiting binding of mitogens to their receptors or by interfering with lymphocyte and AC function. We also have examined the role of class II HLA antigens on CTC by depleting class II HLA-positive cells. As expected, elimination of class II HLA-positive AC with anti-class II HLA MoAb plus complement caused a decrease in proliferation of CTC in response to all the mitogens tested. In contrast, elimination of class II HLA-positive CTC was shown to clearly increase proliferation of CTC, perhaps because this may deplete class II HLA-positive suppressor cells.

Monoclonal antibodies against loggerhead sea turtle, Caretta caretta, IgY isoforms reveal differential contributions to antibody titers and relatedness among other sea turtles.

PubMed

Rodgers, Maria L; Rice, Charles D

2018-05-19

Serum from loggerhead sea turtles, Caretta caretta, was collected from the southeast Atlantic Ocean during routine summer monitoring studies in 2017. Serum immunoglobulin IgY was purified and used to develop IgY isoform-specific monoclonal antibodies (mAb). mAb LH12 was developed against the 66 kDa heavy chain of IgY, mAb LH1 was developed against the truncated heavy chain of approximately 37 kDA, and mAb LH9 was developed against the 23 kDa light chains. mAb LH9 reacts with the light chains of all sea turtles, mAb LH12 reacts with the long heavy chain of all sea turtles within the family Cheloniidae, and mAb LH1 reacts with the truncated form of IgY in both olive and Kemp's ridley turtles. Circulating IgY antibodies against three different marine bacterial pathogens were determined in 16 loggerhead samples using these mAbs. mAb LH12 detects higher titers than mAb LH1, and mAb LH9 detects the highest titers. Copyright © 2018 Elsevier Ltd. All rights reserved.

Chronic treatment of (+)-methamphetamine-induced locomotor effects in rats using one or a combination of two high affinity anti-methamphetamine monoclonal antibodies

PubMed Central

Hambuchen, Michael D.; Rüedi-Bettschen, Daniela; Gunnell, Melinda G.; Hendrickson, Howard; Owens, S. Michael

2016-01-01

ABSTRACT We hypothesized that treatment of methamphetamine (METH) effects with a mixture of 2 high affinity anti-METH monoclonal antibodies (mAb) with differing molecular recognition for METH-like structures could increase efficacy compared to treatment with a single mAb. The antibodies studied were mAb7F9 (METH and amphetamine [AMP] KD = 7.7 and 270 nM) and mAb4G9 (16 nM and 110 nM, respectively) in a 50:50 mixture. Adult male Sprague Dawley Rats were treated with iv saline or a loading dose of mAb7F9-mAb4G9 (141 mg/kg of each mAb) followed by 2 weekly doses (70.5 mg/kg total) on days 7 and 14. METH challenge doses (0.56 mg/kg) were administered 4 hrs and 3 days after each mAb7F9-mAb4G9 treatment, and 7 days after the final treatment (day 21). Locomotor activity (0–4 hrs) and serum METH and AMP concentrations (at 5 hrs) were measured after each METH challenge. MAb7F9-mAb4G9 treatment significantly reduced the duration of locomotor activity after 6 of the 7 METH doses (P < 0.05) and significantly increased serum METH and AMP concentrations. Administering three-fold higher METH doses (1.68 mg/kg) on days 24 and 28 showed mAb7F9-mAb4G9 treatment had negligible effects on the duration of METH-induced locomotor activity. These data were then compared to previous monotherapy data. While mAb7F9-mAb4G9 therapy inhibited the effects of multiple METH challenge doses, the inhibition was not as profound or as long lasting as the effects of mAb7F9 treatment alone. These data demonstrate the importance of both mAb affinity and specificity in the production of effective, long-lasting anti-METH mAb therapies. PMID:27163775

In Vitro Study of Bacteriophage AB3 Endolysin LysAB3 Activity Against Acinetobacter baumannii Biofilm and Biofilm-Bound A. baumannii.

PubMed

Zhang, Jie; Xu, Lu-Lu; Gan, Dan; Zhang, Xingping

2018-06-01

The increase in the prevalence of drug-resistant Acinetobacter baumannii is a serious public health concern, which is closely linked to the formation of biofilm. It is reported that the bacteriophage and its endolysin have a good ability to degrade biofilms. The goals of this study were to compare the ability of A. baumannii bacteriophage AB3, its endolysin AB3, and three antibiotics to degrade A. baumannii biofilm and biofilm-bound A. baumannii and to understand the antibacterial mechanism of LysAB3. The 558-bp sequence of the LysAB3 gene was amplified by polymerase chain reaction (PCR); the fragment was cloned into pET28a (+) to construct the recombinant plasmid pET28a-LysAB3, which was then expressed in E. coli BL21 (DE3) to obtain the LysAB3. Differences in A. baumannii biofilm and biofilm-bound A. baumannii after treatment with bacteriophage AB3, LysAB3 or three antibiotics were examined using the crystal violet staining method and an MTT (3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide) assay. Changes in biofilm morphology and thickness in each treatment group were observed by laser scanning confocal microscopy. In addition, a LysAB3 construct with the amphiphilic peptide structural region removed (LysAB3-D) was assessed for its antibacterial activity. After 24-hour treatment with either bacteriophage AB3 and its LysAB3, A. baumannii biofilms were significantly degraded, and the number of viable biofilm-bound A. baumannii were also significantly decreased. After removing the amphiphilic peptide structure motif from LysAB3, the antibacterial activity decreased from 95.8% to 33.3%. Thus, LysAB3 can effectively degrade A. baumannii biofilm and biofilm-bound A. baumannii in vitro. The antibacterial mechanism of LysAB3 may be associated with the ability of the amphiphilic peptide structural region to enhance the permeability of cytoplasmic membrane of A. baumannii by degradation of bacterial wall peptidoglycan.

The new allelic variant of the subtilase cytotoxin (subAB2) is common among Shiga toxin-producing Escherichia coli strains from large game animals and their meat and meat products.

PubMed

Sánchez, Sergio; Díaz-Sánchez, Sandra; Martínez, Remigio; Llorente, María Teresa; Herrera-León, Silvia; Vidal, Dolors

2013-10-25

Subtilase cytotoxin (SubAB) is an AB5 toxin produced by Shiga toxin (Stx)-producing Escherichia coli (STEC) strains usually lacking the eae gene product intimin. Two allelic variants of SubAB encoding genes have been described: subAB1, located on a plasmid, and subAB2, located on a pathogenicity island (PAI) together with tia gene. While subAB1 has been reported to be more frequent among bovine strains, subAB2 has been mainly associated with strains from small ruminants. We investigated the presence of the two variants of subAB among 59 eae-negative STEC from large game animals (deer and wild boar) and their meat and meat products in order to assess the role of other species in the epidemiology of subAB-positive, eae-negative STEC. For this approach, the strains were PCR-screened for the presence of subAB, including the specific detection of both allelic variants, for the presence of saa, tia and sab, and for stx subtyping. Overall, subAB genes were detected in 71.2% of the strains: 84.1% of the strains from deer and 33.3% of the strains from wild boar. Most of them (97.6%) possessed subAB2 and most of these subAB2-positive strains (92.7%) were also positive for tia and negative for saa, suggesting the presence of the subAB2-harbouring PAI. Subtype stx2b was present in most of the strains (67.8%) and a statistically significant association could be established between subAB2 and stx2b. Our results suggest that large game animals, mainly deer, may represent an important animal reservoir of subAB2-positive, eae-negative STEC, and also highlight the risk of human infection posed by the consumption of large game meat and meat products. Copyright © 2013 Elsevier B.V. All rights reserved.

Identification and prevalence of RND family multidrug efflux pump oqxAB genes in Enterococci isolates from swine manure in China.

PubMed

Yuan, Li; Zhai, Ya-Jun; Wu, Hua; Sun, Hua-Run; He, Zhi-Pei; Wang, Ya-Bin; Pan, Yu-Shan; Kuang, Nan-Nan; Hu, Gong-Zheng

2018-06-01

The resistance/nodulation/cell division (RND) family multidrug efflux pump, OqxAB, has been identified as one of the leading mechanisms of plasmid-mediated quinolone resistance and has become increasingly prevalent among Enterobacteriaceae in recent years. However, oqxAB genes have not yet been reported in Enterococcus isolates. The aim of the present study was to identify the oqxAB genes and investigate their prevalence among Enterococcus from swine manure in China. The oqxAB genes were screened in 87 Enterococcus isolates by PCR. The transferability of the oqxAB genes in Enterococcus was determined by conjugation experiments. The genetic environment of oqxAB genes was investigated by cloning experiments, PCR mapping and sequencing. A high prevalence (86.2 %) of olaquindox resistance was observed in Enterococcus and 98.9 % isolates exhibited multidrug-resistance phenotypes. The occurrence of oqxA and oqxB in Enterococcus was also high (79.3 and 65.5 %, respectively). Sequence analysis of the cloned fragment indicated that the oqxAB cassette was linked to an incomplete Tn5 transposon containing aph(3')-IIa and flanked by IS26 [IS26-oqxAB-IS26-aph(3')-IIa]. The oqxAB-aph(3')-IIa-positive transconjugant or transformant showed resistance or reduced susceptibility to enrofloxacin, ciprofloxacin, olaquindox, mequindox, florfenicol, neomycin and kanamycin. This is the first time that the oqxAB genes have been identified in Enterococcus faecalis from swine manure. The genetic linkage of oqxAB-aph(3')-IIa in Enterococcus has not been described before. The high prevalence of oqxAB genes in Enterococcus suggests that it may constitute a reservoir for oqxAB genes and pose a potential threat to public health.

Antibody to ricin a chain hinders intracellular routing of toxin and protects cells even after toxin has been internalized.

PubMed

Song, Kejing; Mize, R Ranney; Marrero, Luis; Corti, Miriam; Kirk, Jason M; Pincus, Seth H

2013-01-01

Mechanisms of antibody-mediated neutralization are of much interest. For plant and bacterial A-B toxins, A chain mediates toxicity and B chain binds target cells. It is generally accepted and taught that antibody (Ab) neutralizes by preventing toxin binding to cells. Yet for some toxins, ricin included, anti-A chain Abs afford greater protection than anti-B. The mechanism(s) whereby Abs to the A chain neutralize toxins are not understood. We use quantitative confocal imaging, neutralization assays, and other techniques to study how anti-A chain Abs function to protect cells. Without Ab, ricin enters cells and penetrates to the endoplasmic reticulum within 15 min. Within 45-60 min, ricin entering and being expelled from cells reaches equilibrium. These results are consistent with previous observations, and support the validity of our novel methodology. The addition of neutralizing Ab causes ricin accumulation at the cell surface, delays internalization, and postpones retrograde transport of ricin. Ab binds ricin for >6hr as they traffic together through the cell. Ab protects cells even when administered hours after exposure. CONCLUSIONS/KEY FINDINGS: We demonstrate the dynamic nature of the interaction between the host cell and toxin, and how Ab can alter the balance in favor of the cell. Ab blocks ricin's entry into cells, hinders its intracellular routing, and can protect even after ricin is present in the target organelle, providing evidence that the major site of neutralization is intracellular. These data add toxins to the list of pathogenic agents that can be neutralized intracellularly and explain the in vivo efficacy of delayed administration of anti-toxin Abs. The results encourage the use of post-exposure passive Ab therapy, and show the importance of the A chain as a target of Abs.

Cytokine/Antibody complexes: an emerging class of immunostimulants.

PubMed

Mostböck, Sven

2009-01-01

In recent years, complexes formed from a cytokine and antibodies against that respective cytokine (cytokine/Ab complex) have been shown to induce remarkable powerful changes in the immune system. Strong interest exists especially for complexes formed with Interleukin (IL)-2 and anti-IL-2-antibody (IL-2/Ab complex). IL-2/Ab complex activates maturation and proliferation in CD8(+) T cells and natural killer (NK) cells to a much higher degree than conventional IL-2 therapy. In addition, IL-2/Ab complex does not stimulate regulatory T cells as much as IL-2 alone. This suggests the possibility to replace the conventional IL-2 therapy with a therapy using low-dose IL-2/Ab complex. Further synthetic cytokine/Ab complexes are studied currently, including IL-3/Ab complex for its effects on the mast cell population, and IL-4/Ab complex and IL-7/Ab complex for inducing B and T cell expansion and maturation. Cytokine complexes can also be made from a cytokine and its soluble receptor. Pre-association of IL-15 with soluble IL-15 receptor alpha produces a complex with strong agonistic functions that lead to an expansion of CD8(+) T cells and NK cells. However, cytokine/Ab complexes also occur naturally in humans. A multitude of auto-antibodies to cytokines are found in human sera, and many of these auto-antibodies build cytokine/Ab complexes. This review presents naturally occurring auto-antibodies to cytokines and cytokine/Ab complexes in health and disease. It further summarizes recent research on synthetic cytokine/Ab complexes with a focus on the basic mechanisms behind the function of cytokine/Ab complexes.

Engineered domain based assays to identify individual antibodies in oligoclonal combinations targeting the same protein

PubMed Central

Meng, Q.; Garcia-Rodriguez, C.; Manzanarez, G.; Silberg, M.A.; Conrad, F.; Bettencourt, J.; Pan, X.; Breece, T.; To, R.; Li, M.; Lee, D.; Thorner, L.; Tomic, M.T.; Marks, J.D.

2014-01-01

Quantitation of individual mAbs within a combined antibody drug product is required for preclinical and clinical drug development. We have developed two antitoxins (XOMA 3B and XOMA 3E) each consisting of three monoclonal antibodies (mAbs) that neutralize type B and type E botulinum neurotoxin (BoNT/B and BoNT/E) to treat serotype B and E botulism. To develop mAb-specific binding assays for each antitoxin, we mapped the epitopes of the six mAbs. Each mAb bound an epitope on either the BoNT light chain (LC) or translocation domain (HN). Epitope mapping data was used to design LC-HN domains with orthogonal mutations to make them specific for only one mAb in either XOMA 3B or 3E. Mutant LC-HN domains were cloned, expressed, and purified from E. coli. Each mAb bound only to its specific domain with affinity comparable to the binding to holotoxin. Further engineering of domains allowed construction of ELISAs that could characterize the integrity, binding affinity, and identity of each of the six mAbs in XOMA 3B, and 3E without interference from the three BoNT/A mAbs in XOMA 3AB. Such antigen engineering is a general method allowing quantitation and characterization of individual mAbs in a mAb cocktail that bind the same protein. PMID:22922799

Studies of guinea pig immunoglobulin isotype, idiotype and antiidiotype

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

Tirrell, S.M.

1988-01-01

Immunization of Guinea pigs with diphtheria toxoid generated antibodies of the IgG class that were capable of neutralizing native toxin in vivo. Sera from these animals were used to affinity purify idiotypic antibodies (AB1). AB1 vaccines derived from the IgG1 class and from F(ab{prime}){sub 2} of IgG1 + IgG2 (IgG1/2) classes were effective in inducing a syngeneic anti-idiotype (AB2) response. Animals immunized with AB1 consisting of both IgG1/2 did not elicit a detectable AB2 response. Binding of homologous {sup 125}I-F(ab{prime}){sub 2} (AB1) to the antiidiotype was inhibited 90% in the presence of DT.F(ab{prime}){sub 2} derived from preimmune serum or hadmore » no inhibitory effects on the idiotype-antiidiotype interactions. Two groups of outbred guinea pigs were vaccinated with alum absorbed F(ab{prime}){sub 2} of anti-idiotype IgG1/2 (AB2). Of the ten animals inoculated with AB2, three tested positive by RIA against {sup 125}I-DT. Two of the RIA positive sera contained antibodies that neutralized diphtheria toxin in a rabbit intracutaneous assay. Purification of guinea pig IgG by protein A-Sepharose affinity chromatography resulted in the separation of three distinct IgG populations.« less

Characterisation of monoclonal antibodies specific for hamster leukocyte differentiation molecules.

PubMed

Rees, Jennifer; Haig, David; Mack, Victoria; Davis, William C

2017-01-01

Flow cytometry was used to identify mAbs that recognize conserved epitopes on hamster leukocyte differentiation molecules (hLDM) and also to characterize mAbs developed against hLDM. Initial screening of mAbs developed against LDMs in other species yielded mAbs specific for the major histocompatibility (MHC) II molecule, CD4 and CD18. Screening of sets of mAbs developed against hLDM yielded 22 new mAbs, including additional mAbs to MHC II molecules and mAbs that recognize LDMs expressed on all leukocytes, granulocytes, all lymphocytes, all T cells, a subset of T cells, or on all B cells. Based on comparison of the pattern of expression of LDMs expressed on all hamster leukocytes with the patterns of expression of known LDMs in other species, as detected by flow cytometry (FC), four mAbs are predicted to recognize CD11a, CD44, and CD45. Cross comparison of mAbs specific for a subset of hamster T cells with a cross reactive mAb known to recognize CD4 in mice and one recognising CD8 revealed they recognize CD4. The characterization of these mAbs expands opportunities to use hamsters as an additional model species to investigate the mechanisms of immunopathogenesis of infectious diseases. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Development of a mouse-feline chimeric antibody against feline tumor necrosis factor-alpha.

PubMed

Doki, Tomoyoshi; Takano, Tomomi; Hohdatsu, Tsutomu

2016-10-01

Feline infectious peritonitis (FIP) is a fatal inflammatory disease caused by FIP virus infection. Feline tumor necrosis factor (fTNF)-alpha is closely involved in the aggravation of FIP pathology. We previously described the preparation of neutralizing mouse anti-fTNF-alpha monoclonal antibody (mAb 2-4) and clarified its role in the clinical condition of cats with FIP using in vitro systems. However, administration of mouse mAb 2-4 to cat may lead to a production of feline anti-mouse antibodies. In the present study, we prepared a mouse-feline chimeric mAb (chimeric mAb 2-4) by fusing the variable region of mouse mAb 2-4 to the constant region of feline antibody. The chimeric mAb 2-4 was confirmed to have fTNF-alpha neutralization activity. Purified mouse mAb 2-4 and chimeric mAb 2-4 were repeatedly administered to cats, and the changes in the ability to induce feline anti-mouse antibody response were investigated. In the serum of cats treated with mouse mAb 2-4, feline anti-mouse antibody production was induced, and the fTNF-alpha neutralization effect of mouse mAb 2-4 was reduced. In contrast, in cats treated with chimeric mAb 2-4, the feline anti-mouse antibody response was decreased compared to that of mouse mAb 2-4-treated cats.

40 CFR Table 6 to Subpart Jjj of... - Known Organic HAP Emitted From the Production of Thermoplastic Products

Code of Federal Regulations, 2013 CFR

2013-07-01

...-crylate (80-62-6) Styrene (100-42-5) ABS latex ✔ ✔ ✔ ABS using a batch emulsion process ✔ ✔ ✔ ABS using a batch suspension process ✔ ✔ ✔ ABS using a continuous emulsion process ✔ ✔ ✔ ABS using a continuous mass...

40 CFR Table 6 to Subpart Jjj of... - Known Organic HAP Emitted From the Production of Thermoplastic Products

Code of Federal Regulations, 2014 CFR

2014-07-01

...-crylate (80-62-6) Styrene (100-42-5) ABS latex ✔ ✔ ✔ ABS using a batch emulsion process ✔ ✔ ✔ ABS using a batch suspension process ✔ ✔ ✔ ABS using a continuous emulsion process ✔ ✔ ✔ ABS using a continuous mass...

DNA degradation in genetically modified rice with Cry1Ab by food processing methods: implications for the quantification of genetically modified organisms.

PubMed

Xing, Fuguo; Zhang, Wei; Selvaraj, Jonathan Nimal; Liu, Yang

2015-05-01

Food processing methods contribute to DNA degradation, thereby affecting genetically modified organism detection and quantification. This study evaluated the effect of food processing methods on the relative transgenic content of genetically modified rice with Cry1Ab. In steamed rice and rice noodles, the levels of Cry1Ab were ⩾ 100% and <83%, respectively. Frying and baking in rice crackers contributed to a reduction in Pubi and Cry1Ab, while microwaving caused a decrease in Pubi and an increase in Cry1Ab. The processing methods of sweet rice wine had the most severe degradation effects on Pubi and Cry1Ab. In steamed rice and rice noodles, Cry1Ab was the most stable, followed by SPS and Pubi. However, in rice crackers and sweet rice wine, SPS was the most stable, followed by Cry1Ab and Pubi. Therefore, Cry1Ab is a better representative of transgenic components than is Pubi because the levels of Cry1Ab were less affected compared to Pubi. Copyright © 2014 Elsevier Ltd. All rights reserved.

Comparison of In Vitro Activity of Liposomal Nystatin against Aspergillus Species with Those of Nystatin, Amphotericin B (AB) Deoxycholate, AB Colloidal Dispersion, Liposomal AB, AB Lipid Complex, and Itraconazole

PubMed Central

Oakley, Karen L.; Moore, Caroline B.; Denning, David W.

1999-01-01

We compared the in vitro activity of liposomal nystatin (Nyotran) with those of other antifungal agents against 60 Aspergillus isolates. Twelve isolates were itraconazole resistant. For all isolates, geometric mean (GM) MICs (micrograms per milliliter) were 2.30 for liposomal nystatin, 0.58 for itraconazole, 0.86 for amphotericin B (AB) deoxycholate, 9.51 for nystatin, 2.07 for liposomal AB, 2.57 for AB lipid complex, and 0.86 for AB colloidal dispersion. Aspergillus terreus (GM, 8.72 μg/ml; range, 8 to 16 μg/ml) was significantly less susceptible to all of the polyene drugs than all other species (P = 0.0001). PMID:10223948

A novel bicistronic gene design couples stable cell line selection with a fucose switch in a designer CHO host to produce native and afucosylated glycoform antibodies.

PubMed

Roy, Gargi; Martin, Tom; Barnes, Arnita; Wang, Jihong; Jimenez, Rod Brian; Rice, Megan; Li, Lina; Feng, Hui; Zhang, Shu; Chaerkady, Raghothama; Wu, Herren; Marelli, Marcello; Hatton, Diane; Zhu, Jie; Bowen, Michael A

2018-04-01

The conserved glycosylation site Asn 297 of a monoclonal antibody (mAb) can be decorated with a variety of sugars that can alter mAb pharmacokinetics and recruitment of effector proteins. Antibodies lacking the core fucose at Asn 297 (afucosylated mAbs) show enhanced antibody-dependent cell-mediated cytotoxicity (ADCC) and increased efficacy. Here, we describe the development of a robust platform for the manufacture of afucosylated therapeutic mAbs by engineering a Chinese hamster ovary (CHO) host cell line to co-express a mAb with GDP-6-deoxy-D-lyxo-4-hexulose reductase (RMD), a prokaryotic enzyme that deflects an intermediate in the de novo synthesis of fucose to a dead-end product, resulting in the production of afucosylated mAb (GlymaxX™ Technology, ProBioGen). Expression of the mAb and RMD genes was coordinated by co-transfection of separate mAb and RMD vectors or use of an internal ribosome entry site (IRES) element to link the translation of RMD with either the glutamine synthase selection marker or the mAb light chain. The GS-IRES-RMD vector format was more suitable for the rapid generation of high yielding cell lines, secreting afucosylated mAb with titers exceeding 6.0 g/L. These cell lines maintained production of afucosylated mAb over 60 generations, ensuring their suitability for use in large-scale manufacturing. The afucosylated mAbs purified from these RMD-engineered cell lines showed increased binding in a CD16 cellular assay, demonstrating enhancement of ADCC compared to fucosylated control mAb. Furthermore, the afucosylation in these mAbs could be controlled by simple addition of L-fucose in the culture medium, thereby allowing the use of a single cell line for production of the same mAb in fucosylated and afucosylated formats for multiple therapeutic indications.

Asbestos bodies in bronchoalveolar lavage in the 21st century: a time-trend analysis in a clinical population.

PubMed

Nuyts, Valerie; Vanhooren, Hadewijch; Begyn, Sarah; Nackaerts, Kristiaan; Nemery, Benoit

2017-01-01

Asbestos bodies (AB) in bronchoalveolar lavage (BAL) can be detected by light microscopy and their concentration is indicative of past cumulative asbestos exposure. We assessed clinical and exposure characteristics, as well as possible time trends, among patients in whom AB had been quantified in BAL. BAL samples obtained from 578 participants between January 1997 and December 2014 were available for analysis. The processing of samples and the microscopic analysis were performed by a single expert and 76% of samples came from a single tertiary care hospital, allowing clinical and exposure data to be extracted from patient files. The study population (95% males) had a mean age of 62.5 (±12.4) years. AB were detected in 55.2% of the samples, giving a median concentration of 0.5 AB/mL (95th centile: 23.6 AB/mL; highest value: 164.5 AB/mL). The AB concentration exceeded 1 AB/mL in 39.4% and 5 AB/mL in 17.8%. A significant decrease from a geometric mean of 0.93 AB/mL in 1997 to 0.2 AB/mL in 2014 was apparent. High AB concentrations generally corresponded with occupations with (presumed) high asbestos exposure. AB concentrations were higher among patients with asbestosis and pleural plaques, when compared with other disease groups. Nevertheless, a substantial proportion of participants with likely exposure to asbestos did not exhibit high AB counts. This retrospective study of a large clinical population supports the value of counting AB in BAL as a complementary approach to assess past exposure to asbestos. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Impact of SPR biosensor assay configuration on antibody: Neonatal Fc receptor binding data

PubMed Central

Wang, Xiangdan; McKay, Patrick; Dutina, George; Hass, Philip E.; Nijem, Ihsan; Allison, David; Cowan, Kyra J.; Lin, Kevin; Quarmby, Valerie; Yang, Jihong

2017-01-01

ABSTRACT Binding interactions with the neonatal Fc receptor (FcRn) are one determinant of pharmacokinetic properties of recombinant human monoclonal antibody (rhumAb) therapeutics, and a conserved binding motif in the crystallizable fragment (Fc) region of IgG molecules interacts with FcRn. Surface plasmon resonance (SPR) biosensor assays are often used to characterize interactions between FcRn and rhumAb therapeutics. In such assays, generally either the rhumAb (format 1) or the FcRn protein (format 2) is immobilized on a biosensor chip. However, because evidence suggests that, in some cases, the variable domains of a rhumAb may also affect FcRn binding, we evaluated the effect of SPR assay configuration on binding data. We sought to assess FcRn binding properties of 2 rhumAbs (rhumAb1 and rhumAb2) to FcRn proteins using these 2 biosensor assay formats. The two rhumAbs have greater than 99% sequence identity in the Fc domain but differ in their Fab regions. rhumAb2 contains a positively charged patch in the variable domain that is absent in rhumAb1. Our results showed that binding of rhumAb1 to FcRn was independent of biosensor assay configuration, while binding of rhumAb2 to FcRn was highly SPR assay configuration dependent. Further investigations revealed that the format dependency of rhumAb2-FcRn binding is linked to the basic residues that form a positively charged patch in the variable domain of rhumAb2. Our work highlights the importance of analyzing rhumAb-FcRn binding interactions using 2 alternate SPR biosensor assay configurations. This approach may also provide a simple way to identify the potential for non-Fc-driven FcRn binding interactions in otherwise typical IgGs. PMID:28001487

A new type of natural bispecific antibody with potential protective effect in Hashimoto thyroiditis.

PubMed

Li, Wenli; Fan, Gaowei; Chen, Lida; Zhang, Rui; Zhang, Kuo; Sun, Yu; Lin, Guigao; Xie, Jiehong; Wang, Lunan; Li, Jinming

2014-09-01

As a new antibody concept, natural bispecific antibodies (nBsAbs) have been detected in long-term passive immunization and some diseases, but their potential immunomodulatory role remains unclear. Hashimoto thyroiditis (HT) appears to fulfill the condition for nBsAb production but has not yet been characterized. The objective of the study was to identify a new nBsAb against thyroid peroxidase (TPO) and thyroglobulin (Tg) in HT patients and to preliminarily explore its immunomodulatory role. Serum samples were obtained from 136 HT patients, 92 diseased controls, and 99 healthy controls for anti-TPO/Tg nBsAb detection. The relationship between anti-TPO/Tg nBsAb and other clinical parameters was also analyzed. The anti-TPO/Tg nBsAb was detected using a double-antigen sandwich ELISA. Higher nBsAb levels were found to be associated with decreased inflammation in HT patients. The prevalence of anti-TPO/Tg nBsAb in HT was 44.9% (61 of 136), significantly higher than that of diseased controls (2.2%, 2 of 92) (P < .0001) and healthy controls (0%, 0 of 99) (P < .0001). HT patients who were nBsAb positive were prone to have significantly lower levels of serum C-reactive protein and TNF-α compared with the nBsAb-negative individuals (P < .05). The serum amyloid A and interferon-γ levels also showed a similar trend in the two groups. The IgG subclass of anti-TPO/Tg nBsAb was IgG4. Further analysis showed a negative correlation between anti-TPO/Tg nBsAb and serum total IgG4 (r = -0.697, P = .025) in IgG4 thyroiditis patients. A new type of nBsAb against TPO and Tg in HT patients is identified. Our data also indicate a protective effect of anti-TPO/Tg nBsAb in the pathogenesis of HT and extend prior knowledge about nBsAb in diseases.

EIN3-like gene expression during fruit ripening of Cavendish banana (Musa acuminata cv. Grande naine).

PubMed

Mbéguié-A-Mbéguié, Didier; Hubert, Olivier; Fils-Lycaon, Bernard; Chillet, Marc; Baurens, Franc-Christophe

2008-06-01

Ethylene signal transduction initiates with ethylene binding at receptor proteins and terminates in a transcription cascade involving the EIN3/EIL transcription factors. Here, we have isolated four cDNAs homologs of the Arabidopsis EIN3/EIN3-like gene, MA-EILs (Musa acuminata ethylene insensitive 3-like) from banana fruit. Sequence comparison with other banana EIL gene already registered in the database led us to conclude that, at this day, at least five different genes namely MA-EIL1, MA-EIL2/AB266318, MA-EIL3/AB266319, MA-EIL4/AB266320 and AB266321 exist in banana. Phylogenetic analyses included all banana EIL genes within a same cluster consisting of rice OsEILs, a monocotyledonous plant as banana. However, MA-EIL1, MA-EIL2/AB266318, MA-EIL4/AB266320 and AB266321 on one side, and MA-EIL3/AB266319 on the other side, belong to two distant subclusters. MA-EIL mRNAs were detected in all examined banana tissues but at lower level in peel than in pulp. According to tissues, MA-EIL genes were differentially regulated by ripening and ethylene in mature green fruit and wounding in old and young leaves. MA-EIL2/AB266318 was the unique ripening- and ethylene-induced gene; MA-EIL1, MA-EIL4/Ab266320 and AB266321 genes were downregulated, while MA-EIL3/AB266319 presented an unusual pattern of expression. Interestingly, a marked change was observed mainly in MA-EIL1 and MA-EIL3/Ab266319 mRNA accumulation concomitantly with changes in ethylene responsiveness of fruit. Upon wounding, the main effect was observed in MA-EIL4/AB266320 and AB266321 mRNA levels, which presented a markedly increase in both young and old leaves, respectively. Data presented in this study suggest the importance of a transcriptionally step control in the regulation of EIL genes during banana fruit ripening.

Molecular identification and functional analysis of two variants of myeloid differentiation factor 88 (MyD88) from disk abalone (Haliotis discus discus).

PubMed

Priyathilaka, Thanthrige Thiunuwan; Bathige, S D N K; Lee, Seongdo; Lee, Jehee

2018-02-01

Myeloid differentiation factor 88 (MyD88) is a crucial adaptor protein of the Toll-like receptor (TLR)- and interleukin 1 receptor-mediated signaling pathways and is involved in a diverse array of inflammatory responses via NF-κB activation. In the present study, two MyD88 variants were identified from disk abalone (Haliotis discus discus) and designated AbMyD88-2 and AbMyD88-X. The deduced AbMyD88-2 and AbMyD88-X comprised 433 and 354 amino acids with predicted molecular masses of 48.85 kDa and 40.17 kDa, respectively. AbMyD88-2 and AbMyD88-X possessed typical MyD88 domain structural features including an N-terminal death domain (DD) and C-terminal toll interleukin 1 receptor (TIR) domain similar to those in mammals. Expression analysis of AbMyD88-2 and AbMyD88-X mRNA at different early embryonic developmental stages of abalone by qPCR revealed that their constitutive expression at all developmental stages analyzed with the considerably higher values at the 16-cell (AbMyD88-2) and morula stages (AbMyD88-X). In unchallenged disk abalones, AbMyD88-2 was highly expressed in muscles, while AbMyD88-X mRNA was predominantly transcribed in hemocytes. Moreover, AbMyD88-2 and AbMyD88-X mRNA were differentially modulated in abalone hemocytes after a challenge with live bacteria (Vibrio parahaemolyticus, Listeria monocytogenes), virus (viral hemorrhagic septicemia virus), and pathogen-associated molecular patterns (lipopolysaccharides and Poly I:C). Overexpression of AbMyD88-2 and AbMyD88-X in HEK293T cells induced the activation of the NF-κB promoter. AbMyD88-2 and AbMyD88-X involvement in inflammatory responses was characterized by their overexpression in RAW264.7 murine macrophage cells. These results revealed comparatively higher NO (Nitric oxide) production, induction of inflammatory mediator genes (iNOS and COX2), and proinflammatory genes (IL1β, IL6 and TNFα) expression in abalone MyD88s-overexpressing cells than in mock control in the presence or absence of LPS stimulation. Altogether, these results suggest that existence of a MyD88-dependent like signaling pathway in disk abalone and that both AbMyD88-2 and AbMyD88-X might be involved in innate immune and inflammatory responses. Copyright © 2017 Elsevier Ltd. All rights reserved.

Automated Discovery of Simulation Between Programs

DTIC Science & Technology

2014-10-18

relation. These relations enable the refinement-step of SimAbs. We have implemented SimAbs using UFO framework and Z3 SMT-solver and applied it to...step of SimAbs. We implemented SimAbs and AE-VAL on the top of the UFO framework [1, 15] and an SMT-solver Z3 [8], respectively. We have evaluated SimAbs...ut 6 Evaluation We have implemented SimAbs in the UFO framework, and evaluated it on the Software Verification Competition (SVCOMP’14) benchmarks and

Estimation of true serum thyroglobulin concentration using simultaneous measurement of serum antithyroglobulin antibody.

PubMed

Ahn, Byeong-Cheol; Lee, Won Kee; Jeong, Shin Young; Lee, Sang-Woo; Lee, Jaetae

2013-01-01

We investigated the analytical interference of antithyroglobulin antibody (TgAb) to thyroglobulin (Tg) measurement and tried to convert measured Tg concentration to true Tg concentration using a mathematical equation which includes a concentration of TgAb. Methods. Tg was measured by immunoradiometric assay and TgAb by radioimmunoassy. Experimental samples were produced by mixing Tg and TgAb standard solutions or mixing patients' serum with high Tg or high TgAb. Mathematical equations for prediction of expected Tg concentration with measured Tg and TgAb concentrations were deduced. The Tg concentration calculated using the equations was compared with the expected Tg concentration. Results. Measured Tg concentrations of samples having high TgAb were significantly lower than their expected Tg concentration. Magnitude of TgAb interference with the Tg assay showed a positive correlation with concentration of TgAb. Mathematical equations for estimation of expected Tg concentration using measured Tg and TgAb concentrations were successfully deduced and the calculated Tg concentration showed excellent correlation with expected Tg concentration. Conclusions. A mathematic equation for estimation of true Tg concentration using measured Tg and TgAb concentration was deduced. Tg concentration calculated by use of the equation might be more valuable than measured Tg concentration in patients with differentiated thyroid cancer.

Molecular identification of disk abalone (Haliotis discus discus) tetraspanin 33 and CD63: Insights into potent players in the disk abalone host defense system.

PubMed

Priyathilaka, Thanthrige Thiunuwan; Bathige, S D N K; Herath, H M L P B; Lee, Sukkyoung; Lee, Jehee

2017-10-01

Tetraspanins are a superfamily of transmembrane proteins involved in a diverse range of physiological processes including differentiation, adhesion, signal transduction, cell motility, and immune responses. In the present study, two tetraspanins, CD63 and tetraspanin 33 (TSPAN33) from disk abalone (AbCD63 and AbTSPAN33), were identified and characterized at the molecular level. The coding sequences for AbCD63 and AbTSPAN33 encoded polypeptides of 234 and 290 amino acids (aa) with predicted molecular mass of 25.3 and 32.5 kDa, respectively. The deduced AbCD63 and AbTSPAN33 protein sequences were also predicted to have a typical tetraspanin domain architecture, including four transmembrane domains (TM), short N- and C- terminal regions, a short intracellular loop, as well as a large and small extracellular loop. A characteristic CCG motif and cysteine residues, which are highly conserved across CD63 and TSPAN33 proteins of different species, were present in the large extracellular loop of both abalone tetraspanins. Phylogenetic analysis revealed that the AbCD63 and AbTSPAN33 clustered in the invertebrate subclade of tetraspanins, thus exhibiting a close relationship with tetraspanins of other mollusks. The AbCD63 and AbTSPAN33 mRNA transcripts were detected at early embryonic development stages of disk abalone with significantly higher amounts at the trochophore stage, suggesting the involvement of these proteins in embryonic development. Both AbCD63 and AbTSPAN33 were ubiquitously expressed in all the tissues of unchallenged abalones analyzed, with the highest expression levels found in hemocytes. Moreover, significant induction of AbCD63 and AbTSPAN33 mRNA expression was observed in immunologically important tissues, such as hemocytes and gills, upon stimulation with live bacteria (Vibrio parahaemolyticus and Listeria monocytogenes), virus (viral hemorrhagic septicemia virus), and two potent immune stimulators [polyinosinic:polycytidylic acid (poly I:C) and lipopolysaccharide (LPS)]. Collectively, these findings suggest that AbCD63 and AbTSPAN33 are involved in innate immune responses in disk abalone during pathogenic stress. Copyright © 2017 Elsevier Ltd. All rights reserved.

Thyroglobulin autoantibodies switch to immunoglobulin (Ig)G1 and IgG3 subclasses and preserve their restricted epitope pattern after 131I treatment for Graves' hyperthyroidism: the activity of autoimmune disease influences subclass distribution but not epitope pattern of autoantibodies

PubMed Central

Latrofa, F; Ricci, D; Montanelli, L; Piaggi, P; Mazzi, B; Bianchi, F; Brozzi, F; Santini, P; Fiore, E; Marinò, M; Tonacchera, M; Vitti, P

2014-01-01

The subclass distribution of thyroglobulin autoantibodies (TgAb) is debated, whereas their epitope pattern is restricted. Radioidine (131I) treatment for Graves' disease (GD) induces a rise in TgAb levels, but it is unknown whether it modifies subclass distribution and epitope pattern of TgAb as well. We collected sera from GD patients before 131I treatment and 3 and 6 months thereafter. We measured total TgAb, TgAb light chains and TgAb subclasses by enzyme-linked immunosorbent assay (ELISA) in 25 patients. We characterized the TgAb epitope pattern in 30 patients by inhibiting their binding to 125-ITg by a pool of four TgAb-Fab (recognizing Tg epitope regions A, B, C and D) and to Tg in ELISA by each TgAb-Fab. Total TgAb immunoglobulin (Ig)G rose significantly (P = 0·024). TgAb κ chains did not change (P = 0·052), whereas TgAb λ chains increased significantly (P = 0·001) and persistently. We observed a significant rise in IgG1 and IgG3 levels after 131I (P = 0·008 and P = 0·006, respectively), while IgG2 and IgG4 levels did not change. The rise of IgG1 was persistent, that of IgG3 transient. The levels of inhibition of TgAb binding to Tg by the TgAb-Fab pool were comparable. A slight, non-significant reduction of the inhibition by the immune-dominant TgAb-Fab A was observed 3 and 6 months after 131I. We conclude that 131I treatment for GD increases the levels of the complement-activating IgG1 and IgG3 subclasses and does not influence significantly the epitope pattern of TgAb. In autoimmune thyroid disease subclass distribution of autoantibodies is dynamic in spite of a stable epitope pattern. PMID:25134846

40 CFR Table 6 to Subpart Jjj of... - Known Organic HAP Emitted From the Production of Thermoplastic Products

Code of Federal Regulations, 2010 CFR

2010-07-01

...) Styrene (100-42-5) ABS latex ✔ ✔ ✔ ABS using a batch emulsion process ✔ ✔ ✔ ABS using a batch suspension process ✔ ✔ ✔ ABS using a continuous emulsion process ✔ ✔ ✔ ABS using a continuous mass process ✔ ✔ ✔ ASA...

40 CFR Table 6 to Subpart Jjj of... - Known Organic HAP Emitted From the Production of Thermoplastic Products

Code of Federal Regulations, 2012 CFR

2012-07-01

...) Styrene (100-42-5) ABS latex ✔ ✔ ✔ ABS using a batch emulsion process ✔ ✔ ✔ ABS using a batch suspension process ✔ ✔ ✔ ABS using a continuous emulsion process ✔ ✔ ✔ ABS using a continuous mass process ✔ ✔ ✔ ASA...

40 CFR Table 6 to Subpart Jjj of... - Known Organic HAP Emitted From the Production of Thermoplastic Products

Code of Federal Regulations, 2011 CFR

2011-07-01

...) Styrene (100-42-5) ABS latex ✔ ✔ ✔ ABS using a batch emulsion process ✔ ✔ ✔ ABS using a batch suspension process ✔ ✔ ✔ ABS using a continuous emulsion process ✔ ✔ ✔ ABS using a continuous mass process ✔ ✔ ✔ ASA...

Undoing Gender Through Legislation and Schooling: the Case of AB 537 and AB 394 IN California, USA

NASA Astrophysics Data System (ADS)

Knotts, Greg

2009-11-01

This article investigates California laws AB 537: The Student Safety and Violence Prevention Act of 2000, and the recently enacted AB 394: Safe Place to Learn Act. Both demand that gender identity and sexual orientation be added to the lexicon of anti-harassment protection in public education. However, despite these progressive measures, schools have an unconscious acceptance of heteronormativity and gendered norms, which undermines both the spirit and language of these laws. This paper examines how California schools can both change standard practices and realise the transformative social change that laws like AB 537 and AB 394 can instigate. I assert that the systemic implementation of these laws, through the adoption, enforcement and evaluation of existing AB 537 Task Force Recommendations, is necessary for their success. My second assertion is that AB 537 and AB 394 have the potential to change and reconstitute gender-based and heteronormative standards at school sites.

Disease-enhancing antibodies improve the efficacy of bacterial toxin-neutralizing antibodies

PubMed Central

Chow, Siu-Kei; Smith, Cameron; MacCarthy, Thomas; Pohl, Mary Ann; Bergman, Aviv; Casadevall, Arturo

2013-01-01

SUMMARY During infection, humoral immunity produces a polyclonal response with various immunoglobulins recognizing different epitopes within the microbe or toxin. Despite this diverse response, the biological activity of an antibody (Ab) is usually assessed by the action of a monoclonal population. We demonstrate that a combination of monoclonal antibodies (mAbs) that are individually disease-enhancing or neutralizing to Bacillus anthracis protective antigen (PA), a component of anthrax toxin, results in significantly augmented protection against the toxin. This boosted protection is Fc gamma receptor (FcγR)-dependent and involves the formation of stoichiometrically defined mAb-PA complexes that requires immunoglobulin bivalence and simultaneous interaction between PA and the two mAbs. The formation of these mAb-PA complexes inhibits PA oligomerization, resulting in protection. These data suggest that functional assessments of single Abs may inaccurately predict how the same Abs will operate in polyclonal preparations and imply that potentially therapeutic mAbs may be overlooked in single Ab screens. PMID:23601104

Therapeutic monoclonal antibodies and the need for targeted pharmacovigilance in India

PubMed Central

Kalaivani, M; Singh, Abhishank; Kalaiselvan, V

2015-01-01

A growing number of innovative mAb therapeutics are on the global market, and biosimilar versions have now also been approved, including in India. Although efficacy and safety is demonstrated prior to approval, targeted pharmacovigilance is essential for the identification and assessment of risk for any mAb products. We analyzed the ADR data related to mAbs reported to the NCC-PvPI through the spontaneous reporting system Vigiflow during April 2011 to February 2014 to identify mAbs with the highest number of ADR including fatal/serious ADR. Only 0.72% reports were related to mAbs. Although 15 mAbs are approved in the country, only 6 mAbs were reported through Vigiflow. Rituximab was highly reported, and no fatal/serious ADR related to any mAbs were reported during the study period. Our study shows that PvPI is effective and robust system in the detection and assessment of risks associated with the use of mAbs. PMID:25523367

Antibacterial effects of fibrin glue-antibiotic mixtures.

PubMed

Kram, H B; Bansal, M; Timberlake, O; Shoemaker, W C

1991-02-01

In the present in vitro study, we investigated the duration of action and antibacterial effects of nonautologous fibrin glue (FG) combined with antibiotics (Abs) including Ciprofloxacin, Teicoplanin, Cefoxitin, and Gentamicin; the effect of FG alone on bacterial growth was also evaluated. The rate of Ab diffusion from combined FG-Ab clots was evaluated by separate elution with pooled human serum (HS) and normal saline (NS); supernatants were removed daily and assayed for active concentrations of each Ab. The effects of FG and combined FG-Ab clots on bacterial growth were evaluated by inoculating brain-heart infusion (BHI) with Staphylococcus aureus, followed by the addition of FG or FG-Ab clots; a separate set of studies was also performed with the addition of mouse lung homogenate (MLH) as well. The addition of Ab to FG clots resulted in continuous diffusion of the Ab into the surrounding HS or NS for up to 5 to 7 days; however, more than two-thirds of the Ab diffused out within 2 to 3 days regardless of the Ab used. The antibacterial effects of FG-Ciprofloxacin and FG-Teicoplanin clots were significant compared to those of FG clots not containing Ab. In addition, the presence of FG clots (in BHI) resulted in a reduction in bacterial growth compared to that of BHI alone. The addition of MLH to BHI resulted in increased bacterial growth, but this effect was inhibited by the presence of combined FG-Ab clots.(ABSTRACT TRUNCATED AT 250 WORDS)

Engineered domain-based assays to identify individual antibodies in oligoclonal combinations targeting the same protein.

PubMed

Meng, Q; Garcia-Rodriguez, C; Manzanarez, G; Silberg, M A; Conrad, F; Bettencourt, J; Pan, X; Breece, T; To, R; Li, M; Lee, D; Thorner, L; Tomic, M T; Marks, J D

2012-11-15

Quantitation of individual monoclonal antibodies (mAbs) within a combined antibody drug product is required for preclinical and clinical drug development. We have developed two antitoxins, XOMA 3B and XOMA 3E, each consisting of three mAbs that neutralize type B and type E botulinum neurotoxin (BoNT/B and BoNT/E) to treat serotype B and E botulism. To develop mAb-specific binding assays for each antitoxin, we mapped the epitopes of the six mAbs. Each mAb bound an epitope on either the BoNT light chain (LC) or translocation domain (H(N)). Epitope mapping data were used to design LC-H(N) domains with orthogonal mutations to make them specific for only one mAb in either XOMA 3B or XOMA 3E. Mutant LC-H(N) domains were cloned, expressed, and purified from Escherichia coli. Each mAb bound only to its specific domain with affinity comparable to the binding to holotoxin. Further engineering of domains allowed construction of enzyme-linked immunosorbent assays (ELISAs) that could characterize the integrity, binding affinity, and identity of each of the six mAbs in XOMA 3B and 3E without interference from the three BoNT/A mAbs in XOMA 3AB. Such antigen engineering is a general method allowing quantitation and characterization of individual mAbs in a mAb cocktail that bind the same protein. Copyright © 2012 Elsevier Inc. All rights reserved.

Influence of Immunogenicity on the Efficacy of Long-Term Treatment with TNFα Blockers in Rheumatoid Arthritis and Spondyloarthritis Patients

PubMed Central

2015-01-01

Objective. To analyze the clinical relevance of the levels of TNFα blockers and anti-drug antibodies (anti-drug Ab) in patients with rheumatoid arthritis (RA) and spondyloarthritis (SpA) treated with adalimumab (ADA), etanercept (ETA), or infliximab (INF) for a prolonged period of time. Methods. Clinical characteristics (disease activity, and adverse events), serum TNFα blockers, and anti-drug Ab levels were evaluated in 62 RA and 81 SpA patients treated with TNFα blockers for a median of 28 months. Results. Anti-ADA Ab were detected in 1 (4.0%) and anti-INF Ab in 14 out of 57 (24.6%) RA and SpA patients. Patient with anti-ADA Ab and 57.1% patients with anti-INF Ab were considered nonresponders to treatment. Anti-ETA Ab were not found in any of 61 ETA treated patients. Anti-ADA and anti-INF Ab levels differ between responders and nonresponders (P > 0.05). Three (5.3%) patients with high serum anti-INF Ab levels developed infusion related reactions. Patients with anti-INF Ab more often required changing to another biologic drug (OR 11.43 (95% CI 1.08–120.93)) and treatment discontinuation (OR 9.28 (95% CI 1.64–52.52)). Conclusion. Patients not responding to treatment had higher serum anti-ADA and anti-INF Ab concentrations. Anti-INF Ab formation is related to increased risk of infusion related reactions, changing to another biologic drug, and treatment discontinuation. PMID:26064930

Immunologic advances in monoclonal antibody therapy: implications for oncology nursing.

PubMed

Karius, D; Marriott, M A

1997-04-01

To provide an overview of monoclonal antibody (MoAb) formation, therapeutic and diagnostic uses of MoAbs, and the implications for oncology nurses. Books and Journal articles (including research studies). Clinical trials have demonstrated the diagnostic and therapeutic potential of MoAb therapy. Advances in hybridoma technology and gene-splicing techniques have led to the formation of chimeric MoAbs, which exhibit decreased immunogenicity in the recipient. Clinical limitations with MoAb therapy include cross-reactivity with normal tissues, heterogeneity of antigen expression, presence of circulating antigen, antigenic modulation, tumor size and vascularity, and the anti-antibody response. MoAbs currently are used for diagnostic purposes and in phase I, II, and III clinical trials for cancer treatment. As research progresses, MoAbs are likely to be incorporated into the mainstream of cancer therapy as have other biologic response modifiers. Current uses of MoAb therapy in clinical trials involve nurses in many roles, including clinical nurse specialist, staff nurse, and research nurse. As more oncology nurses encounter MoAb therapy in practice, they will have to have an increased understanding of basic immunologic principles and the expertise to manage the unique toxicities associated with MoAb therapy.

Dynamics of attentional bias to threat in anxious adults: bias towards and/or away?

PubMed

Zvielli, Ariel; Bernstein, Amit; Koster, Ernst H W

2014-01-01

The aim of the present study was to question untested assumptions about the nature of the expression of Attentional Bias (AB) towards and away from threat stimuli. We tested the idea that high trait anxious individuals (N = 106; M(SD)age = 23.9(3.2) years; 68% women) show a stable AB towards multiple categories of threatening information using the emotional visual dot probe task. AB with respect to five categories of threat stimuli (i.e., angry faces, attacking dogs, attacking snakes, pointed weapons, violent scenes) was evaluated. In contrast with current theories, we found that 34% of participants expressed AB towards threat stimuli, 20.8% AB away from threat stimuli, and 34% AB towards some categories of threat stimuli and away from others. The multiple observed expressions of AB were not an artifact of a specific criterion AB score cut-off; not specific to certain categories of threat stimuli; not an artifact of differences in within-subject variability in reaction time; nor accounted for by individual differences in anxiety-related variables. Findings are conceptualized as reflecting the understudied dynamics of AB expression, with implications for AB measurement and quantification, etiology, relations, and intervention research.

Dynamics of Attentional Bias to Threat in Anxious Adults: Bias towards and/or Away?

PubMed Central

Zvielli, Ariel; Bernstein, Amit; Koster, Ernst H. W.

2014-01-01

The aim of the present study was to question untested assumptions about the nature of the expression of Attentional Bias (AB) towards and away from threat stimuli. We tested the idea that high trait anxious individuals (N = 106; M(SD)age = 23.9(3.2) years; 68% women) show a stable AB towards multiple categories of threatening information using the emotional visual dot probe task. AB with respect to five categories of threat stimuli (i.e., angry faces, attacking dogs, attacking snakes, pointed weapons, violent scenes) was evaluated. In contrast with current theories, we found that 34% of participants expressed AB towards threat stimuli, 20.8% AB away from threat stimuli, and 34% AB towards some categories of threat stimuli and away from others. The multiple observed expressions of AB were not an artifact of a specific criterion AB score cut-off; not specific to certain categories of threat stimuli; not an artifact of differences in within-subject variability in reaction time; nor accounted for by individual differences in anxiety-related variables. Findings are conceptualized as reflecting the understudied dynamics of AB expression, with implications for AB measurement and quantification, etiology, relations, and intervention research. PMID:25093664

Decompression tables for inside chamber attendants working at altitude.

PubMed

Bell, James; Thombs, Paul A; Davison, William J; Weaver, Lindell K

2014-01-01

Hyperbaric oxygen (HBO2) multiplace chamber inside attendants (IAs) are at risk for decompression sickness (DCS). Standard decompression tables are formulated for sea-level use, not for use at altitude. At Presbyterian/St. Luke's Medical Center (Denver, Colorado, 5,924 feet above sea level) and Intermountain Medical Center (Murray, Utah, 4,500 feet), the decompression obligation for IAs is managed with U.S. Navy Standard Air Tables corrected for altitude, Bühlmann Tables, and the Nobendem© calculator. IAs also breathe supplemental oxygen while compressed. Presbyterian/St. Luke's (0.83 atmospheres absolute/atm abs) uses gauge pressure, uncorrected for altitude, at 45 feet of sea water (fsw) (2.2 atm abs) for routine wound care HBO2 and 66 fsw (2.8 atm abs) for carbon monoxide/cyanide poisoning. Presbyterian/St. Luke's provides oxygen breathing for the IAs at 2.2 atm abs. At Intermountain (0.86 atm abs), HBO2 is provided at 2.0 atm abs for routine treatments and 3.0 atm abs for carbon monoxide poisoning. Intermountain IAs breathe intermittent 50% nitrogen/50% oxygen at 3.0 atm abs and 100% oxygen at 2.0 atm abs. The chamber profiles include a safety stop. From 1990-2013, Presbyterian/St. Luke's had 26,900 total IA exposures: 25,991 at 45 fsw (2.2 atm abs) and 646 at 66 fsw (2.8 atm abs); there have been four cases of IA DCS. From 2008-2013, Intermountain had 1,847 IA exposures: 1,832 at 2 atm abs and 15 at 3 atm abs, with one case of IA DCS. At both facilities, DCS incidents occurred soon after the chambers were placed into service. Based on these results, chamber inside attendant risk for DCS at increased altitude is low when the inside attendants breathe supplemental oxygen.

Diagnostic algorithm for relapsing acquired demyelinating syndromes in children.

PubMed

Hacohen, Yael; Mankad, Kshitij; Chong, W K; Barkhof, Frederik; Vincent, Angela; Lim, Ming; Wassmer, Evangeline; Ciccarelli, Olga; Hemingway, Cheryl

2017-07-18

To establish whether children with relapsing acquired demyelinating syndromes (RDS) and myelin oligodendrocyte glycoprotein antibodies (MOG-Ab) show distinctive clinical and radiologic features and to generate a diagnostic algorithm for the main RDS for clinical use. A panel reviewed the clinical characteristics, MOG-Ab and aquaporin-4 (AQP4) Ab, intrathecal oligoclonal bands, and Epstein-Barr virus serology results of 110 children with RDS. A neuroradiologist blinded to the diagnosis scored the MRI scans. Clinical, radiologic, and serologic tests results were compared. The findings showed that 56.4% of children were diagnosed with multiple sclerosis (MS), 25.4% with neuromyelitis optica spectrum disorder (NMOSD), 12.7% with multiphasic disseminated encephalomyelitis (MDEM), and 5.5% with relapsing optic neuritis (RON). Blinded analysis defined baseline MRI as typical of MS in 93.5% of children with MS. Acute disseminated encephalomyelitis presentation was seen only in the non-MS group. Of NMOSD cases, 30.7% were AQP4-Ab positive. MOG-Ab were found in 83.3% of AQP4-Ab-negative NMOSD, 100% of MDEM, and 33.3% of RON. Children with MOG-Ab were younger, were less likely to present with area postrema syndrome, and had lower disability, longer time to relapse, and more cerebellar peduncle lesions than children with AQP4-Ab NMOSD. A diagnostic algorithm applicable to any episode of CNS demyelination leads to 4 main phenotypes: MS, AQP4-Ab NMOSD, MOG-Ab-associated disease, and antibody-negative RDS. Children with MS and AQP4-Ab NMOSD showed features typical of adult cases. Because MOG-Ab-positive children showed notable and distinctive clinical and MRI features, they were grouped into a unified phenotype (MOG-Ab-associated disease), included in a new diagnostic algorithm. © 2017 American Academy of Neurology.

Radiolabeled F(ab')2-cetuximab for theranostic purposes in colorectal and skin tumor-bearing mice models.

PubMed

Bellaye, P-S; Moreau, M; Raguin, O; Oudot, A; Bernhard, C; Vrigneaud, J-M; Dumont, L; Vandroux, D; Denat, F; Cochet, A; Brunotte, F; Collin, B

2018-05-17

This study aimed to investigate theranostic strategies in colorectal and skin cancer based on fragments of cetuximab, an anti-EGFR mAb, labeled with radionuclide with imaging and therapeutic properties, 111 In and 177 Lu, respectively. We designed F(ab') 2 -fragments of cetuximab radiolabeled with 111 In and 177 Lu. 111 In-F(ab') 2 -cetuximab tumor targeting and biodistribution were evaluated by SPECT in BalbC nude mice bearing primary colorectal tumors. The efficacy of 111 In-F(ab') 2 -cetuximab to assess therapy efficacy was performed on BalbC nude mice bearing colorectal tumors receiving 17-DMAG, an HSP90 inhibitor. Therapeutic efficacy of the radioimmunotherapy based on 177 Lu-F(ab') 2 -cetuximab was evaluated in SWISS nude mice bearing A431 tumors. Radiolabeling procedure did not change F(ab') 2 -cetuximab and cetuximab immunoreactivity nor affinity for HER1 in vitro. 111 In-DOTAGA-F(ab') 2 -cetuximab exhibited a peak tumor uptake at 24 h post-injection and showed a high tumor specificity determined by a significant decrease in tumor uptake after the addition of an excess of unlabeled-DOTAGA-F(ab') 2 -cetuximab. SPECT imaging of 111 In-DOTAGA-F(ab') 2 -cetuximab allowed an accurate evaluation of tumor growth and successfully predicted the decrease in tumor growth induced by 17-DMAG. Finally, 177 Lu-DOTAGA-F(ab') 2 -cetuximab radioimmunotherapy showed a significant reduction of tumor growth at 4 and 8 MBq doses. 111 In-DOTAGA-F(ab') 2 -cetuximab is a reliable and stable tool for specific in vivo tumor targeting and is suitable for therapy efficacy assessment. 177 Lu-DOTAGA-F(ab') 2 -cetuximab is an interesting theranostic tool allowing therapy and imaging.

An Optical Waveguide Humidity Detector.

DTIC Science & Technology

1985-10-31

films: (1) AB#11; (2) AB#13-, (3) AB#14. 14 ............................................... nl ,, I ,, . . . . m 1500 S1000- w z b. os,, ooo 0 500 -b...4c are for the following films: (1) AB#11. (2) AB#13; (3) AB#14. -1 15 1500 a. b. < 1000- w C. z o 500 0~0 w 20 40 60 80 REL. HUMIDITY (%) (c) Fig. 4...400*- . . . . . . . . . . * .\\ 2787 6- // CL 74 z 0 72 A I’- ., / // _0 .- O - IL, - f 7 I I I II .2 .4 .6 .8 1 NORM. FACTOR (N.F.) Fig. 6

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

Solanki, Ashish K.; Rathore, Yogendra S.; Badmalia, Maulik D.

Asymmetric disposition of Fab arms in the structures solved for the broadly neutralizing monoclonal antibody (nmAb) IgG1 b12 raised the question of whether the unusual shape observed for b12 is common for all IgG1 mAbs or if there is a difference in the overall shape of nmAbs versus non-nmAbs. In this paper, we compared small angle x-ray scattering (SAXS) data-based models and limited proteolysis profiles of some IgG1 mAbs known to be having and lacking HIV-1 neutralizing potency. In non-nmAbs, the Fab arms were found to be symmetrically disposed in space relative to central Fc, but in most nmAbs, themore » Fab arms were asymmetrically disposed, as seen for IgG1 b12. The only exceptions were 2G12 and 4E10, where both Fab arms were closed above Fc, suggesting some Fab-Fc and/or Fab-Fab interaction in the nmAbs that constrained extension of the Fab-Fc linker. Interestingly, these observations were correlated with differential proteolysis profiles of the mAbs by papain. Under conditions when papain could cut both Fab arms of non-nmAbs, only one Fab arm could be removed from neutralizing ones (except for 2G12 and 4E10). Chromatography and small angle x-ray scattering results of papain-digested products revealed that 1) the Fab-Fc or Fab-Fab interactions in unliganded mAbs are retained in digested products, and 2) whereas anti-gp120 non-nmAbs could bind two gp120 molecules, nmAbs could bind only one gp120. Finally, additional experiments showed that except for 2G12 and 4E10, unopen shapes of nmAbs remain uninfluenced by ionic strength but can be reversibly opened by low pH of buffer accompanied by loss of ligand binding ability.« less

The FhaB/FhaC two-partner secretion system is involved in adhesion of Acinetobacter baumannii AbH12O-A2 strain

PubMed Central

Pérez, A.; Merino, M.; Rumbo-Feal, S.; Álvarez-Fraga, L.; Vallejo, J. A.; Beceiro, A.; Ohneck, E. J.; Mateos, J.; Fernández-Puente, P.; Actis, L. A.; Poza, M.; Bou, G.

2017-01-01

ABSTRACT Acinetobacter baumannii is a hospital-acquired pathogen that shows an extraordinary capacity to stay in the hospital environment. Adherence of the bacteria to eukaryotic cells or to abiotic surfaces is the first step for establishing an infection. The A. baumannii strain AbH12O-A2 showed an exceptional ability to adhere to A549 epithelial cells. The AbFhaB/FhaC 2-partner secretion (TPS) system involved in adhesion was discovered after the screening of the recently determined A. baumannii AbH12O-A2 strain genome (CP009534.1). The AbFhaB is a large exoprotein which transport to the bacterial surface is mediated by the AbFhaC protein. In the present study, the role of this TPS system in the AbH12O-A2 adherence phenotype was investigated. The functional inactivation of this 2-partner secretion system was addressed by analyzing the outer membrane vesicles (OMV) proteomic profile from the wild-type strain and its derivative mutant AbH12O-A2ΔfhaC demonstrating that AbFhaB is no longer detected in the absence of AbFhaC. Scanning electron microscopy (SEM) and adhesion experiments demonstrated that inactivation of the AbFhaB/FhaC system significantly decreases bacterial attachment to A549 alveolar epithelial cells. Moreover, it has been demonstrated that this 2-partner secretion system is involved in fibronectin-mediated adherence of the A. baumannii AbH12O-A2 isolate. Finally, we report that the AbFhaB/FhaC system is involved in virulence when tested using invertebrate and vertebrate hosts. These data suggest the potential role that this AbFhaB/FhaC secretion system could play in the pathobiology of A. baumannii. PMID:27858524

The FhaB/FhaC two-partner secretion system is involved in adhesion of Acinetobacter baumannii AbH12O-A2 strain.

PubMed

Pérez, A; Merino, M; Rumbo-Feal, S; Álvarez-Fraga, L; Vallejo, J A; Beceiro, A; Ohneck, E J; Mateos, J; Fernández-Puente, P; Actis, L A; Poza, M; Bou, G

2017-08-18

Acinetobacter baumannii is a hospital-acquired pathogen that shows an extraordinary capacity to stay in the hospital environment. Adherence of the bacteria to eukaryotic cells or to abiotic surfaces is the first step for establishing an infection. The A. baumannii strain AbH12O-A2 showed an exceptional ability to adhere to A549 epithelial cells. The AbFhaB/FhaC 2-partner secretion (TPS) system involved in adhesion was discovered after the screening of the recently determined A. baumannii AbH12O-A2 strain genome (CP009534.1). The AbFhaB is a large exoprotein which transport to the bacterial surface is mediated by the AbFhaC protein. In the present study, the role of this TPS system in the AbH12O-A2 adherence phenotype was investigated. The functional inactivation of this 2-partner secretion system was addressed by analyzing the outer membrane vesicles (OMV) proteomic profile from the wild-type strain and its derivative mutant AbH12O-A2ΔfhaC demonstrating that AbFhaB is no longer detected in the absence of AbFhaC. Scanning electron microscopy (SEM) and adhesion experiments demonstrated that inactivation of the AbFhaB/FhaC system significantly decreases bacterial attachment to A549 alveolar epithelial cells. Moreover, it has been demonstrated that this 2-partner secretion system is involved in fibronectin-mediated adherence of the A. baumannii AbH12O-A2 isolate. Finally, we report that the AbFhaB/FhaC system is involved in virulence when tested using invertebrate and vertebrate hosts. These data suggest the potential role that this AbFhaB/FhaC secretion system could play in the pathobiology of A. baumannii.

Thyroglobulin (Tg) Testing Revisited: Tg Assays, TgAb Assays, and Correlation of Results With Clinical Outcomes.

PubMed

Netzel, Brian C; Grebe, Stefan K G; Carranza Leon, B Gisella; Castro, M Regina; Clark, Penelope M; Hoofnagle, Andrew N; Spencer, Carole A; Turcu, Adina F; Algeciras-Schimnich, Alicia

2015-08-01

Measurement of thyroglobulin (Tg) by mass spectrometry (Tg-MS) is emerging as a tool for accurate Tg quantification in patients with anti-Tg autoantibodies (TgAbs). The objective of the study was to perform analytical and clinical evaluations of two Tg-MS assays in comparison with immunometric Tg assays (Tg-IAs) and Tg RIAs (Tg-RIAs) in a cohort of thyroid cancer patients. A total of 589 samples from 495 patients, 243 TgAb-/252 TgAb+, were tested by Beckman, Roche, Siemens-Immulite, and Thermo-Brahms Tg and TgAb assays, two Tg-RIAs, and two Tg-MS assays. The frequency of TgAb+ was 58%, 41%, 27%, and 39% for Roche, Beckman, Siemens-Immulite, and Thermo-Brahms, respectively. In TgAb- samples, clinical sensitivities and specificities of 100% and 74%-100%, respectively, were observed across all assays. In TgAb+ samples, all Tg-IAs demonstrated assay-dependent Tg underestimation, ranging from 41% to 86%. In TgAb+ samples, the use of a common cutoff (0.5 ng/mL) for the Tg-MS, three Tg-IAs, and the USC-RIA improved the sensitivity for the Tg-MSs and Tg-RIAs when compared with the Tg-IAs. In up to 20% of TgAb+ cases, Tg-IAs failed to detect Tg that was detectable by Tg-MS. In Tg-RIAs false-high biases were observed in TgAb+ samples containing low Tg concentrations. Tg-IAs remain the method of choice for Tg quantitation in TgAb- patients. In TgAb+ patients with undetectable Tg by immunometric assay, the Tg-MS will detect Tg in up to 20% additional cases. The Tg-RIA will detect Tg in approximately 35% cases, but a significant proportion of these will be clinical false-positive results. The undetectable Tg-MS seen in approximately 40% of TgAb+ cases in patients with disease need further evaluation.

A Multiplex Microsphere-Based Immunoassay Increases the Sensitivity of SIV-Specific Antibody Detection in Serum Samples and Mucosal Specimens Collected from Rhesus Macaques Infected with SIVmac239.

PubMed

Powell, Rebecca L R; Ouellette, Ian; Lindsay, Ross W; Parks, Christopher L; King, C Richter; McDermott, Adrian B; Morrow, Gavin

2013-06-01

Results from recent HIV-1 vaccine studies have indicated that high serum antibody (Ab) titers may not be necessary for Ab-mediated protection, and that Abs localized to mucosal sites might be critical for preventing infection. Enzyme-linked immunosorbent assay (ELISA) has been used for decades as the gold standard for Ab measurement, though recently, highly sensitive microsphere-based assays have become available, with potential utility for improved detection of Abs. In this study, we assessed the Bio-Plex(®) Suspension Array System for the detection of simian immunodeficiency virus (SIV)-specific Abs in rhesus macaques (RMs) chronically infected with SIV, whose serum or mucosal SIV-specific Ab titers were negative by ELISA. We developed a SIVmac239-specific 4-plex bead array for the simultaneous detection of Abs binding to Env, Gag, Pol, and Nef. The 4-plex assay was used to quantify SIV-specific serum IgG and rectal swab IgA titers from control (SIV-naive) and SIVmac239-infected RMs. The Bio-Plex assay specifically detected anti-SIV Abs in specimens from SIV-infected animals for all four analytes when compared to SIV-naive control samples (p≤0.04). Furthermore, in 70% of Env and 79% of Gag ELISA-negative serum samples, specific Ab was detected using the Bio-Plex assay. Similarly, 71% of Env and 48% of Gag ELISA-negative rectal swab samples were identified as positive using the Bio-Plex assay. Importantly, assay specificity (i.e., probability of true positives) was comparable to ELISA (94%-100%). The results reported here indicate that microsphere-based methods provide a substantial improvement over ELISA for the detection of Ab responses, aid in detecting specific Abs when analyzing samples containing low levels of Abs, such as during the early stages of a vaccine trial, and may be valuable in attempts to link protective efficacy of vaccines with induced Ab responses.

Antibody to Ricin A Chain Hinders Intracellular Routing of Toxin and Protects Cells Even after Toxin Has Been Internalized

PubMed Central

Song, Kejing; Mize, R. Ranney; Marrero, Luis; Corti, Miriam; Kirk, Jason M.; Pincus, Seth H.

2013-01-01

Background Mechanisms of antibody-mediated neutralization are of much interest. For plant and bacterial A-B toxins, A chain mediates toxicity and B chain binds target cells. It is generally accepted and taught that antibody (Ab) neutralizes by preventing toxin binding to cells. Yet for some toxins, ricin included, anti-A chain Abs afford greater protection than anti-B. The mechanism(s) whereby Abs to the A chain neutralize toxins are not understood. Methodology/Principal Findings We use quantitative confocal imaging, neutralization assays, and other techniques to study how anti-A chain Abs function to protect cells. Without Ab, ricin enters cells and penetrates to the endoplasmic reticulum within 15 min. Within 45–60 min, ricin entering and being expelled from cells reaches equilibrium. These results are consistent with previous observations, and support the validity of our novel methodology. The addition of neutralizing Ab causes ricin accumulation at the cell surface, delays internalization, and postpones retrograde transport of ricin. Ab binds ricin for >6hr as they traffic together through the cell. Ab protects cells even when administered hours after exposure. Conclusions/Key Findings We demonstrate the dynamic nature of the interaction between the host cell and toxin, and how Ab can alter the balance in favor of the cell. Ab blocks ricin’s entry into cells, hinders its intracellular routing, and can protect even after ricin is present in the target organelle, providing evidence that the major site of neutralization is intracellular. These data add toxins to the list of pathogenic agents that can be neutralized intracellularly and explain the in vivo efficacy of delayed administration of anti-toxin Abs. The results encourage the use of post-exposure passive Ab therapy, and show the importance of the A chain as a target of Abs. PMID:23638075

Global Shape and Ligand Binding Efficiency of the HIV-1-neutralizing Antibodies Differ from Those of Antibodies That Cannot Neutralize HIV-1*

PubMed Central

Solanki, Ashish K.; Rathore, Yogendra S.; Badmalia, Maulik D.; Dhoke, Reema R.; Nath, Samir K.; Nihalani, Deepak; Ashish

2014-01-01

Asymmetric disposition of Fab arms in the structures solved for the broadly neutralizing monoclonal antibody (nmAb) IgG1 b12 raised the question of whether the unusual shape observed for b12 is common for all IgG1 mAbs or if there is a difference in the overall shape of nmAbs versus non-nmAbs. We compared small angle x-ray scattering (SAXS) data-based models and limited proteolysis profiles of some IgG1 mAbs known to be having and lacking HIV-1 neutralizing potency. In non-nmAbs, the Fab arms were found to be symmetrically disposed in space relative to central Fc, but in most nmAbs, the Fab arms were asymmetrically disposed, as seen for IgG1 b12. The only exceptions were 2G12 and 4E10, where both Fab arms were closed above Fc, suggesting some Fab-Fc and/or Fab-Fab interaction in the nmAbs that constrained extension of the Fab-Fc linker. Interestingly, these observations were correlated with differential proteolysis profiles of the mAbs by papain. Under conditions when papain could cut both Fab arms of non-nmAbs, only one Fab arm could be removed from neutralizing ones (except for 2G12 and 4E10). Chromatography and small angle x-ray scattering results of papain-digested products revealed that 1) the Fab-Fc or Fab-Fab interactions in unliganded mAbs are retained in digested products, and 2) whereas anti-gp120 non-nmAbs could bind two gp120 molecules, nmAbs could bind only one gp120. Additional experiments showed that except for 2G12 and 4E10, unopen shapes of nmAbs remain uninfluenced by ionic strength but can be reversibly opened by low pH of buffer accompanied by loss of ligand binding ability. PMID:25331945

Global Shape and Ligand Binding Efficiency of the HIV-1-neutralizing Antibodies Differ from Those of Antibodies That Cannot Neutralize HIV-1

DOE PAGES

Solanki, Ashish K.; Rathore, Yogendra S.; Badmalia, Maulik D.; ...

2014-10-20

Asymmetric disposition of Fab arms in the structures solved for the broadly neutralizing monoclonal antibody (nmAb) IgG1 b12 raised the question of whether the unusual shape observed for b12 is common for all IgG1 mAbs or if there is a difference in the overall shape of nmAbs versus non-nmAbs. In this paper, we compared small angle x-ray scattering (SAXS) data-based models and limited proteolysis profiles of some IgG1 mAbs known to be having and lacking HIV-1 neutralizing potency. In non-nmAbs, the Fab arms were found to be symmetrically disposed in space relative to central Fc, but in most nmAbs, themore » Fab arms were asymmetrically disposed, as seen for IgG1 b12. The only exceptions were 2G12 and 4E10, where both Fab arms were closed above Fc, suggesting some Fab-Fc and/or Fab-Fab interaction in the nmAbs that constrained extension of the Fab-Fc linker. Interestingly, these observations were correlated with differential proteolysis profiles of the mAbs by papain. Under conditions when papain could cut both Fab arms of non-nmAbs, only one Fab arm could be removed from neutralizing ones (except for 2G12 and 4E10). Chromatography and small angle x-ray scattering results of papain-digested products revealed that 1) the Fab-Fc or Fab-Fab interactions in unliganded mAbs are retained in digested products, and 2) whereas anti-gp120 non-nmAbs could bind two gp120 molecules, nmAbs could bind only one gp120. Finally, additional experiments showed that except for 2G12 and 4E10, unopen shapes of nmAbs remain uninfluenced by ionic strength but can be reversibly opened by low pH of buffer accompanied by loss of ligand binding ability.« less

Autoantibodies to human tryptophan hydroxylase and aromatic L-amino acid decarboxylase.

PubMed

Dal Pra, Chiara; Chen, Shu; Betterle, Corrado; Zanchetta, Renato; McGrath, Vivienne; Furmaniak, Jadwiga; Rees Smith, Bernard

2004-03-01

To assess the prevalence of autoantibodies (Abs) to tryptophan hydroxylase (TPH) and aromatic l-amino acid decarboxylase (AADC) in patients with different autoimmune diseases and to analyse their respective epitopes. TPH and AADC Abs were measured in an immunoprecipitation assay using (35)S-labelled full-length and fragments of TPH and AADC. Patients with different autoimmune adrenal diseases (n=84), non-adrenal autoimmune diseases (n=37), idiopathic vitiligo (n=8) and 56 healthy blood donors were studied. Fourteen of twenty-three (61%) of patients with autoimmune polyglandular syndrome (APS) type I and 1/34 (3%) of patients with isolated Addison's disease (AD) were positive for TPH Abs. None of the patients with APS type II (n=27), coeliac disease (n=10), autoimmune thyroid disease (AITD) (n=11), type 1 diabetes mellitus (DM) (n=16) or idiopathic vitiligo (n=8) was positive for TPH Abs. AADC Abs were detected in 12/23 (52%) patients with APS type I, in 1/29 (3%) patients with APS type II and 1/34 (3%) patients with isolated AD. None of the patients with coeliac disease, type 1 DM, AITD or idiopathic vitiligo was positive for AADC Abs. TPH Abs were found to interact with the C-terminal amino acids (aa) 308-423, central aa 164-205 and N-terminal aa 1-105 of the TPH molecule. AADC Ab binding epitopes were within the C-terminal aa 382-483, the central aa 243-381 and the N-terminal aa 1-167. Our study suggests that TPH Abs and AADC Abs react with several different epitopes and that different epitopes are recognized by different sera. The prevalence of TPH Abs and AADC Abs in patients with APS type I in our study is in agreement with previous reports. TPH Abs and AADC Abs were found very rarely in patients with other forms of autoimmune adrenal disease and were not detected in patients with non-adrenal autoimmune diseases.

Staged induction of HIV-1 glycan–dependent broadly neutralizing antibodies

DOE PAGES

Bonsignori, Mattia; Kreider, Edward F.; Fera, Daniela; ...

2017-03-15

A preventive HIV-1 vaccine should induce HIV-1–specific broadly neutralizing antibodies (bnAbs). However, bnAbs generally require high levels of somatic hypermutation (SHM) to acquire breadth, and current vaccine strategies have not been successful in inducing bnAbs. Because bnAbs directed against a glycosylated site adjacent to the third variable loop (V3) of the HIV-1 envelope protein require limited SHM, the V3-glycan epitope is an attractive vaccine target. By studying the cooperation among multiple V3-glycan B cell lineages and their coevolution with autologous virus throughout 5 years of infection, we identify key events in the ontogeny of a V3- glycan bnAb. Two autologousmore » neutralizing antibody lineages selected for virus escape mutations and consequently allowed initiation and affinity maturation of a V3-glycan bnAb lineage. The nucleotide substitution required to initiate the bnAb lineage occurred at a low-probability site for activation-induced cytidine deaminase activity. Cooperation of B cell lineages and an improbable mutation critical for bnAb activity defined the necessary events leading to breadth in this V3- glycan bnAb lineage. These findings may, in part, explain why initiation of V3-glycan bnAbs is rare, and suggest an immunization strategy for inducing similar V3-glycan bnAbs.« less

Domain-based assays of individual antibody concentrations in an oligoclonal combination targeting a single protein.

PubMed

Meng, Q; Li, M; Silberg, M A; Conrad, F; Bettencourt, J; To, R; Huang, C; Ma, J; Meyer, K; Shimizu, R; Cao, L; Tomic, M T; Marks, J D

2012-02-15

Quantitation of individual monoclonal antibodies (mAbs) within a combined antibody drug product is required for preclinical and clinical drug development, including pharmacokinetic (PK), toxicology, stability, and biochemical characterization studies of such drugs. We have developed an antitoxin, XOMA 3AB, consisting of three recombinant mAbs that potently neutralize the known subtypes of type A botulinum neurotoxin (BoNT/A). The three mAbs bind nonoverlapping BoNT/A epitopes with high affinity. XOMA 3AB is being developed as a treatment for botulism resulting from BoNT/A. To develop antibody-specific assays, we cloned, expressed, and purified BoNT/A domains from Escherichia coli. Each mAb bound only to its specific domain with affinity comparable to the binding to holotoxin. mAb-specific domains were used to develop an enzyme-linked immunosorbent assay (ELISA) for characterization of the integrity and binding activity of the three mAbs in the drug product. An electrochemiluminescence bridging assay that is robust to interference from components in serum was also developed, and we demonstrate that it can be used for PK assays. This type of antigen engineering to generate mAb-specific domains is a general method allowing quantitation and characterization of individual mAbs in a mAb cocktail that binds the same protein and is superior to anti-idiotype approaches. Copyright © 2011 Elsevier Inc. All rights reserved.

Abnormal IgG4 antibody response to aeroallergens in allergic patients.

PubMed

Jeannin, P; Delneste, Y; Tillie-Leblond, I; Wallaert, B; carlier, A; Pestel, J; Tonnel, A B

1994-01-01

Various studies have suggested the involvement of immunoglobulin G4 (IgG4) antibodies (Ab) in the physiopathology of allergic disorders. Recently, an abnormal IgG4 Ab production in response to immunization has been reported in some atopic patients. Thus, in order to evidence in allergic patients, a potential abnormal IgG4 Ab response to aeroallergens following natural exposure, we compared, in 34 patients sensitive to Dermatophagoides pteronyssinus and in 16 healthy subjects, the IgG4 Ab response to D. pteronyssinus, grass pollen and cat dander, using a solid-phase radioimmunoassay. Since some patients were also sensitive to grass pollen and/or to cat dander, we analyzed, in all patients, the IgG4 Ab responses both towards the allergen(s) they were sensitive to (sensitizing allergen) or not (unrelated allergen). The results showed that 90% of the patients produced levels of antisensitizing allergen(s) IgG4 Ab significantly higher than the controls; this IgG4 Ab response was correlated with the corresponding specific IgE Ab level. In addition, among these patients, around 40% presented high levels of IgG4 Ab to the unrelated allergen(s). Thus, in allergic patients, while specific IgE Ab define the nature of the sensitizing allergen, the presence of IgG4 Ab directed against various allergens seems in relation with an abnormal isotype regulation associated with atopic disorders.

Staged induction of HIV-1 glycan–dependent broadly neutralizing antibodies

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

Bonsignori, Mattia; Kreider, Edward F.; Fera, Daniela

A preventive HIV-1 vaccine should induce HIV-1–specific broadly neutralizing antibodies (bnAbs). However, bnAbs generally require high levels of somatic hypermutation (SHM) to acquire breadth, and current vaccine strategies have not been successful in inducing bnAbs. Because bnAbs directed against a glycosylated site adjacent to the third variable loop (V3) of the HIV-1 envelope protein require limited SHM, the V3-glycan epitope is an attractive vaccine target. By studying the cooperation among multiple V3-glycan B cell lineages and their coevolution with autologous virus throughout 5 years of infection, we identify key events in the ontogeny of a V3- glycan bnAb. Two autologousmore » neutralizing antibody lineages selected for virus escape mutations and consequently allowed initiation and affinity maturation of a V3-glycan bnAb lineage. The nucleotide substitution required to initiate the bnAb lineage occurred at a low-probability site for activation-induced cytidine deaminase activity. Cooperation of B cell lineages and an improbable mutation critical for bnAb activity defined the necessary events leading to breadth in this V3- glycan bnAb lineage. These findings may, in part, explain why initiation of V3-glycan bnAbs is rare, and suggest an immunization strategy for inducing similar V3-glycan bnAbs.« less

Effectiveness of antilock braking systems in reducing motorcycle fatal crash rates.

PubMed

Teoh, Eric R

2011-04-01

Overbraking and underbraking have been shown to be common factors in motorcycle crashes. Antilock braking systems (ABS) prevent wheels from locking during braking and may make riders less reluctant to apply full braking force. The objective of this study was to evaluate the effect of ABS in fatal motorcycle crashes. Motorcycle drivers involved in fatal crashes per 10,000 registered vehicle years were compared for 13 motorcycle models with optional ABS and those same models without the option during 2003-2008. Motorcycles with optional ABS were included only if the presence of the option could be identified from the vehicle identification number. The rate of fatal motorcycle crashes per 10,000 registered vehicle years was 37 percent lower for ABS models than for their non-ABS versions. ABS appears to be highly effective in preventing fatal motorcycle crashes based on some early adopters of motorcycle ABS technology.

Negative tail fusions can improve ruggedness of single domain antibodies.

PubMed

Goldman, Ellen R; Brozozog-Lee, P Audrey; Zabetakis, Dan; Turner, Kendrick B; Walper, Scott A; Liu, Jinny L; Anderson, George P

2014-03-01

Single-domain antibodies (sdAbs), the recombinantly expressed binding domains derived from the heavy-chain-only antibodies found in camelids and sharks, are valued for their ability to refold after heat denaturation. However, some sdAbs are prone to aggregation on extended heating at high concentration. Additionally, sdAbs prepared cytoplasmically often lack the conserved disulfide bond found in variable heavy domains, which both decreases their melting point and can decrease their ability to refold. Genetic fusions of sdAbs with the acid tail of α-synuclein (ATS) resulted in constructs that had enhanced ability to resist aggregation. In addition, almost complete refolding was observed even in the absence of the disulfide bond. These sdAb-ATS fusions expand the utility of sdAbs. They provide sdAbs that are resistant to aggregation, and enable the production of re-foldable sdAbs in the reducing environment of the cytoplasm. Published by Elsevier Inc.

Anti-GD2 mAbs and next-generation mAb-based agents for cancer therapy

PubMed Central

Perez Horta, Zulmarie; Goldberg, Jacob L; Sondel, Paul M

2016-01-01

Tumor-specific monoclonal antibodies (mAbs) have demonstrated efficacy in the clinic, becoming an important approach for cancer immunotherapy. Due to its limited expression on normal tissue, the GD2 disialogangloside expressed on neuroblastoma cells is an excellent candidate for mAb therapy. In 2015, dinutuximab (an anti-GD2 mAb) was approved by the US FDA and is currently used in a combination immunotherapeutic regimen for the treatment of children with high-risk neuroblastoma. Here, we review the extensive preclinical and clinical development of anti-GD2 mAbs and the different mechanisms by which they mediate tumor cell killing. In addition, we discuss different mAb-based strategies that capitalize on the targeting ability of anti-GD2 mAbs to potentially deliver, as monotherapy, or in combination with other treatments, improved antitumor efficacy. PMID:27485082

Acoustic Performance of 3D Printed Nanocomposite Earmuff

PubMed Central

Ahmadi, Saeid; Nassiri, Parvin; Ghasemi, Ismaeil; Monazzam Ep, Mohammad R.

2016-01-01

Introduction: Hearing protection devices are one of the primary noise reduction tools in developing countries. This study is intended to produce and apply acrylonitrile butadiene styrene (ABS)/clay nanocomposites to fabricate a laboratory single cup earmuffs and then compare it with double cup and single cup pure ABS earmuffs in terms of noise attenuation performance and comfort. In addition, the noise attenuation performance of single cup pure ABS earmuffs is compared with double cup pure ABS earmuffs. Methods: ABS/nanoclay filament was fabricated using a twin screw extruder. A three dimensional (3D) printing machine and a 3D model of earcup, designed by solid work software, were applied to print single and double cup earmuffs using ABS/nanoclay composite and pure ABS filaments. Finally, using an acoustic test fixture, objective noise attenuation test was performed on three different types of earmuffs, including with and without nano material and a secondary cup. Moreover, earmuffs weight was measured as a comfort component. Results: Insertion loss and calculated noise reduction rating (NRR) of single cup ABS/nanoclay earmuffs (NRR=19.4 dB) and double cup pure ABS earmuffs (NRR=18.93 dB) were improved in comparison with single cup pure ABS earmuffs (NRR=15.7 dB). Additionally, both single cup earmuffs were significantly lighter than double cup earmuffs. Although single cup nano and double cup earmuffs had nearly the same attenuation performance, single cup nano earmuffs were 74 gr lighter than double cup earmuffs, so with reference to comfort, single cup nano earmuffs will probably be more acceptable. Conclusions: From this survey it might be concluded that, even though single cup ABS/nanoclay earmuffs was lighter than double cup pure ABS earmuffs, it had approximately more attenuation performance in comparison with double cup pure ABS earmuffs. Consequently, users are probably more prone to wear light- weight single cup ABS/nanoclay earmuffs as a result of improved comfort. In short, ABS/nanoclay composite can be considered a good choice in products with the necessity of high acoustic performance and low weight. PMID:26234977

Addition of an extra immunoglobulin domain to two anti-rodent TNF monoclonal antibodies substantially increased their potency.

PubMed

Scallon, Bernard; Cai, Ann; Radewonuk, Jennifer; Naso, Michael

2004-05-01

The functional valency of a monoclonal antibody (mAb) has important influences on such things as antigen avidity, Fc-mediated immune effector functions, and clearance of immune complexes. cV1q, a neutralizing rat/mouse chimeric anti-mouse tumor necrosis factor (TNF) monoclonal antibody (mAb), and Rt108, a neutralizing mouse anti-rat TNF (anti-raTNF) mAb, appear to be functionally monovalent for TNF-binding despite containing two antigen binding sites. The functional monovalency of these two independent anti-rodent TNF mAbs is presumably a result of steric hindrance from one TNF molecule binding to one Fab arm that prevents binding of a second TNF molecule to the other Fab arm. To test whether this steric hindrance could be overcome by introducing extra space and flexibility between the Fab arms, these mAbs were engineered to contain an extra CH1 immunoglobulin domain between the CH1 and hinge domains of their heavy chains. In vitro binding data showed that, compared to the original mAbs, the modified mAbs (S-mAbs) had greater capability of binding two TNF molecules simultaneously. In vitro activity assays showed that, compared to the original mAbs, the S-mAbs had significantly greater TNF-neutralization potency, with the S-mAb version of cV1q (S-cV1q) being 200-fold more effective at blocking mouse TNF (muTNF) and the S-mAb version of Rt108 (S-Rt108) being 20-fold more effective at blocking raTNF. Similar results were observed in vivo, where S-cV1q was between 100- and 500-fold more protective than cV1q in mice challenged with endotoxin. These data reveal that introduction of another constant region immunoglobulin domain into two unrelated mAbs dramatically enhanced their neutralization potency. Other mAbs may also show more potent activity using this engineering approach, particularly mAbs that recognize homopolymeric antigens.

Identification of unique B virus (Macacine Herpesvirus 1) epitopes of zoonotic and macaque isolates using monoclonal antibodies

PubMed Central

Vasireddi, Mugdha; Patrusheva, Irina; Seoh, Hyuk-Kyu; Filfili, Chadi N.; Wildes, Martin J.; Oh, Jay

2017-01-01

Our overall aim is to develop epitope-based assays for accurate differential diagnosis of B virus zoonotic infections in humans. Antibodies to cross-reacting epitopes on human-simplexviruses continue to confound the interpretation of current assays where abundant antibodies exist from previous infections with HSV types 1 and 2. To find B virus-specific epitopes we cloned ten monoclonal antibodies (mAbs) from the hybridomas we produced. Our unique collection of rare human sera from symptomatic and asymptomatic patients infected with B virus was key to the evaluation and identification of the mAbs as reagents in competition ELISAs (mAb-CE). The analysis of the ten mAbs revealed that the target proteins for six mAbs was glycoprotein B of which two are reactive to simian simplexviruses and not to human simplexviruses. Two mAbs reacted specifically with B virus glycoprotein D, and two other mAbs were specific to VP13/14 and gE-gI complex respectively. The mAbs specific to VP13/14 and gE-gI are strain specific reacting with B virus isolates from rhesus and Japanese macaques and not with isolates from cynomolgus and pigtail macaques. The mAb-CE revealed that a high proportion of naturally B virus infected rhesus macaques and two symptomatic humans possess antibodies to epitopes of VP13/14 protein and on the gE-gI complex. The majority of sera from B virus infected macaques and simplexvirus-infected humans competed with the less specific mAbs. These experiments produced a novel panel of mAbs that enabled B virus strain identification and confirmation of B virus infected macaques by the mAb-CE. For human sera the mAb-CE could be used only for selected cases due to the selective B virus strain-specificity of the mAbs against VP13/14 and gE/gI. To fully accomplish our aim to provide reagents for unequivocal differential diagnosis of zoonotic B virus infections, additional mAbs with a broader range of specificities is critical. PMID:28783746

Differential intra-endothelial delivery of polymer nanocarriers targeted to distinct PECAM-1 epitopes

PubMed Central

Garnacho, Carmen; Albelda, Steven M.; Muzykantov, Vladimir R.; Muro, Silvia

2008-01-01

Coupling drug carriers to antibodies for targeting endothelial cells (ECs) may improve treatment of vascular and pulmonary diseases. Selecting antibodies that deliver carriers to the cell surface or intracellularly may further optimize specifcity of interventions. We studied antibody-directed targeting of nanocarriers to platelet–endothelial cell adhesion molecule (PECAM)-1, an endothelial glycoprotein containing 6 Ig-like extracellular domains. PECAM-1 antibodies bind to ECs without internalization, but ECs internalize by endocytosis nanocarriers carrying multiple copies of anti-PECAM (anti-PECAM/NCs). To determine whether binding and intracellular transport of anti-PECAM/NCs depend on the epitope engaged, we targeted five PECAM-1 epitopes: mAb35, mAb37 and mAb62 (membrane-distal Ig domain 1), mAbGi34 (Ig domains 2/3), and mAb4G6 (membrane-proximal Ig domain 6). The antibodies bound to ECs regardless of the epitope proximity to the plasmalemma, whereas 130 nm diameter nanocarriers only targeted effectively distal domains (mAb4G6/NCs did not bind to ECs). ECs internalized mAb35, mAb62, and mAbGi34 carriers regardless of their size (0.13 to 5 µm diameter), yet they did not internalize mAb37/NCs. After internalization, mAb62/NCs trafficked to lysosomes within 2–3 h, whereas mAb35/NCs had prolonged residence in pre-lysosomal vesicles. Therefore, endothelial binding, endocytosis, and intracellular transport of anti-PECAM/NCs are epitope-specific. This paradigm will guide the design of endothelial drug delivery systems providing specific cellular localizations. PMID:18606202

Adsorption of transgenic insecticidal Cry1Ab protein to SiO2. 2. Patch-controlled electrostatic attraction.

PubMed

Madliger, Michael; Sander, Michael; Schwarzenbach, René P

2010-12-01

Adsorption governs the fate of Cry proteins from genetically modified Bt crops in soils. The effect of ionic strength (I) on the adsorption of Cry1Ab (isoelectric point IEP(Cry1Ab) ≈ 6) to negatively charged quartz (SiO(2)) and positively charged poly-L-lysine (PLL) was investigated at pH 5 to 8, using quartz crystal microbalance with dissipation monitoring and optical waveguide lightmode spectroscopy. Cry1Ab adsorbed via positively and negatively charged surface patches to SiO(2) and PLL, respectively. This patch controlled electrostatic attraction (PCEA) explains the observed increase in Cry1Ab adsorption to sorbents that carried the same net charge as the protein (SiO(2) at pH > IEP(Cry1Ab) and PLL at pH < IEP(Cry1Ab)) with decreasing I. In contrast, the adsorption of two reference proteins, BSA and HEWL, with different adsorption mechanism, were little affected by similar changes of I. Consistent with PCEA, Cry1Ab desorption from SiO(2) at pH > IEP(Cry1Ab) increased with increasing I and pH. Weak Cry1Ab-SiO(2) PCEA above pH 7 resulted in reversible, concentration dependent adsorption. Solution depletion experiments showed that PCEA also governed Cry1Ab adsorption to SiO(2) particles at environmentally relevant concentrations (a few ng mL(-1)). These results imply that models describing Cry1Ab adsorption to charged surfaces in soils need to account for the nonuniform surface charge distribution of the protein.

The clinicopathological relevance of pretransplant anti-angiotensin II type 1 receptor antibodies in renal transplantation.

PubMed

Lee, Juhan; Huh, Kyu Ha; Park, Yongjung; Park, Borae G; Yang, Jaeseok; Jeong, Jong Cheol; Lee, Joongyup; Park, Jae Berm; Cho, Jang-Hee; Lee, Sik; Ro, Han; Han, Seung-Yeup; Kim, Myoung Soo; Kim, Yu Seun; Kim, Sung Joo; Kim, Chan-Duck; Chung, Wookyung; Park, Sung-Bae; Ahn, Curie

2017-07-01

Anti-angiotensin II type 1 receptor antibodies (AT1R-Abs) have been suggested as a risk factor for graft failure and acute rejection (AR). However, the prevalence and clinical significance of pretransplant AT1R-Abs have seldom been evaluated in Asia. In this multicenter, observational cohort study, we tested the AT1R-Abs in pretransplant serum samples obtained from 166 kidney transplant recipients. Statistical analysis was used to set a threshold AT1R-Abs level at 9.05 U/mL. Pretransplant AT1R-Abs were detected in 98/166 (59.0%) of the analyzed recipients. No graft loss or patient death was reported during the study period. AT1R-Abs (+) patients had a significantly higher incidence of biopsy-proven AR than AT1R-Abs (-) patients (27.6 versus 10.3%, P = 0.007). Recipients with pretransplant AT1R-Abs had a 3.2-fold higher risk of AR within a year of transplantation (P = 0.006). Five study subjects developed microcirculation inflammation (score ≥2). Four of them were presensitized to AT1R-Abs. In particular, three patients had a high titer of anti-AT1R-Abs (>22.7 U/mL). Pretransplant AT1R-Abs is an independent risk factor for AR, especially acute cellular rejection, and is possibly associated with the risk of antibody-mediated injury. Pretransplant assessment of AT1R-Abs may be useful for stratifying immunologic risks. © The Author 2015. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

Evolution of the Quorum network and the mobilome (plasmids and bacteriophages) in clinical strains of Acinetobacter baumannii during a decade.

PubMed

López, M; Rueda, A; Florido, J P; Blasco, L; Fernández-García, L; Trastoy, R; Fernández-Cuenca, F; Martínez-Martínez, L; Vila, J; Pascual, A; Bou, G; Tomas, M

2018-02-06

In this study, we compared eighteen clinical strains of A. baumannii belonging to the ST-2 clone and isolated from patients in the same intensive care unit (ICU) in 2000 (9 strains referred to collectively as Ab_GEIH-2000) and 2010 (9 strains referred to collectively as Ab_GEIH-2010), during the GEIH-REIPI project (Umbrella BioProject PRJNA422585). We observed two main molecular differences between the Ab_GEIH-2010 and the Ab_GEIH-2000 collections, acquired over the course of the decade long sampling interval and involving the mobilome: i) a plasmid harbouring genes for bla OXA 24/40 ß-lactamase and abKA/abkB proteins of a toxin-antitoxin system; and ii) two temperate bacteriophages, Ab105-1ϕ (63 proteins) and Ab105-2ϕ (93 proteins), containing important viral defence proteins. Moreover, all Ab_GEIH-2010 strains contained a Quorum functional network of Quorum Sensing (QS) and Quorum Quenching (QQ) mechanisms, including a new QQ enzyme, AidA, which acts as a bacterial defence mechanism against the exogenous 3-oxo-C12-HSL. Interestingly, the infective capacity of the bacteriophages isolated in this study (Ab105-1ϕ and Ab105-2ϕ) was higher in the Ab_GEIH-2010 strains (carrying a functional Quorum network) than in the Ab_GEIH-2000 strains (carrying a deficient Quorum network), in which the bacteriophages showed little or no infectivity. This is the first study about the evolution of the Quorum network and the mobilome in clinical strains of Acinetobacter baumannii during a decade.

Predictions Regarding the Performance of Field Emission Cathodes in Radio Frequency Guns

DTIC Science & Technology

2010-01-01

b2b by equation 2.3. Finally, ηb = ab/fb as defined in equation 2.5. Next...2.11) with η (r, z) = √ r2 + (z − fb)2 + √ r2 + (z + fb) 2 2fb where, summarizing from above, fb = √ a2 b − b2b and ηb = ab fb = ab √ a2 b − b2b ...1 2 x, to rewrite the equations as ze ≈ ab [ 1− 1 2 ( r bb )2 ] zc ≈ cr [ 1− 1 2 ( r cr )2 ] + ab − cr. or ze ≈ ab − 1 2 abr 2 b2b zc ≈ ab − 1 2 r2

Antilock braking system effectiveness in prevention of road traffic crashes in Iran

PubMed Central

2013-01-01

Background Anti-lock Brake System (ABS) helps the equipped vehicles to stop under heavy braking, in a shorter distance and with a better control of direction. It was expected that this technology will reduce the rate of fatal road traffic crashes (RTC); however, the outcome is controversial in the real world. The aim of this study is to compare the claimed annual incidence rate and financial losses due to RTCs in ABS vs. non-ABS personal vehicles in Iran. Methods A telephone survey among drivers of two similar models of personal vehicles was arranged. The studied vehicles were of the same brand and type; but only one of them was equipped with ABS. The number of RTCs, subsequent financial loss, and drivers’ knowledge and perception about ABS were sought. The sample consisted of drivers of 1232 ABS and 3123 non-ABS vehicles. Results The annual incidence rate of RTC involving another vehicle was 145.1 (134.8-155.9) per 1000 vehicle-years and there was not a statistically significant difference between study groups. The incidence of RTC with another vehicle due to brake failure was 50.3 (42.9-58.5) for 1000 non ABS vehicle-years and 30.0 (21.2-41.2) for 1000 ABS equipped vehicle-years. The difference was statistically significant after adjustment for the driver and vehicle's age and the daily driving time. The attributable risk of RTC for non-ABS vehicles was 20/1000 vehicles and the excess fraction was 39.8%. The mean financial loss due to reported RTCs was $987.9 ± $1547.3 US Dollars and there was not a statistically significant difference between study groups. While 61.1% of ABS vehicle drivers reported situations in which they believed the ABS had prevented a crash, 44.1% of them however, they did not know how to use ABS efficiently. Conclusions Law enforcement to maintain safe distance and adhere to speed limit while driving, is needed to raise the effectiveness of ABS. This is as necessary as considering mandatory outfitting of ABS. Safety authorities should first consider the global experience and local evidence, before adopting any specific policy in this regard. The drivers need to learn the right way to use ABS for maximum effectiveness. PMID:23642097

Antilock braking system effectiveness in prevention of road traffic crashes in Iran.

PubMed

Khorasani-Zavareh, Davoud; Shoar, Saeed; Saadat, Soheil

2013-05-04

Anti-lock Brake System (ABS) helps the equipped vehicles to stop under heavy braking, in a shorter distance and with a better control of direction. It was expected that this technology will reduce the rate of fatal road traffic crashes (RTC); however, the outcome is controversial in the real world. The aim of this study is to compare the claimed annual incidence rate and financial losses due to RTCs in ABS vs. non-ABS personal vehicles in Iran. A telephone survey among drivers of two similar models of personal vehicles was arranged. The studied vehicles were of the same brand and type; but only one of them was equipped with ABS. The number of RTCs, subsequent financial loss, and drivers' knowledge and perception about ABS were sought. The sample consisted of drivers of 1232 ABS and 3123 non-ABS vehicles. The annual incidence rate of RTC involving another vehicle was 145.1 (134.8-155.9) per 1000 vehicle-years and there was not a statistically significant difference between study groups.The incidence of RTC with another vehicle due to brake failure was 50.3 (42.9-58.5) for 1000 non ABS vehicle-years and 30.0 (21.2-41.2) for 1000 ABS equipped vehicle-years. The difference was statistically significant after adjustment for the driver and vehicle's age and the daily driving time. The attributable risk of RTC for non-ABS vehicles was 20/1000 vehicles and the excess fraction was 39.8%. The mean financial loss due to reported RTCs was $987.9 ± $1547.3 US Dollars and there was not a statistically significant difference between study groups. While 61.1% of ABS vehicle drivers reported situations in which they believed the ABS had prevented a crash, 44.1% of them however, they did not know how to use ABS efficiently. Law enforcement to maintain safe distance and adhere to speed limit while driving, is needed to raise the effectiveness of ABS. This is as necessary as considering mandatory outfitting of ABS. Safety authorities should first consider the global experience and local evidence, before adopting any specific policy in this regard. The drivers need to learn the right way to use ABS for maximum effectiveness.

Antitumour effects of single or combined monoclonal antibodies directed against membrane antigens expressed by human B cells leukaemia

PubMed Central

2011-01-01

Background The increasing availability of different monoclonal antibodies (mAbs) opens the way to more specific biologic therapy of cancer patients. However, despite the significant success of therapy in breast and ovarian carcinomas with anti-HER2 mAbs as well as in non-Hodkin B cell lymphomas with anti-CD20 mAbs, certain B cell malignancies such as B chronic lymphocytic leukaemia (B-CLL) respond poorly to anti-CD20 mAb, due to the low surface expression of this molecule. Thus, new mAbs adapted to each types of tumour will help to develop personalised mAb treatment. To this aim, we analyse the biological and therapeutic properties of three mAbs directed against the CD5, CD71 or HLA-DR molecules highly expressed on B-CLL cells. Results The three mAbs, after purification and radiolabelling demonstrated high and specific binding capacity to various human leukaemia target cells. Further in vitro analysis showed that mAb anti-CD5 induced neither growth inhibition nor apoptosis, mAb anti-CD71 induced proliferation inhibition with no early sign of cell death and mAb anti-HLA-DR induced specific cell aggregation, but without evidence of apoptosis. All three mAbs induced various degrees of ADCC by NK cells, as well as phagocytosis by macrophages. Only the anti-HLA-DR mAb induced complement mediated lysis. Coincubation of different pairs of mAbs did not significantly modify the in vitro results. In contrast with these discrete and heterogeneous in vitro effects, in vivo the three mAbs demonstrated marked anti-tumour efficacy and prolongation of mice survival in two models of SCID mice, grafted either intraperitoneally or intravenously with the CD5 transfected JOK1-5.3 cells. This cell line was derived from a human hairy cell leukaemia, a type of malignancy known to have very similar biological properties as the B-CLL, whose cells constitutively express CD5. Interestingly, the combined injection of anti-CD5 with anti-HLA-DR or with anti-CD71 led to longer mouse survival, as compared to single mAb injection, up to complete inhibition of tumour growth in 100% mice treated with both anti-HLA-DR and anti-CD5. Conclusions Altogether these data suggest that the combined use of two mAbs, such as anti-HLA-DR and anti-CD5, may significantly enhance their therapeutic potential. PMID:21504579

Enhancing antibody patent protection using epitope mapping information

PubMed Central

Deng, Xiaoxiang; Storz, Ulrich; Doranz, Benjamin J.

2018-01-01

ABSTRACT As the $100B therapeutic monoclonal antibody (mAb) market continues to grow, developers of therapeutic mAbs increasingly face the need to strengthen patent protection of their products and enforce their patents in courts. In view of changes in the patent law landscape, patent applications are strategically using information on the precise binding sites of their mAbs, i.e., the epitopes, to support patent novelty, non-obviousness, subject matter, and a tightened written description requirement for broad genus antibody claims. Epitope data can also allow freedom-to-operate for second-generation mAbs by differentiation from patented first-generation mAbs. Numerous high profile court cases, including Amgen v. Sanofi over rival mAbs that block PCSK9 activity, have been centered on epitope mapping claims, highlighting the importance of epitopes in determining broad mAb patent rights. Based on these cases, epitope mapping claims must describe a sufficiently large number of mAbs that share an epitope, and each epitope must be described at amino acid resolution. Here, we review current best practices for the use of epitope information to overcome the increasing challenges of patenting mAbs, and how the quality, conformation, and resolution of epitope residue data can influence the breadth and strength of mAb patents. PMID:29120697

Understanding Youth Antisocial Behavior Using Neuroscience through a Developmental Psychopathology Lens: Review, Integration, and Directions for Research

PubMed Central

Hyde, Luke W.; Shaw, Daniel S.; Hariri, Ahmad R.

2013-01-01

Youth antisocial behavior (AB) is an important public health concern impacting perpetrators, victims, and society. Functional neuroimaging is becoming a more common and useful modality for understanding neural correlates of youth AB. Although there has been a recent increase in neuroimaging studies of youth AB and corresponding theoretical articles on the neurobiology of AB, there has been little work critically examining the strengths and weaknesses of individual studies and using this knowledge to inform the design of future studies. Additionally, research on neuroimaging and youth AB has not been integrated within the broader framework of developmental psychopathology. Thus, this paper provides an in-depth review of the youth AB functional neuroimaging literature with the following goals: 1. to evaluate how this literature has informed our understanding of youth AB, 2. to evaluate current neuroimaging studies of youth AB from a developmental psychopathology perspective with a focus on integrating research from neuroscience and developmental psychopathology, as well as placing this research in the context of other related areas (e.g., psychopathy, molecular genetics), and 3. to examine strengths and weaknesses of neuroimaging and behavioral studies of youth AB to suggest how future studies can develop a more informed and integrated understanding of youth AB. PMID:24273368

Revealing strategies of quorum sensing in Azospirillum brasilense strains Ab-V5 and Ab-V6.

PubMed

Fukami, Josiane; Abrantes, Julia Laura Fernandes; Del Cerro, Pablo; Nogueira, Marco Antonio; Ollero, Francisco Javier; Megías, Manuel; Hungria, Mariangela

2018-01-01

Azospirillum brasilense is an important plant-growth promoting bacterium (PGPB) that requires several critical steps for root colonization, including biofilm and exopolysaccharide (EPS) synthesis and cell motility. In several bacteria these mechanisms are mediated by quorum sensing (QS) systems that regulate the expression of specific genes mediated by the autoinducers N-acyl-homoserine lactones (AHLs). We investigated QS mechanisms in strains Ab-V5 and Ab-V6 of A. brasilense, which are broadly used in commercial inoculants in Brazil. Neither of these strains carries a luxI gene, but there are several luxR solos that might perceive AHL molecules. By adding external AHLs we verified that biofilm and EPS production and cell motility (swimming and swarming) were regulated via QS in Ab-V5, but not in Ab-V6. Differences were observed not only between strains, but also in the specificity of LuxR-type receptors to AHL molecules. However, Ab-V6 was outstanding in indole acetic acid (IAA) synthesis and this molecule might mimic AHL signals. We also applied the quorum quenching (QQ) strategy, obtaining transconjugants of Ab-V5 and Ab-V6 carrying a plasmid with acyl-homoserine lactonase. When maize (Zea mays L.) was inoculated with the wild-type and transconjugant strains, plant growth was decreased with the transconjugant of Ab-V5-confirming the importance of an AHL-mediated QS system-but did not affect plant growth promotion by Ab-V6.

Synthesis of a novel polyamidoamine dendrimer conjugating with alkali blue as a lymphatic tracer and study on the lymphatic targeting in vivo.

PubMed

Yang, Rui; Xia, Suxia; Ye, Tiantian; Yao, Jianhua; Zhang, Ruizhi; Wang, Shujun; Wang, Siling

2016-09-01

In this study, a novel lymphatic tracer polyamidoamin-alkali blue (PAMAM-AB) was synthesized in order to evaluate the intra-lymphatic targeting ability and lymphatic tropism of PAMAM-AB after subcutaneous administration. UV-Vis, FT-IR, NMR and HPLC characterization were performed to prove the successful synthesis of PAMAM-AB. The calculated AB payload of PAMAM-AB conjugate was seven per dendrimer molecule (27.16% by weight). Hydrolysis stability of PAMAM-AB in vitro was evaluated, which was stable in PBS and human plasma. Lymphatic tracing were studied to determine the blue-stained intensity of PAMAM-AB in right popliteral lymph nodes (PLNs), iliac lymph nodes (ILNs) and para-aortic lymph nodes (PALNs) after subcutaneous administration. The pharmacokinetics and biodistribution of PAMAM-AB in mice were investigated. PLNs, ILNs and PALNs could be obviously blue-stained within 10 min after PAMAM-AB administration, and displayed a more rapid lymphatic absorption, a higher AUC value in lymph nodes and a longer lymph nodes residence time compared with methylene blue solution (MB-S), MB water-in-oil microemulsion (MB-ME), MB multiple microemulsion (MB-MME). Enhanced lymphatic drainage from the injection site and uptake into lymph of PAMAM-AB indicated that PAMAM-AB possesses the double function of lymphatic tracing and lymphatic targeting, and suggested the potential for the development of lymphatic targeting vectors or as a lymphatic tracer in its own right.

DR5 mAb-conjugated, DTIC-loaded immuno-nanoparticles effectively and specifically kill malignant melanoma cells in vivo.

PubMed

Ding, Baoyue; Zhang, Wei; Wu, Xin; Wang, Jeffrey; Xie, Chen; Huang, Xuan; Zhan, Shuyu; Zheng, Yongxia; Huang, Yueyan; Xu, Ningyin; Ding, Xueying; Gao, Shen

2016-08-30

We combined chemo- and immunotherapies by constructing dual therapeutic function immuno-nanoparticles (NPs) consisting of death receptor 5 monoclonal antibody (DR5 mAb)-conjugated nanoparticles loaded with dacarbazine (DTIC) (DTIC-NPs-DR5 mAb). We determined the in vivo targeting specificity of DTIC-NPs-DR5 mAb by evaluating distribution in tumor-bearing nude mice using a real-time imaging system. Therapeutic efficacy was assessed in terms of its effect on tumor volume, survival time, histomorphology, microvessel density (MVD), and apoptotic index (AI). Systemic toxicity was evaluated by measuring white blood cells (WBC) counts, alanine aminotransferase (ALT) levels, and creatinine clearance (CR).In vivo and ex vivo imaging indicates that DR5 mAb modification enhanced the accumulation of NPs within the xenograft tumor. DTIC-NPs-DR5 mAb inhibited tumor growth more effectively than DTIC or DR5 mAb alone, indicating that combining DTIC and DR5 mAb through pharmaceutical engineering achieves a better therapeutic effect. Moreover, the toxicity of DTIC-NPs-DR5 mAb was much lower than that of DTIC, implying that DR5 mAb targeting reduces nonspecific uptake of DTIC into normal tissue and thus decreases toxic side effects. These results demonstrate that DTIC-NPs-DR5 mAb is a safe and effective nanoparticle formulation with the potential to improve the efficacy and specificity of melanoma treatment.

DR5 mAb-conjugated, DTIC-loaded immuno-nanoparticles effectively and specifically kill malignant melanoma cells in vivo

PubMed Central

Wang, Jeffrey; Xie, Chen; Huang, Xuan; Zhan, Shuyu; Zheng, Yongxia; Huang, Yueyan; Xu, Ningyin; Ding, Xueying; Gao, Shen

2016-01-01

We combined chemo- and immunotherapies by constructing dual therapeutic function immuno-nanoparticles (NPs) consisting of death receptor 5 monoclonal antibody (DR5 mAb)-conjugated nanoparticles loaded with dacarbazine (DTIC) (DTIC-NPs-DR5 mAb). We determined the in vivo targeting specificity of DTIC-NPs-DR5 mAb by evaluating distribution in tumor-bearing nude mice using a real-time imaging system. Therapeutic efficacy was assessed in terms of its effect on tumor volume, survival time, histomorphology, microvessel density (MVD), and apoptotic index (AI). Systemic toxicity was evaluated by measuring white blood cells (WBC) counts, alanine aminotransferase (ALT) levels, and creatinine clearance (CR).In vivo and ex vivo imaging indicates that DR5 mAb modification enhanced the accumulation of NPs within the xenograft tumor. DTIC-NPs-DR5 mAb inhibited tumor growth more effectively than DTIC or DR5 mAb alone, indicating that combining DTIC and DR5 mAb through pharmaceutical engineering achieves a better therapeutic effect. Moreover, the toxicity of DTIC-NPs-DR5 mAb was much lower than that of DTIC, implying that DR5 mAb targeting reduces nonspecific uptake of DTIC into normal tissue and thus decreases toxic side effects. These results demonstrate that DTIC-NPs-DR5 mAb is a safe and effective nanoparticle formulation with the potential to improve the efficacy and specificity of melanoma treatment. PMID:27494835

Fluorescence dye-based detection of mAb aggregates in CHO culture supernatants.

PubMed

Paul, Albert Jesuran; Schwab, Karen; Prokoph, Nina; Haas, Elena; Handrick, René; Hesse, Friedemann

2015-06-01

Product yields, efficacy, and safety of monoclonal antibodies (mAbs) are reduced by the formation of higher molecular weight aggregates during upstream processing. In-process characterization of mAb aggregate formation is a challenge since there is a lack of a fast detection method to identify mAb aggregates in cell culture. In this work, we present a rapid method to characterize mAb aggregate-containing Chinese hamster ovary (CHO) cell culture supernatants. The fluorescence dyes thioflavin T (ThT) and 4-4-bis-1-phenylamino-8-naphthalene sulfonate (Bis-ANS) enabled the detection of soluble as well as large mAb aggregates. Partial least square (PLS) regression models were used to evaluate the linearity of the dye-based mAb aggregate detection in buffer down to a mAb aggregate concentration of 2.4 μg mL(-1). Furthermore, mAb aggregates were detected in bioprocess medium using Bis-ANS and ThT. Dye binding to aggregates was stable for 60 min, making the method robust and reliable. Finally, the developed method using 10 μmol L(-1) Bis-ANS enabled discrimination between CHO cell culture supernatants containing different levels of mAb aggregates. The method can be adapted for high-throughput screening, e.g., to screen for cell culture conditions influencing mAb product quality, and hence can contribute to the improvement of production processes of biopharmaceuticals in mammalian cell culture.

Antibiotic stewardship through the EU project "ABS International".

PubMed

Allerberger, Franz; Frank, Annegret; Gareis, Roland

2008-01-01

The increasing problem of antimicrobial resistance requires implementation of antibiotic stewardship (ABS) programs. The project "ABS International--implementing antibiotic strategies for appropriate use of antibiotics in hospitals in member states of the European Union" was started in September 2006 in Austria, Belgium, the Czech Republic, Germany, Hungary, Italy, Poland, Slovenia and Slovakia. A training program for national ABS trainers was prepared and standard templates for ABS tools (antibiotic list, guides for antibiotic treatment and surgical prophylaxis, antibiotic-related organization) and valid process measures, as well as quality indicators for antibiotic use were developed. Specific ABS tools are being implemented in up to five healthcare facilities in each country. Although ABS International clearly focuses on healthcare institutions, future antimicrobial stewardship programs must also cover public education and antibiotic prescribing in primary care.

Developmental relations between behavioral inhibition, anxiety, and attention biases to threat and positive information

PubMed Central

White, Lauren K.; Degnan, Kathryn A.; Henderson, Heather A.; Pérez-Edgar, Koraly; Walker, Olga L.; Shechner, Tomer; Leibenluft, Ellen; Bar-Haim, Yair; Pine, Daniel S.; Fox, Nathan A.

2016-01-01

The current study examined relations between behavioral inhibition (BI) assessed in toddlerhood (n=268) and attention biases (AB) to threat and positive faces and maternal reported anxiety assessed when children were 5 and 7-years old. Results revealed that BI predicted anxiety at age 7 in children with AB toward threat, away from positive, or with no bias, at age 7; BI did not predict anxiety for children displaying AB away from threat or toward positive. Five-year AB did not moderate the link between BI and 7-year anxiety. No direct association between AB and BI or anxiety was detected; moreover, children did not show stable AB across development. These findings extend our understanding of the developmental links between BI, AB, and anxiety. PMID:28042902

Characterization of virulence factor regulation by SrrAB, a two-component system in Staphylococcus aureus.

PubMed

Pragman, Alexa A; Yarwood, Jeremy M; Tripp, Timothy J; Schlievert, Patrick M

2004-04-01

Workers in our laboratory have previously identified the staphylococcal respiratory response AB (SrrAB), a Staphylococcus aureus two-component system that acts in the global regulation of virulence factors. This system down-regulates production of agr RNAIII, protein A, and toxic shock syndrome toxin 1 (TSST-1), particularly under low-oxygen conditions. In this study we investigated the localization and membrane orientation of SrrA and SrrB, transcription of the srrAB operon, the DNA-binding properties of SrrA, and the effect of SrrAB expression on S. aureus virulence. We found that SrrA is localized to the S. aureus cytoplasm, while SrrB is localized to the membrane and is properly oriented to function as a histidine kinase. srrAB has one transcriptional start site which results in either an srrA transcript or a full-length srrAB transcript; srrB must be cotranscribed with srrA. Gel shift assays of the agr P2, agr P3, protein A (spa), TSST-1 (tst), and srr promoters revealed SrrA binding at each of these promoters. Analysis of SrrAB-overexpressing strains by using the rabbit model of bacterial endocarditis demonstrated that overexpression of SrrAB decreased the virulence of the organisms compared to the virulence of isogenic strains that do not overexpress SrrAB. We concluded that SrrAB is properly localized and oriented to function as a two-component system. Overexpression of SrrAB, which represses agr RNAIII, TSST-1, and protein A in vitro, decreases virulence in the rabbit endocarditis model. Repression of these virulence factors is likely due to a direct interaction between SrrA and the agr, tst, and spa promoters.

Serum p53 antibody as a potential tumor marker in extrahepatic cholangiocarcinoma.

PubMed

Okada, Rei; Shimada, Hideaki; Otsuka, Yuichiro; Tsuchiya, Masaru; Ishii, Jun; Katagiri, Toshio; Maeda, Tetsuya; Kubota, Yoshihisa; Nemoto, Tetsuo; Kaneko, Hironori

2017-12-01

Only a few studies have evaluated the clinicopathological significance of the p53 protein expression and s-p53-Abs level in patients with cholangiocarcinoma. We therefore analyzed the clinicopathological and prognostic significance of s-p53-Abs in patients with extrahepatic cholangiocarcinoma. We prospectively evaluated s-p53-Abs levels before and after surgery in 61 patients with extrahepatic cholangiocarcinoma to determine the relationship between clinicopathological factors and the prognostic significance of s-p53-Abs. Among a total of 61 primary extrahepatic cholangiocarcinoma cases, 23% were positive for s-p53-Abs. Combination of s-p53-Abs with the conventional serum markers carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) significantly increased the rate of positive extrahepatic cholangiocarcinoma cases (57% for CEA and/or CA19-9 vs. 75% for CEA and/or CA19-9 and/or s-p53-Abs, P = 0.035). There were no significant differences in clinicopathological factors between the p53-seropositive and p53-seronegative patients. An immunohistochemical analysis showed the presence of significant associations between the intensity (P = 0.003) and extent (P = 0.001) of p53 immunoreactivity and p53-seropositivitly. Although s-p53-Abs was not a significant prognostic factor for the survival in either univariate or multivariate analyses, p53 immunoreactivity was independently associated with a poor survival. Among patients positive for s-p53-Abs before surgery, the s-p53-Abs levels were reduced after surgery in most. These findings suggested that s-p53-Abs might be associated with p53 immunoreactivity. In addition, s-p53-Abs may be useful for a diagnosis, but was not useful for predicting tumor recurrence or the survival. This study was registered as UMIN000014530.

Utility of Bronchoalveolar Lavage for the Diagnosis of Asbestos-Related Diseases.

PubMed

Cruz, María Jesús; Curull, Victor; Pijuan, Lara; Álvarez-Simón, Daniel; Sánchez-Font, Albert; de Gracia, Javier; Culebras, Mario; Ferrer, Jaume

2017-06-01

Bronchoalveolar lavage (BAL) analysis has been proposed as an objective technique for confirming asbestos exposure. However, the reliability and diagnostic yield of this procedure has not been studied in Spain. The aim of this study was to assess the usefulness of the analysis of asbestos bodies (AB) in bronchoalveolar lavage (BAL) for the diagnosis of asbestos-related diseases (ARD). BAL samples from 72 patients (66 male, mean age 66 years) undergoing bronchoscopy were analyzed. Lung tissue from 23 of these patients was also analyzed. Asbestos exposure was assessed by anamnesis and a review of the patient's medical records. BAL and lung samples were processed and AB count was determined by light microscopy. The accepted threshold value to diagnose asbestos-related diseases was 1 AB/ml BAL or 1000 AB/gr dry tissue. Thirty-nine patients reported exposure to asbestos. Of these, 13 (33%) presented AB values above 1 AB/ml BAL. In the 33 non-exposed patients, 5 (15%) presented AB values above 1 AB/ml BAL. There was a significant difference between the AB levels of exposed and non-exposed patients (P=.006). The ROC curve showed that a value of 0.5 AB/ml BAL achieved the most satisfactory sensitivity, 46%, and a specificity of 83%. The correlation between AB levels in BAL and lung was 0.633 (P=.002). BAL study provides objective evidence of exposure to asbestos. The good correlation between the AB counts in BAL and lung tissue indicates that both techniques are valid for the analysis of asbestos content. Copyright © 2016 SEPAR. Publicado por Elsevier España, S.L.U. All rights reserved.

The effect of arch height on kinematic coupling during walking.

PubMed

Wilken, Jason; Rao, Smita; Saltzman, Charles; Yack, H John

2011-03-01

The purpose of the current study was to assess kinematic coupling within the foot in individuals across a range of arch heights. Seventeen subjects participated in this study. Weight-bearing lateral radiographs were used to measure the arch height, defined as angle between the 1st metatarsal and the calcaneus. A kinematic model including the 1st metatarsal, lateral forefoot, calcaneus and tibia was used to assess foot kinematics during walking. Four coupling ratios were calculated: calcaneus frontal to forefoot transverse plane motion (Calcaneal EV/Forefoot AB), calcaneus frontal to transverse plane motion (Calcaneus EV/AB), forefoot sagittal to transverse plane motion (Forefoot DF/AB), and 1st metatarsal sagittal to transverse plane motion (1st Metatarsal DF/AB). Pearson product moment correlations were used to assess the relationship between arch height and coupling ratios. Mean (SD) radiographic arch angles of 129.8 (12.1) degrees with a range from 114 to 153 were noted, underscoring the range of arch heights in this cohort. Arch height explained approximately 3%, 38%, 12% and 1% of the variance in Calcaneal EV/Forefoot AB, Calcaneus EV/AB, Forefoot DF/AB and 1st Metatarsal DF/AB respectively. Calcaneal EV/Forefoot AB, Calcaneus EV/AB, Forefoot DF/AB and 1st Metatarsal DF/AB coupling ratios of 1.84 ± 0.80, 0.56 ± 0.35, 0.96 ± 0.27 and 0.43 ± 0.21 were noted, consistent with the twisted foot plate model, windlass mechanism and midtarsal locking mechanisms. Arch height had a small and modest relationship with kinematic coupling ratios during walking. Copyright © 2010 Elsevier Ltd. All rights reserved.

Aquaporin-4 antibody titration in NMO patients treated with rituximab: A retrospective study.

PubMed

Valentino, Paola; Marnetto, Fabiana; Granieri, Letizia; Capobianco, Marco; Bertolotto, Antonio

2017-03-01

We undertook an observational retrospective study to investigate the usefulness of aquaporin-4 (AQP4) antibodies (Ab) titration in the management of patients with neuromyelitis optica (NMO) treated with rituximab (RTX) by studying (1) the correlation between AQP4-Ab titer and disease activity, (2) the influence of RTX on antibody levels, and (3) the association between AQP4-Ab levels and responsiveness to RTX. A cell-based assay was used for AQP4-Ab titration in 322 serum samples from 7 patients with NMO treated with RTX (median follow-up 65 months), according to a treatment-to-target approach. Serum samples were collected every month following standardized procedures. (1) In group analysis, AQP4-Ab titers correlated with the disease activity, showing higher titers during and preceding relapses than during remission. However, in individual analysis, an increase in AQP4-Ab titers and CD19+ B cells did not always precede a relapse. (2) A reduction of AQP4-Ab titers in the short-term and long-term period was observed during RTX treatment. (3) Reduction of AQP4-Ab titers was observed in responder patients both 3 months after RTX infusion and in the long-term follow-up. In one nonresponder patient, AQP4-Ab levels never decreased during the treatment period. Titration of AQP4-Abs could be useful in the clinical management of patients with NMO treated with RTX: titration before each reinfusion and 3 months after each reinfusion may provide information about responsiveness to RTX. Although a relationship among AQP4-Ab levels, disease activity, and response to RTX was observed, the usefulness of AQP4-Ab titration to predict relapses is limited.

Influence of two barrier membranes on staged guided bone regeneration and osseointegration of titanium implants in dogs: part 1. Augmentation using bone graft substitutes and autogenous bone.

PubMed

Schwarz, Frank; Mihatovic, Ilja; Golubovic, Vladimir; Hegewald, Andrea; Becker, Jürgen

2012-01-01

To assess the influence of two barrier membranes and two bone graft substitutes mixed with autogenous bone (AB) on staged guided bone regeneration and osseointegration of titanium implants in dogs. Four saddle-type defects each were prepared in the upper jaw of six fox hounds and randomly filled with a natural bone mineral (NBM)+AB and a biphasic calcium phosphate (SBC)+AB and allocated to either an in situ gelling polyethylene glycol (PEG) or a collagen membrane (CM). At 8 weeks, modSLA titanium implants were inserted and left to heal in a submerged position. At 8+2 weeks, dissected blocks were processed for histomorphometrical analysis (e.g., treated area [TA], bone-to-implant contact [BIC]). The mean TA values (mm(2) ) and BIC values (%) tended to be higher in the PEG groups(TA: NBM+AB [10.4 ± 2.5]; SBC+AB [10.4 ± 5.8]/BIC: NBM+AB [86.4 ± 20.1]; SBC+AB [80.1 ± 21.5]) when compared with the corresponding CM groups (TA: NBM+AB [9.7 ± 4.8]; SBC+AB [7.8 ± 4.3]/BIC: NBM+AB [71.3 ± 20.8]; SBC+AB [72.4 ± 20.3]). A significant difference was observed for the mean TA values in the SBC+AB groups. It was concluded that all augmentation procedures investigated supported bone regeneration and staged osseointegration of modSLA titanium implants. However, the application of PEG may be associated with increased TA values. © 2011 John Wiley & Sons A/S.

The attentional blink is related to phonemic decoding, but not sight-word recognition, in typically reading adults.

PubMed

Tyson-Parry, Maree M; Sailah, Jessica; Boyes, Mark E; Badcock, Nicholas A

2015-10-01

This research investigated the relationship between the attentional blink (AB) and reading in typical adults. The AB is a deficit in the processing of the second of two rapidly presented targets when it occurs in close temporal proximity to the first target. Specifically, this experiment examined whether the AB was related to both phonological and sight-word reading abilities, and whether the relationship was mediated by accuracy on a single-target rapid serial visual processing task (single-target accuracy). Undergraduate university students completed a battery of tests measuring reading ability, non-verbal intelligence, and rapid automatised naming, in addition to rapid serial visual presentation tasks in which they were required to identify either two (AB task) or one (single target task) target/s (outlined shapes: circle, square, diamond, cross, and triangle) in a stream of random-dot distractors. The duration of the AB was related to phonological reading (n=41, β=-0.43): participants who exhibited longer ABs had poorer phonemic decoding skills. The AB was not related to sight-word reading. Single-target accuracy did not mediate the relationship between the AB and reading, but was significantly related to AB depth (non-linear fit, R(2)=.50): depth reflects the maximal cost in T2 reporting accuracy in the AB. The differential relationship between the AB and phonological versus sight-word reading implicates common resources used for phonemic decoding and target consolidation, which may be involved in cognitive control. The relationship between single-target accuracy and the AB is discussed in terms of cognitive preparation. Copyright © 2015 Elsevier Ltd. All rights reserved.

Resistance to Bacillus thuringiensis Toxin Cry2Ab in Trichoplusia ni Is Conferred by a Novel Genetic Mechanism

PubMed Central

Song, Xiaozhao; Kain, Wendy; Cassidy, Douglas

2015-01-01

The resistance to the Bacillus thuringiensis (Bt) toxin Cry2Ab in a greenhouse-originated Trichoplusia ni strain resistant to both Bt toxins Cry1Ac and Cry2Ab was characterized. Biological assays determined that the Cry2Ab resistance in the T. ni strain was a monogenic recessive trait independent of Cry1Ac resistance, and there existed no significant cross-resistance between Cry1Ac and Cry2Ab in T. ni. From the dual-toxin-resistant T. ni strain, a strain resistant to Cry2Ab only was isolated, and the Cry2Ab resistance trait was introgressed into a susceptible laboratory strain to facilitate comparative analysis of the Cry2Ab resistance with the susceptible T. ni strain. Results from biochemical analysis showed no significant difference between the Cry2Ab-resistant and -susceptible T. ni larvae in midgut proteases, including caseinolytic proteolytic activity and zymogram profile and serine protease activities, in midgut aminopeptidase and alkaline phosphatase activity, and in midgut esterases and hemolymph plasma melanization activity. For analysis of genetic linkage of Cry2Ab resistance with potential Cry toxin receptor genes, molecular markers for the midgut cadherin, alkaline phosphatase (ALP), and aminopeptidase N (APN) genes were identified between the original greenhouse-derived dual-toxin-resistant and the susceptible laboratory T. ni strains. Genetic linkage analysis showed that the Cry2Ab resistance in T. ni was not genetically associated with the midgut genes coding for the cadherin, ALP, and 6 APNs (APN1 to APN6) nor associated with the ABC transporter gene ABCC2. Therefore, the Cry2Ab resistance in T. ni is conferred by a novel but unknown genetic mechanism. PMID:26025894

Subtilase cytotoxin-coding genes in verotoxin-producing Escherichia coli strains from sheep and goats differ from those from cattle.

PubMed

Orden, José A; Horcajo, Pilar; de la Fuente, Ricardo; Ruiz-Santa-Quiteria, José A; Domínguez-Bernal, Gustavo; Carrión, Javier

2011-12-01

Subtilase cytotoxin (SubAB) from verotoxin (VT)-producing Escherichia coli (VTEC) strains was first described in the 98NK2 strain and has been associated with human disease. However, SubAB has recently been found in two VT-negative E. coli strains (ED 591 and ED 32). SubAB is encoded by two closely linked, cotranscribed genes (subA and subB). In this study, we investigated the presence of subAB genes in 52 VTEC strains isolated from cattle and 209 strains from small ruminants, using PCR. Most (91.9%) VTEC strains from sheep and goats and 25% of the strains from healthy cattle possessed subAB genes. The presence of subAB in a high percentage of the VTEC strains from small ruminants might increase the pathogenicity of these strains for human beings. Some differences in the results of PCRs and in the association with some virulence genes suggested the existence of different variants of subAB. We therefore sequenced the subA gene in 12 strains and showed that the subA gene in most of the subAB-positive VTEC strains from cattle was almost identical (about 99%) to that in the 98NK2 strain, while the subA gene in most of the subAB-positive VTEC strains from small ruminants was almost identical to that in the ED 591 strain. We propose the terms subAB1 to describe the SubAB-coding genes resembling that in the 98NK2 strain and subAB2 to describe those resembling that in the ED 591 strain.

Design and Validation of a Novel Generic Platform for the Production of Tetravalent IgG1-like Bispecific Antibodies.

PubMed

Golay, Josée; Choblet, Sylvie; Iwaszkiewicz, Justyna; Cérutti, Pierre; Ozil, Annick; Loisel, Séverine; Pugnière, Martine; Ubiali, Greta; Zoete, Vincent; Michielin, Olivier; Berthou, Christian; Kadouche, Jean; Mach, Jean-Pierre; Duonor-Cérutti, Martine

2016-04-01

We have designed and validated a novel generic platform for production of tetravalent IgG1-like chimeric bispecific Abs. The VH-CH1-hinge domains of mAb2 are fused through a peptidic linker to the N terminus of mAb1 H chain, and paired mutations at the CH1-CL interface mAb1 are introduced that force the correct pairing of the two different free L chains. Two different sets of these CH1-CL interface mutations, called CR3 and MUT4, were designed and tested, and prototypic bispecific Abs directed against CD5 and HLA-DR were produced (CD5xDR). Two different hinge sequences between mAb1 and mAb2 were also tested in the CD5xDR-CR3 or -MUT4 background, leading to bispecific Ab (BsAbs) with a more rigid or flexible structure. All four Abs produced bound with good specificity and affinity to CD5 and HLA-DR present either on the same target or on different cells. Indeed, the BsAbs were able to efficiently redirect killing of HLA-DR(+) leukemic cells by human CD5(+) cytokine-induced killer T cells. Finally, all BsAbs had a functional Fc, as shown by their capacity to activate human complement and NK cells and to mediate phagocytosis. CD5xDR-CR3 was chosen as the best format because it had overall the highest functional activity and was very stable in vitro in both neutral buffer and in serum. In vivo, CD5xDR-CR3 was shown to have significant therapeutic activity in a xenograft model of human leukemia. Copyright © 2016 by The American Association of Immunologists, Inc.

Thermal behavior of vehicle plastic blends contained acrylonitrile-butadiene-styrene (ABS) in pyrolysis using TG-FTIR.

PubMed

Liu, Guicai; Liao, Yanfen; Ma, Xiaoqian

2017-03-01

As important plastic blends in End-of-Life vehicles (ELV), pyrolysis profiles of ABS/PVC, ABS/PA6 and ABS/PC were investigated using thermogravimetric-Fourier transform infrared spectrometer (TG-FTIR). Also, CaCO 3 was added as plastic filler to discuss its effects on the pyrolysis of these plastics. The results showed that the interaction between ABS and PVC made PVC pyrolysis earlier and HCl emission slightly accelerated. The mixing of ABS and PA6 made their decomposition temperature closer, and ketones in PA6 pyrolysis products were reduced. The presence of ABS made PC pyrolysis earlier, and phenyl compounds in PC pyrolysis products could be transferred into alcohol or H 2 O. The interaction between ABS and other polymers in pyrolysis could be attributed to the intermolecular radical transfer, and free radicals from the polymer firstly decomposed led to a fast initiation the decomposition of the other polymer. As plastic filler, CaCO 3 promoted the thermal decomposition of PA6 and PC, and had no obvious effects on ABS and PVC pyrolysis process. Also, CaCO 3 made the pyrolysis products from PA6 and PC further decomposed into small-molecule compounds like CO 2 . The kinetics analysis showed that isoconversional method like Starink method was more suitable for these polymer blends. Starink method showed the average activation energy of ABS50/PVC50, ABS50/PA50 and ABS50/PC50 was 186.63kJ/mol, 239.61kJ/mol and 248.95kJ/mol, respectively, and the interaction among them could be reflected by the activation energy variation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Antibody response to actinomyces antigen and dental caries experience: implications for caries susceptibility.

PubMed

Levine, Martin; Owen, Willis L; Avery, Kevin T

2005-06-01

Fluoridated dentifrices reduce dental caries in subjects who perform effective oral hygiene. Actinomyces naeslundii increases in teeth-adherent microbial biofilms (plaques) in these subjects, and a well-characterized serum immunoglobulin G (IgG) antibody response (Actinomyces antibody [A-Ab]) is also increased. Other studies suggest that a serum IgG antibody response to streptococcal d-alanyl poly(glycerophosphate) (S-Ab) may indicate caries experience associated strongly with gingival health and exposure to fluoridated water. The aim of this study was to investigate relationships between A-Ab response, oral hygiene, S-Ab response, and caries experience. Measurements were made of A-Ab and S-Ab concentrations, caries experience (number of decayed, missing, and filled teeth [DMFT], number of teeth surfaces [DMFS], and number of decayed teeth needing treated [DT]), exposure to fluoridated water (Flu), mean clinical pocket depth (PD; in millimeters), and extent of plaque (PL) and gingival bleeding on probing (BOP). A-Ab concentration, the dependent variable in a multiple regression analysis, increased with S-Ab concentration and decreased with PL and DMFT adjusted for Flu (R(2) = 0.51, P < 0.002). Residual associations with age, DMFS, DT, and BOP were not significant. In addition, an elevated A-Ab response, defined from immunoprecipitation and immunoassay measurements, indicated a significant, 30% reduction in DMFT after adjustment for significant age and Flu covariance (analysis of variance with covariance F statistic = 10.6, P < 0.003; S-Ab response and interactions not significant). Thus, an elevated A-Ab response indicates less caries in subjects performing effective oral hygiene using fluoridated dentifrices. Conversely, a low A-Ab response is suggestive of decreased A. naeslundii binding to saliva-coated apatite and greater caries experience, as reported by others.

Mutational Analysis of the TnrA-Binding Sites in the Bacillus subtilis nrgAB and gabP Promoter Regions

PubMed Central

Wray, Lewis V.; Zalieckas, Jill M.; Ferson, Amy E.; Fisher, Susan H.

1998-01-01

Transcription of the Bacillus subtilis nrgAB promoter is activated during nitrogen-limited growth by the TnrA protein. A common inverted repeat, TGTNAN7TNACA (TnrA site), is centered 49 to 51 bp upstream of the transcriptional start sites for the TnrA-regulated nrgAB, gabP P2, and nas promoters. Oligonucleotide-directed mutagenesis of the nrgAB promoter region showed that conserved nucleotides within the TnrA site, the A+T-rich region between the two TnrA half-sites, and an upstream A tract are all required for high-level activation of nrgAB expression. Mutations that alter the relative distance between the two half-sites of the nrgAB TnrA site abolish nitrogen regulation of nrgAB expression. Spacer mutations that change the relative distance between the TnrA site and −35 region of the nrgAB promoter reveal that activation of nrgAB expression occurs only when the TnrA site is located 49 to 51 bp upstream of the transcriptional start site. Mutational analysis of the conserved nucleotides in the gabP P2 TnrA site showed that this sequence is also required for nitrogen-regulated gabP P2 expression. The TnrA protein, expressed in an overproducing Escherichia coli strain, had a 625-fold-higher affinity for the wild-type nrgAB promoter DNA than for a mutated nrgAB promoter DNA fragment that is unable to activate nrgAB expression in vivo. These results indicate that the proposed TnrA site functions as the binding site for the TnrA protein. TnrA was found to activate nrgAB expression during late exponential growth in nutrient sporulation medium containing glucose, suggesting that cells become nitrogen limited during growth in this medium. PMID:9603886

Humanized immunoglobulin mice: models for HIV vaccine testing and studying the broadly neutralizing antibody problem

PubMed Central

Verkoczy, Laurent

2018-01-01

A vaccine that can effectively prevent HIV-1 transmission remains paramount to ending the HIV pandemic, but to do so, will likely need to induce broad neutralizing antibody (bnAb) responses. A major technical hurdle towards achieving this goal has been a shortage of animal models with the ability to systematically pinpoint roadblocks to bnAb induction and to rank vaccine strategies based on their ability to stimulate bnAb development. Over the past six years, immunoglobulin (Ig) knock-in (KI) technology has been leveraged to express bnAbs in mice, an approach that has enabled elucidation of various B-cell tolerance mechanisms limiting bnAb production and evaluation of strategies to circumvent such processes. From these studies, in conjunction with the wealth of information recently obtained regarding the evolutionary pathways and paratopes/epitopes of multiple bnAbs, it has become clear that the very features of bnAbs desired for their function will be problematic to elicit by traditional vaccine paradigms, necessitating more iterative testing of new vaccine concepts. To meet this need, novel bnAb KI models have now been engineered to express either inferred pre-rearranged V(D)J exons (or unrearranged germline V, D, or J segments that can be assembled into functional rearranged V(D)J exons) encoding predecessors of mature bnAbs One encouraging approach that has materialized from studies using such newer models is sequential administration of immunogens designed to bind progressively more mature bnAb predecessors. In this review, insights into the regulation and induction of bnAbs based on the use of KI models will be discussed, as will new Ig KI approaches for higher-throughput production and/or altering expression of bnAbs in vivo, so as to further enable vaccine-guided bnAb induction studies. PMID:28413022

[Impact of an |A|B|S|-training initiative on |A|B|S|-structural quality of participating hospitals].

PubMed

Christoph, Anna; Ehm, Christine; de With, Katja

2015-01-01

The "ABS-training initiative" was funded by the German Ministry of Health as part of the German Antimicrobial Resistance Strategy (Deutsche Antibiotika-Resistenz-Strategie, DART) from 2009 until early 2014. The initiative was designed for clinicians and clinical pharmacists and contains several training units covering antiinfectives, infectious diseases and ABS strategies including the conduction of a research project at the participants' hospital. Participants who complete the four-weeks training initiative will become a certified "ABS Expert". 281 ABS Experts were asked to take part in a survey (staff for ABS, surveillance data about agents and consumption, ABS activity) to estimate the influence of the ABS-training initiative on the ABS-structural quality. The evaluation was performed using GrafStat (V 4.255), statistical software package for the evaluation of surveys. Ninety-two ABS Experts representing 92 hospitals participated in a questionnaire-based survey before and after completing the training initiative. Forty (44 %) hospitals appointed an ABS representative (+22 %) after completing the training initiative. Antibiotic surveillance data available as a report increased from 34 (40 %) to 54 (60 %) and correct data presentation (DDD or RDD/100 days) from 7 (8 %) to 40 (43 %). Proactive auditing of antiinfective prescribing improved from 54 (60 %) to 71 (78 %) in intensive care units, and from 28 (31 %) to 53 (58 %) on normal wards. Availability of local guidelines increased from 36 (39 %) to 52 (57 %). The "ABS Training Initiative" had a positive impact on ABS-structural quality regarding nomination of ABS-teams, surveillance data of antibiotic consumption, implementation of proactive auditing of antiinfective prescribing and availability of local guidelines. However, there is optimization potential in many sectors. The short time period between pre- and post-assessment and the ongoing personnel or time constraints need to be taken into account. Copyright © 2015. Published by Elsevier GmbH.

Comparison of F(ab')2 versus Fab antivenom for pit viper envenomation: A prospective, blinded, multicenter, randomized clinical trial

PubMed Central

Ruha, Anne-Michelle; Seifert, Steven A.; Morgan, David L.; Lewis, Brandon J.; Arnold, Thomas C.; Clark, Richard F.; Meggs, William J.; Toschlog, Eric A.; Borron, Stephen W.; Figge, Gary R.; Sollee, Dawn R.; Shirazi, Farshad M.; Wolk, Robert; de Chazal, Ives; Quan, Dan; García-Ubbelohde, Walter; Alagón, Alejandro; Gerkin, Richard D.; Boyer, Leslie V.

2015-01-01

Background. Crotalidae Polyvalent Immune Fab (Ovine) has been the only antivenom commercially available in the US since 2007 for treatment of Crotalinae envenomation. Late coagulopathy can occur or recur after clearance of Fab antivenom, often after hospital discharge, lasting in some cases more than 2 weeks. There have been serious, even fatal, bleeding complications associated with recurrence phenomena. Frequent follow-up is required, and additional intervention or hospitalization is often necessary. F(ab')2 immunoglobulin derivatives have longer plasma half life than do Fab. We hypothesized that F(ab')2 antivenom would be superior to Fab in the prevention of late coagulopathy following treatment of patients with Crotalinae envenomation. Methods. We conducted a prospective, double-blind, randomized clinical trial, comparing late coagulopathy in snakebitten patients treated with F(ab')2 with maintenance doses [F(ab')2/F(ab')2], or F(ab')2 with placebo maintenance doses [F(ab')2/placebo], versus Fab with maintenance doses [Fab/Fab]. The primary efficacy endpoint was coagulopathy (platelet count < 150 K/mm3, fibrinogen level < 150 mg/dL) between end of maintenance dosing and day 8. Results. 121 patients were randomized at 18 clinical sites and received at least one dose of study drug. 114 completed the study. Of these, 11/37 (29.7%) in the Fab/Fab cohort experienced late coagulopathy versus 4/39 (10.3%, p < 0.05) in the F(ab')2/F(ab')2 cohort and 2/38 (5.3%, p < 0.05) in the F(ab')2/placebo cohort. The lowest heterologous protein exposure was with F(ab')2/placebo. No serious adverse events were related to study drug. In each study arm, one patient experienced an acute serum reaction and one experienced serum sickness. Conclusions. In this study, management of coagulopathic Crotalinae envenomation with longer-half-life F(ab')2 antivenom, with or without maintenance dosing, reduced the risk of subacute coagulopathy and bleeding following treatment of envenomation. PMID:25361165

Peptide-based Antibodies against Glutathione-binding Domains Suppress Superoxide Production Mediated by Mitochondrial Complex I*

PubMed Central

Chen, Jingfeng; Chen, Chwen-Lih; Rawale, Sharad; Chen, Chun-An; Zweier, Jay L.; Kaumaya, Pravin T. P.; Chen, Yeong-Renn

2010-01-01

Complex I (NQR) is a critical site of superoxide () production and the major host of redox protein thiols in mitochondria. In response to oxidative stress, NQR-derived protein thiols at the 51- and 75-kDa subunits are known to be reversibly S-glutathionylated. Although several glutathionylated domains from NQR 51 and 75 kDa have been identified, their roles in the regulatory functions remain to be explored. To gain further insights into protein S-glutathionylation of complex I, we used two peptides of S-glutathionylated domain (200GAGAYICGEETALIESIEGK219 of 51-kDa protein and 361VDSDTLCTEEVFPTAGAGTDLR382 of 75-kDa protein) as chimeric epitopes incorporating a “promiscuous” T-cell epitope to generate two polyclonal antibodies, AbGSCA206 and AbGSCB367. Binding of AbGSCA206 and AbGSCB367 inhibited NQR-mediated generation by 37 and 57%, as measured by EPR spin-trapping. To further provide an appropriate control, two peptides of non-glutathionylated domain (21SGDTTAPKKTSFGSLKDFDR40 of 51-kDa peptide and 100WNILTNSEKTKKAREGVMEFL120 of 75-kDa peptide) were synthesized as chimeric epitopes to generate two polyclonal antibodies, Ab51 and Ab75. Binding of A51 did not affect NQR-mediated generation to a significant level. However, binding of Ab75 inhibited NQR-mediated generation by 35%. None of AbGSCA206, AbGSCB367, Ab51, or Ab75 showed an inhibitory effect on the electron transfer activity of NQR, suggesting that antibody binding to the glutathione-binding domain decreased electron leakage from the hydrophilic domain of NQR. When heart tissue homogenates were immunoprecipitated with Ab51 or Ab75 and probed with an antibody against glutathione, protein S-glutathionylation was enhanced in post-ischemic myocardium at the NQR 51-kDa subunit, but not at the 75-kDa subunit, indicating that the 51-kDa subunit of flavin subcomplex is more sensitive to oxidative stress resulting from myocardial infarction. PMID:19940158

Investigations of immunogenic, allergenic and adjuvant properties of Cry1Ab protein after intragastric exposure in a food allergy model in mice.

PubMed

Andreassen, Monica; Bøhn, Thomas; Wikmark, Odd-Gunnar; Bodin, Johanna; Traavik, Terje; Løvik, Martinus; Nygaard, Unni Cecilie

2016-05-04

In genetically modified (GM) crops there is a risk that the inserted genes may introduce new allergens and/or adjuvants into the food and feed chain. The MON810 maize, expressing the insecticidal Cry1Ab toxin, is grown in many countries worldwide. In animal models, intranasal and intraperitoneal immunisations with the purified Cry1Ab proteins have induced immune responses, and feeding trials with Cry1Ab-containing feed have revealed some altered immune responses. Previous investigations have primarily measured antibody responses to the protein, while investigations of clinical food allergy symptoms, or allergy promotion (adjuvant effect) associated with the Cry1Ab protein are largely missing. We aimed to investigate immunogenic, allergenic and adjuvant properties of purified Cry1Ab toxin (trypCry1Ab, i.e., trypsin activated Cry1Ab) in a mouse model of food allergy. Female C3H/HeJ mice were immunized by intragastric gavage of 10 μg purified, trypsin activated Cry1Ab toxin (trypCry1Ab) alone or together with the food allergen lupin. Cholera toxin was added as a positive control for adjuvant effect to break oral tolerance. Clinical symptoms (anaphylaxis) as well as humoral and cellular responses were assessed. In contrast to results from previous airway investigations, we observed no indication of immunogenic properties of trypCry1Ab protein after repeated intragastric exposures to one dose, with or without CT as adjuvant. Moreover, the results indicated that trypCry1Ab given by the intragastric route was not able to promote allergic responses or anaphylactic reactions against the co-administered allergen lupin at the given dose. The study suggests no immunogenic, allergenic or adjuvant capacity of the given dose of trypCry1Ab protein after intragastric exposure of prime aged mice.

Family distribution of anti-F(ab')2 antibodies in relatives of patients with systemic lupus erythematosus.

PubMed Central

Silvestris, F; Searles, R P; Bankhurst, A D; Williams, R C

1985-01-01

Recently we reported an inverse relationship between the levels of anti-F(ab')2 antibodies and disease activity in systemic lupus erythematosus (SLE). The present study focused on anti-F(ab')2 antibodies in unaffected relatives of SLE patients. Sixty sera from first degree family members from 11 SLE families and 49 sera from 8 control families were studied. Percentage of SLE family members with anti-DNA antibodies (15%) was higher than than control family sera (8%, P less than 0.05). Anti-F(ab')2 antibodies were measured using ELISA assays. The SLE family sera had higher amounts of anti-F(ab')2 antibodies than the normal control family group (P = 0.0051). In an effort to determine if anti-F(ab')2 antibodies found in high titres in the sera of some SLE family members had specificity for the F(ab')2 fragment of anti-DNA antibodies of the SLE relative patients, DNA-anti-DNA inhibition experiments were performed using anti-F(ab')2 prepared from the relative in parallel with anti-F(ab')2 prepared from normal controls with equivalent high titres of serum anti-F(ab')2. Inhibition exhibited by anti-F(ab')2 of first degree relatives was higher than that obtained from control normal donors (P less than 0.02). Such differences in inhibition were not recorded using a control tetanus toxoid-anti-tetanus toxoid assay. In direct binding ELISA experiments, peroxidase-conjugated anti-F(ab')2 antibodies from the same first degree relative showed high relative specificity against purified anti-DNA antibodies of his SLE proband when compared to those obtained against different anti-DNA antibodies isolated from unrelated SLE patients (P less than 0.001). Such a substantial difference was not observed in parallel experiments using peroxidase conjugated anti-F(ab')2 antibodies from normal controls unrelated to SLE subjects. PMID:3874025

Comparison of F(ab')2 versus Fab antivenom for pit viper envenomation: a prospective, blinded, multicenter, randomized clinical trial.

PubMed

Bush, Sean P; Ruha, Anne-Michelle; Seifert, Steven A; Morgan, David L; Lewis, Brandon J; Arnold, Thomas C; Clark, Richard F; Meggs, William J; Toschlog, Eric A; Borron, Stephen W; Figge, Gary R; Sollee, Dawn R; Shirazi, Farshad M; Wolk, Robert; de Chazal, Ives; Quan, Dan; García-Ubbelohde, Walter; Alagón, Alejandro; Gerkin, Richard D; Boyer, Leslie V

2015-01-01

Crotalidae Polyvalent Immune Fab (Ovine) has been the only antivenom commercially available in the US since 2007 for treatment of Crotalinae envenomation. Late coagulopathy can occur or recur after clearance of Fab antivenom, often after hospital discharge, lasting in some cases more than 2 weeks. There have been serious, even fatal, bleeding complications associated with recurrence phenomena. Frequent follow-up is required, and additional intervention or hospitalization is often necessary. F(ab')2 immunoglobulin derivatives have longer plasma half life than do Fab. We hypothesized that F(ab')2 antivenom would be superior to Fab in the prevention of late coagulopathy following treatment of patients with Crotalinae envenomation. We conducted a prospective, double-blind, randomized clinical trial, comparing late coagulopathy in snakebitten patients treated with F(ab')2 with maintenance doses [F(ab')2/F(ab')2], or F(ab')2 with placebo maintenance doses [F(ab')2/placebo], versus Fab with maintenance doses [Fab/Fab]. The primary efficacy endpoint was coagulopathy (platelet count < 150 K/mm(3), fibrinogen level < 150 mg/dL) between end of maintenance dosing and day 8. 121 patients were randomized at 18 clinical sites and received at least one dose of study drug. 114 completed the study. Of these, 11/37 (29.7%) in the Fab/Fab cohort experienced late coagulopathy versus 4/39 (10.3%, p < 0.05) in the F(ab')2/F(ab')2 cohort and 2/38 (5.3%, p < 0.05) in the F(ab')2/placebo cohort. The lowest heterologous protein exposure was with F(ab')2/placebo. No serious adverse events were related to study drug. In each study arm, one patient experienced an acute serum reaction and one experienced serum sickness. In this study, management of coagulopathic Crotalinae envenomation with longer-half-life F(ab')2 antivenom, with or without maintenance dosing, reduced the risk of subacute coagulopathy and bleeding following treatment of envenomation.

IgY Antibodies Protect against Human Rotavirus Induced Diarrhea in the Neonatal Gnotobiotic Piglet Disease Model

PubMed Central

Vega, Celina G.; Bok, Marina; Vlasova, Anastasia N.; Chattha, Kuldeep S.; Fernández, Fernando M.; Wigdorovitz, Andrés; Parreño, Viviana G.; Saif, Linda J.

2012-01-01

Group A Rotaviruses are the most common cause of severe, dehydrating diarrhea in children worldwide. The aim of the present work was to evaluate protection against rotavirus (RV) diarrhea conferred by the prophylactic administration of specific IgY antibodies (Ab) to gnotobiotic piglets experimentally inoculated with virulent Wa G1P[8] human rotavirus (HRV). Chicken egg yolk IgY Ab generated from Wa HRV hyperimmunized hens specifically recognized (ELISA) and neutralized Wa HRV in vitro. Supplementation of the RV Ab free cow milk diet with Wa HRV-specific egg yolk IgY Ab at a final ELISA Ab titer of 4096 (virus neutralization –VN- titer = 256) for 9 days conferred full protection against Wa HRV associated diarrhea and significantly reduced virus shedding. This protection was dose-dependent. The oral administration of semi-purified passive IgY Abs from chickens did not affect the isotype profile of the pig Ab secreting cell (ASC) responses to Wa HRV infection, but it was associated with significantly fewer numbers of HRV–specific IgA ASC in the duodenum. We further analyzed the pigś immune responses to the passive IgY treatment. The oral administration of IgY Abs induced IgG Ab responses to chicken IgY in serum and local IgA and IgG Ab responses to IgY in the intestinal contents of neonatal piglets in a dose dependent manner. To our knowledge, this is the first study to show that IgY Abs administered orally as a milk supplement passively protect neonatal pigs against an enteric viral pathogen (HRV). Piglets are an animal model with a gastrointestinal physiology and an immune system that closely mimic human infants. This strategy can be scaled-up to inexpensively produce large amounts of polyclonal IgY Abs from egg yolks to be applied as a preventive and therapeutic passive Ab treatment to control RV diarrhea. PMID:22880110

Word frequency and the attentional blink: the effects of target difficulty on retrieval and consolidation processes.

PubMed

Wierda, Stefan M; Taatgen, Niels A; van Rijn, Hedderik; Martens, Sander

2013-01-01

When a second target (T2) is presented in close succession of a first target (T1) within a stream of non-targets, people often fail to detect T2-a deficit known as the attentional blink (AB). Two types of theories can be distinguished that have tried to account for this phenomenon. Whereas attentional-control theories suggest that protection of consolidation processes induces the AB, limited-resource theories claim that the AB is caused by a lack of resources. According to the latter type of theories, increasing difficulty of one or both targets should increase the magnitude of the AB. Similarly, attentional-control theories predict that a difficult T1 increases the AB due to prolonged processing. However, the prediction for T2 is not as straightforward. Prolonged processing of T2 could cause conflicts and increase the AB. However, if consolidation of T2 is postponed without loss of identity, the AB might be attenuated. Participants performed an AB task that consisted of a stream of distractor non-words and two target words. Difficulty of T1 and T2 was manipulated by varying word-frequency. Overall performance for high-frequency words was better than for low-frequency words. When T1 was highly frequent, the AB was reduced. The opposite effect was found for T2. When T2 was highly frequent, performance during the AB period was relatively worse than for a low-frequency T2. A threaded-cognition model of the AB was presented that simulated the observed pattern of behavior by taking changes in the time-course of retrieval and consolidation processes into account. Our results were replicated in a subsequent ERP study. The finding that a difficult low-frequency T2 reduces the magnitude of the AB is at odds with limited-resource accounts of the AB. However, it was successfully accounted for by the threaded-cognition model, thus providing an explanation in terms of attentional control.

In vitro and in vivo evaluation of osteoinductivity and bone fusion ability of an activin a/BMP2 chimera (AB204): a comparison study between AB204 and rhBMP-2.

PubMed

Zheng, Guang Bin; Lee, Jae Hyup; Jin, Yuan-Zhe

2017-12-01

This study compared osteoinductivity and osteogenic capacity between AB204 and rhBMP-2 using hMSCs in vitro and a beagle's posterolateral spinal fusion model. Cultured hMSCs were treated with AB204 or rhBMP-2 with low to high doses. Three male beagles were performed posterolateral spinal fusion with biphasic calcium phosphate (2 ml) + AB204 or rhBMP-2 (20, 50 or 200 µg). They were euthanized after 8 weeks. The fusion rate and bone formation of spine samples were examined. AB204 had higher alkaline phosphatase activity, mineralization and osteogenic-related gene expression than rhBMP-2. Fusion rates in all rhBMP-2 groups were 0. They were 100% for 50 μg and 200 μg AB204 groups. Therefore, AB204 showed higher osteogenicity than rhBMP-2. It could be a better bone graft substitute.

What Are the Primary Limitations in B-Cell Affinity Maturation, and How Much Affinity Maturation Can We Drive with Vaccination?

PubMed Central

2017-01-01

A key goal of HIV-1 vaccine development is the induction of broadly neutralizing antibodies (bnAbs) targeted to the vulnerable regions of the HIV envelope. BnAbs develop overtime in ∼50%of HIV-1-infected individuals. However, to date, no vaccines have induced bnAbs and few or none of these vaccine-elicited HIV-1 antibodies carry the high frequencies of V(D)J mutations characteristic of bnAbs. Do the high frequencies of mutations characteristic of naturally induced bnAbs represent a fundamental barrier to the induction of bnAbs by vaccines? Recent studies suggest that high frequencies of V(D)J mutations can be achieved by serial vaccination strategies. Rather, it appears that, in the absence of HIV-1 infection, physiologic immune tolerance controls, including a germinal center process termed affinity reversion, may limit vaccine-driven bnAb development by clonal elimination or selecting for mutations incompatible with bnAb activity. PMID:28630077

[In vitro hepatic targeting tendency of galactosyl-anti CD3 McAb-TILS].

PubMed

Jiang, P; He, S; Zhang, C

1999-03-01

This study was undertaken to enhance the hepatic targeting tendency of tumor infiltrating lymphocytes (TILs) and hence to lower the recurrence rate of primary liver cancer after hepatectomy. Galactosyl-anti CD3McAb-TILs were prepared and then were incubated together with hepatocytes. Their interaction through asialoglycoprotein receptor-mediated mechanism was observed under the inverted phase contrast microscope. The carbohydrate density of galactosyl-anti CD3McAb and the combining rate of galactosyl-anti CD3McAb with TILs were measured. The results revealed that galactosyl-anti CD3 McAb-TIL obviously were adhered to hepatocytes. The carbohydrate density of galactosyl-anti CD3McAb was 62.18, and the combining rate of galactosyl-anti CD3McAb with TILs was 97.9%. The results suggested that in vitro hepatic targeting tendency of galactosyl-anti CD3McAb-TILs was satisfactory, and that the carbohydrate density of 62.18 and the combining rate of 97.9% ensured the effective use of TILs.

Long noncoding RNA AB074169 inhibits cell proliferation via modulation of KHSRP-mediated p21 expression in papillary thyroid carcinoma.

PubMed

Gou, Qiheng; Gao, Linbo; Nie, Xinwen; Pu, Wenchen; Zhu, Jingqiang; Wang, Yichao; Liu, Xuesha; Tan, Shuangyan; Zhou, Jian-Kang; Gong, Yanqiu; He, Juan; Wu, Ke; Xie, Yuxin; Zhao, Wanjun; Dai, Lunzhi; Liu, Lunxu; Xiang, Rong; Wei, Yu-Quan; Zhang, Lin; Peng, Yong

2018-05-07

Long noncoding RNAs (lncRNAs) are emerging as a novel class of regulators in gene expression associated with tumorigenesis. However, the role of lncRNAs in papillary thyroid carcinoma (PTC) is poorly understood. Here we conducted global lncRNA profiling and identified lncRNA AB074169 (lncAB) as significantly downregulated in PTC. Decreased expression of lncAB in PTC was caused by CpG hypermethylation within its gene promoter. Functional studies showed that lncAB overexpression led to cell cycle arrest and tumor growth inhibition in vitro and in vivo, whereas lncAB knockdown promoted cell proliferation. Mechanistic analyses revealed that lncAB bound KH-type splicing regulatory protein (KHSRP) and also decreased expression of KHSRP, thus increasing CDKN1a (p21) expression and decreasing CDK2 expression to repress cell proliferation. Taken together, these findings demonstrate that lncAB functions as a tumor suppressor during PTC tumorigenesis. Copyright ©2018, American Association for Cancer Research.

Functional Relevance of Improbable Antibody Mutations for HIV Broadly Neutralizing Antibody Development.

PubMed

Wiehe, Kevin; Bradley, Todd; Meyerhoff, R Ryan; Hart, Connor; Williams, Wilton B; Easterhoff, David; Faison, William J; Kepler, Thomas B; Saunders, Kevin O; Alam, S Munir; Bonsignori, Mattia; Haynes, Barton F

2018-06-13

HIV-1 broadly neutralizing antibodies (bnAbs) require high levels of activation-induced cytidine deaminase (AID)-catalyzed somatic mutations for optimal neutralization potency. Probable mutations occur at sites of frequent AID activity, while improbable mutations occur where AID activity is infrequent. One bottleneck for induction of bnAbs is the evolution of viral envelopes (Envs) that can select bnAb B cell receptors (BCR) with improbable mutations. Here we define the probability of bnAb mutations and demonstrate the functional significance of key improbable mutations in three bnAb B cell lineages. We show that bnAbs are enriched for improbable mutations, which implies that their elicitation will be critical for successful vaccine induction of potent bnAb B cell lineages. We discuss a mutation-guided vaccine strategy for identification of Envs that can select B cells with BCRs that have key improbable mutations required for bnAb development. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Monoclonal and anti-idiotypic anti-EBV/C3d receptor antibodies detect two binding sites, one for EBV and one for C3d on glycoprotein 140, the EBV/C3dR, expressed on human B lymphocytes.

PubMed

Barel, M; Fiandino, A; Delcayre, A X; Lyamani, F; Frade, R

1988-09-01

Glycoprotein (gp) 140, the EBV/C3dR of B lymphocytes, is a membrane site involved in human cell regulation. To analyze the specificities of the binding sites for EBV and for C3d on the gp 140 molecule, two distinct approaches were used. First, anti-EBV/C3dR mAb were prepared against highly purified EBV/C3dR. Nine anti-EBV/C3dR mAb were obtained. Four of these anti-EBV/C3dR mAb inhibited C3d binding but not EBV binding on gp 140, whereas four others exerted an inverse effect. These differences could not be due to differences in isotype, antibody concentration, affinity constant, and number of molecules bound on cell surface, as these parameters were identical for the nine used mAb. Second, polyclonal anti-idiotypic antibodies (Ab2) were prepared against F(ab)'2 fragments of polyclonal anti-EBV/C3dR (Ab1). Ab2 recognized the variable portion of Ab1 as controlled by immunoblotting experiments. Ab2, which did not react with the cell surface, inhibited Ab1 binding on Raji cells. Ab2 mimicked the EBV/C3dR by its properties to bind to particle-bound C3d and EBV, preventing their binding on Raji cell surface. C3d binding specificities contained in Ab2 were isolated by affinity chromatography on C3b/C3bi-Sepharose. These specificities, being the internal image of C3d binding site of EBV/C3dR, reacted with Ab1 and inhibited particle-bound C3d binding on Raji cells but did not react with EBV. Taken together, these data support strongly that gp 140, the EBV/C3dR, carried two distinct binding sites, one for EBV and one for C3d.

[Conformational Fingerprinting Using Monoclonal Antibodies
(on the Example of Angiotensin I-Converting Enzyme-ACE)].

PubMed

Danilov, S M

2017-01-01

During the past 30 years my laboratory has generated 40+ monoclonal antibodies (mAbs) directed to structural and conformational epitopes on human ACE as well as ACE from rats, mice and other species. These mAbs were successfully used for detection and quantification of ACE by ELISA, Western blotting, flow cytometry and immunohistochemistry. In all these applications mainly single mAbs were used. We hypothesized that we can obtain a completely new kind of information about ACE structure and function if we use the whole set of mAbs directed to different epitopes on the ACE molecule. When we finished epitope mapping of all mAbs to ACE (and especially, those recognizing conformational epitopes), we realized that we had obtained a new tool to study ACE. First, we demonstrated that binding of some mAbs is very sensitive to local conformational changes on the ACE surface-due to local denaturation, inactivation, ACE inhibitor or mAbs binding or due to diseases. Second, we were able to detect, localize and characterize several human ACE mutations. And, finally, we established a new concept - conformational fingerprinting of ACE using mAbs that in turn allowed us to obtain evidence for tissue specificity of ACE, which has promising scientific and diagnostic perspectives. The initial goal for the generation of mAbs to ACE 30 years ago was obtaining mAbs to organ-specific endothelial cells, which could be used for organ-specific drug delivery. Our systematic work on characterization of mAbs to numerous epitopes on ACE during these years has lead not only to the generation of the most effective mAbs for specific drug/gene delivery into the lung capillaries, but also to the establishment of the concept of conformational fingerprinting of ACE, which in turn gives a theoretical base for the generation of mAbs, specific for ACE from different organs. We believe that this concept could be applicable for any glycoprotein against which there is a set of mAbs to different epitopes.

Immunohistochemical detection of p53 protein in ameloblastoma types.

PubMed

el-Sissy, N A

1999-05-01

Overexpression of p53 protein in unicystic ameloblastoma (uAB) is denser than in the conventional ameloblastoma (cAB) type, indicating increased wild type p53--suppressing the growth potential of uAB and denoting the early event of neoplastic transformation, probably of a previous odontogenic cyst. Overexpression of p53 in borderline cAB and malignant ameloblastoma (mAB) types might reflect a mutational p53 protein playing an oncogenic role, promoting tumour growth. Overexpression of p53 protein could be a valid screening method for predicting underlying malignant genetic changes in AB types, through increased frequency of immunoreactive cells or increased staining density.

Diagnostic value of Tg and TgAb for metastasis following ablation in patients with differentiated thyroid carcinoma coexistent with Hashimoto thyroiditis.

PubMed

Chai, Hong; Zhu, Zhao-Jin; Chen, Ze-Quan; Yu, Yong-Li

2016-08-01

This study was designed to investigate the clinical value of serum thyroglobulin (Tg) and antithyroglobulin antibody (TgAb) measurements and the cutoff value after ablation in differentiated thyroid carcinoma (DTC) complicated by Hashimoto thyroiditis (HT) with metastasis. We measured serum Tg and TgAb levels and evaluated the disease status in 164 cases of DTC coexistent with HT in pathologically confirmed patients after surgery and post-remnant ablation during a 3-year follow-up. All Tg and TgAb levels were assessed by chemiluminescent immunoassay (IMA). Receiver operating characteristic (ROC) curve analysis was used to evaluate the prognostic value of Tg and TgAb for disease metastasis. The relationship between Tg and TgAb was analyzed using the scatter diagram distribution method. We found that the cutoff values of Tg and TgAb were 1.48 µg/L and 45 kIU/L, respectively. The area under the ROC curve (AUC) of Tg and TgAb was 0.907 and 0.650, respectively. In DTC coexistent with HT patients, the optimal cutoff value correlated with metastasis in Tg and TgAb was 1.48 µg/L and 45 kIU/L, respectively.

Multiple measures of dispositional global/local bias predict attentional blink magnitude.

PubMed

Dale, Gillian; Arnell, Karen M

2015-07-01

When the second of two targets (T2) is presented temporally close to the first target (T1) in a rapid serial visual presentation stream, accuracy to identify T2 is markedly reduced-an attentional blink (AB). While most individuals show an AB, Dale and Arnell (Atten Percept Psychophys 72(3):602-606, 2010) demonstrated that individual differences in dispositional attentional focus predicted AB performance, such that individuals who showed a natural bias toward the global level of Navon letter stimuli were less susceptible to the AB and showed a smaller AB effect. For the current study, we extended the findings of Dale and Arnell (Atten Percept Psychophys 72(3):602-606, 2010) through two experiments. In Experiment 1, we examined the relationship between dispositional global/local bias and the AB using a highly reliable hierarchical shape task measure. In Experiment 2, we examined whether three distinct global/local measures could predict AB performance. In both experiments, performance on the global/local tasks predicted subsequent AB performance, such that individuals with a greater preference for the global information showed a reduced AB. This supports previous findings, as well as recent models which discuss the role of attentional breadth in selective attention.

The adnAB Locus, Encoding a Putative Helicase-Nuclease Activity, Is Essential in Streptomyces

PubMed Central

Zhang, Lingli; Nguyen, Hoang Chuong; Chipot, Ludovic; Piotrowski, Emilie; Bertrand, Claire

2014-01-01

Homologous recombination is a crucial mechanism that repairs a wide range of DNA lesions, including the most deleterious ones, double-strand breaks (DSBs). This multistep process is initiated by the resection of the broken DNA ends by a multisubunit helicase-nuclease complex exemplified by Escherichia coli RecBCD, Bacillus subtilis AddAB, and newly discovered Mycobacterium tuberculosis AdnAB. Here we show that in Streptomyces, neither recBCD nor addAB homologues could be detected. The only putative helicase-nuclease-encoding genes identified were homologous to M. tuberculosis adnAB genes. These genes are conserved as a single copy in all sequenced genomes of Streptomyces. The disruption of adnAB in Streptomyces ambofaciens and Streptomyces coelicolor could not be achieved unless an ectopic copy was provided, indicating that adnAB is essential for growth. Both adnA and adnB genes were shown to be inducible in response to DNA damage (mitomycin C) and to be independently transcribed. Introduction of S. ambofaciens adnAB genes in an E. coli recB mutant restored viability and resistance to UV light, suggesting that Streptomyces AdnAB could be a functional homologue of RecBCD and be involved in DNA damage resistance. PMID:24837284

Differential induction of anti-V3 crown antibodies with cradle- and ladle-binding modes in response to HIV-1 envelope vaccination.

PubMed

Balasubramanian, Preetha; Kumar, Rajnish; Williams, Constance; Itri, Vincenza; Wang, Shixia; Lu, Shan; Hessell, Ann J; Haigwood, Nancy L; Sinangil, Faruk; Higgins, Keith W; Liu, Lily; Li, Liuzhe; Nyambi, Phillipe; Gorny, Miroslaw K; Totrov, Maxim; Nadas, Arthur; Kong, Xiang-Peng; Zolla-Pazner, Susan; Hioe, Catarina E

2017-03-07

The V3 loop in the HIV envelope gp120 is one of the immunogenic sites targeted by Abs. The V3 crown in particular has conserved structural elements recognized by cross-reactive neutralizing Abs, indicating its potential contribution in protection against HIV. Crystallographic analyses of anti-V3 crown mAbs in complex with the V3 peptides have revealed that these mAbs recognize the conserved sites on the V3 crown via two distinct strategies: a cradle-binding mode (V3C) and a ladle-binding (V3L) mode. However, almost all of the anti-V3 crown mAbs studied in the past were isolated from chronically HIV-infected individuals. The extents to which the two types of anti-V3 crown Abs are generated by vaccination are unknown. This study analyzed the prevalence of V3C-type and V3L-type Ab responses in HIV-infected individuals and in HIV envelope-immunized humans and animals using peptide mimotopes that distinguish the two Ab types. The results show that both V3L-type and V3C-type Abs were generated by the vast majority of chronically HIV-infected humans, although the V3L-type were more prevalent. In contrast, only one of the two V3 Ab types was elicited in vaccinated humans or animal models, irrespective of HIV-1 envelope clades, envelope constructs (oligomeric or monomeric), and protocols (DNA plus protein or protein alone) used for vaccinations. The V3C-type Abs were produced by vaccinated humans, macaques, and rabbits, whereas the V3L-type Abs were made by mice. The V3C-type and V3L-type Abs generated by the vaccinations were able to mediate virus neutralization. These data indicate the restricted repertoires and the species-specific differences in the functional V3-specific Ab responses induced by the HIV envelope vaccines. The study implies the need for improving immunogen designs and vaccination strategies to broaden the diversity of Abs in order to target the different conserved epitopes in the V3 loop and, by extension, in the entire HIV envelope. Published by Elsevier Ltd.

Cell Culture Systems To Study Human Herpesvirus 6A/B Chromosomal Integration.

PubMed

Gravel, Annie; Dubuc, Isabelle; Wallaschek, Nina; Gilbert-Girard, Shella; Collin, Vanessa; Hall-Sedlak, Ruth; Jerome, Keith R; Mori, Yasuko; Carbonneau, Julie; Boivin, Guy; Kaufer, Benedikt B; Flamand, Louis

2017-07-15

Human herpesviruses 6A/B (HHV-6A/B) can integrate their viral genomes in the telomeres of human chromosomes. The viral and cellular factors contributing to HHV-6A/B integration remain largely unknown, mostly due to the lack of efficient and reproducible cell culture models to study HHV-6A/B integration. In this study, we characterized the HHV-6A/B integration efficiencies in several human cell lines using two different approaches. First, after a short-term infection (5 h), cells were processed for single-cell cloning and analyzed for chromosomally integrated HHV-6A/B (ciHHV-6A/B). Second, cells were infected with HHV-6A/B and allowed to grow in bulk for 4 weeks or longer and then analyzed for the presence of ciHHV-6. Using quantitative PCR (qPCR), droplet digital PCR, and fluorescent in situ hybridization, we could demonstrate that HHV-6A/B integrated in most human cell lines tested, including telomerase-positive (HeLa, MCF-7, HCT-116, and HEK293T) and telomerase-negative cell lines (U2OS and GM847). Our results also indicate that inhibition of DNA replication, using phosphonoacetic acid, did not affect HHV-6A/B integration. Certain clones harboring ciHHV-6A/B spontaneously express viral genes and proteins. Treatment of cells with phorbol ester or histone deacetylase inhibitors triggered the expression of many viral genes, including U39 , U90 , and U100 , without the production of infectious virus, suggesting that the tested stimuli were not sufficient to trigger full reactivation. In summary, both integration models yielded comparable results and should enable the identification of viral and cellular factors contributing to HHV-6A/B integration and the screening of drugs influencing viral gene expression, as well as the release of infectious HHV-6A/B from the integrated state. IMPORTANCE The analysis and understanding of HHV-6A/B genome integration into host DNA is currently limited due to the lack of reproducible and efficient viral integration systems. In the present study, we describe two quantitative cell culture viral integration systems. These systems can be used to define cellular and viral factors that play a role in HHV-6A/B integration. Furthermore, these systems will allow us to decipher the conditions resulting in virus gene expression and excision of the integrated viral genome resulting in reactivation. Copyright © 2017 American Society for Microbiology.

Deletion of nfnAB in Thermoanaerobacterium saccharolyticum and Its Effect on Metabolism

DOE PAGES

Lo, Jonathan; Zheng, Tianyong; Olson, Daniel G.; ...

2015-06-29

NfnAB catalyzes the reversible transfer of electrons from reduced ferredoxin and NADH to 2 NADP +. The NfnAB complex has been hypothesized to be the main enzyme for ferredoxin oxidization in strains of Thermoanaerobacterium saccharolyticum engineered for increased ethanol production. NfnAB complex activity was detectable in crude cell extracts of T. saccharolyticum. In this paper, activity was also detected using activity staining of native PAGE gels. The nfnAB gene was deleted in different strains of T. saccharolyticum to determine its effect on end product formation. In wild-type T. saccharolyticum, deletion of nfnAB resulted in a 46% increase in H 2more » formation but otherwise little change in other fermentation products. In two engineered strains with 80% theoretical ethanol yield, loss of nfnAB caused two different responses: in one strain, ethanol yield decreased to about 30% of the theoretical value, while another strain had no change in ethanol yield. Biochemical analysis of cell extracts showed that the ΔnfnAB strain with decreased ethanol yield had NADPH-linked alcohol dehydrogenase (ADH) activity, while the ΔnfnAB strain with unchanged ethanol yield had NADH-linked ADH activity. Deletion of nfnAB caused loss of NADPH-linked ferredoxin oxidoreductase activity in all cell extracts. Significant NADH-linked ferredoxin oxidoreductase activity was seen in all cell extracts, including those that had lost nfnAB. This suggests that there is an unidentified NADH:ferredoxin oxidoreductase (distinct from nfnAB) playing a role in ethanol formation. The NfnAB complex plays a key role in generating NADPH in a strain that had become reliant on NADPH-ADH activity. Importance: Thermophilic anaerobes that can convert biomass-derived sugars into ethanol have been investigated as candidates for biofuel formation. Many anaerobes have been genetically engineered to increase biofuel formation; however, key aspects of metabolism remain unknown and poorly understood. One example is the mechanism for ferredoxin oxidation and transfer of electrons to NAD(P) +. The electron-bifurcating enzyme complex NfnAB is known to catalyze the reversible transfer of electrons from reduced ferredoxin and NADH to 2 NADP + and is thought to play key roles linking NAD(P)(H) metabolism with ferredoxin metabolism. Finally, we report the first deletion of nfnAB and demonstrate a role for NfnAB in metabolism and ethanol formation in Thermoanaerobacterium saccharolyticum and show that this may be an important feature among other thermophilic ethanologenic anaerobes.« less

Domain based assays of individual antibody concentrations in an oligoclonal combination targeting a single protein

PubMed Central

Meng, Q.; Li, M.; Silberg, M.A.; Conrad, F.; Bettencourt, J.; To, R.; Huang, C.; Ma, J.; Meyer, K.; Shimizu, R.; Cao, L.; Tomic, M.T.; Marks, J.D.

2014-01-01

Quantitation of individual mAbs within a combined antibody drug product is required for preclinical and clinical drug development including pharmacokinetics (PK), toxicology, stability and biochemical characterization studies of such drugs. We have developed an antitoxin (XOMA 3AB) consisting of three recombinant monoclonal antibodies (mAbs) that potently neutralizes the known subtypes of type A botulinum neurotoxin (BoNT/A). The three mAbs bind non-overlapping BoNT/A epitopes with high affinity. XOMA3AB is being developed as a treatment for botulism resulting from BoNT/A. To develop antibody-specific assays, we cloned, expressed, and purified BoNT/A domains from E. coli. Each mAb bound only to its specific domain with affinity comparable to the binding to holotoxin. MAb specific domains were used to develop an ELISA for characterization of the integrity and binding activity of the three mAbs in the drug product. An electrochemiluminescence bridging assay was also developed that is robust to interference from components in serum and we demonstrate that it can be used for PK assays. This type of antigen engineering to generate mAb-specific domains is a general method allowing quantitation and characterization of individual mAbs in a mAb cocktail that bind the same protein and is superior to anti-idiotype approaches. PMID:22037290

Performance evaluation of phage-displayed synthetic human single-domain antibody libraries: A retrospective analysis.

PubMed

Henry, Kevin A; Tanha, Jamshid

2018-05-01

Fully human synthetic single-domain antibodies (sdAbs) are desirable therapeutic molecules but their development is a considerable challenge. Here, using a retrospective analysis of in-house historical data, we examined the parameters that impact the outcome of screening phage-displayed synthetic human sdAb libraries to discover antigen-specific binders. We found no evidence for a differential effect of domain type (V H or V L ), library randomization strategy, incorporation of a stabilizing disulfide linkage or sdAb display format (monovalent vs. multivalent) on the probability of obtaining any antigen-binding human sdAbs, instead finding that the success of library screens was primarily related to properties of target antigens, especially molecular mass. The solubility and binding affinity of sdAbs isolated from successful screens depended both on properties of the sdAb libraries (primarily domain type) and the target antigens. Taking attrition of sdAbs with major manufacturability concerns (aggregation; low expression) and sdAbs that do not recognize native cell-surface antigens as independent probabilities, we calculate the overall likelihood of obtaining ≥1 antigen-binding human sdAb from a single library-target screen as ~24%. Successful library-target screens should be expected to yield ~1.3 human sdAbs on average, each with average binding affinity of ~2 μM. Copyright © 2018 Elsevier B.V. All rights reserved.

HHV-6A/B Integration and the Pathogenesis Associated with the Reactivation of Chromosomally Integrated HHV-6A/B.

PubMed

Collin, Vanessa; Flamand, Louis

2017-06-26

Unlike other human herpesviruses, human herpesvirus 6A and 6B (HHV-6A/B) infection can lead to integration of the viral genome in human chromosomes. When integration occurs in germinal cells, the integrated HHV-6A/B genome can be transmitted to 50% of descendants. Such individuals, carrying one copy of the HHV-6A/B genome in every cell, are referred to as having inherited chromosomally-integrated HHV-6A/B (iciHHV-6) and represent approximately 1% of the world's population. Interestingly, HHV-6A/B integrate their genomes in a specific region of the chromosomes known as telomeres. Telomeres are located at chromosomes' ends and play essential roles in chromosomal stability and the long-term proliferative potential of cells. Considering that the integrated HHV-6A/B genome is mostly intact without any gross rearrangements or deletions, integration is likely used for viral maintenance into host cells. Knowing the roles played by telomeres in cellular homeostasis, viral integration in such structure is not likely to be without consequences. At present, the mechanisms and factors involved in HHV-6A/B integration remain poorly defined. In this review, we detail the potential biological and medical impacts of HHV-6A/B integration as well as the possible chromosomal integration and viral excision processes.

Factorial Validity of the Movement Assessment Battery for Children-2nd Edition (MABC-2) in 7-16-Year-Olds.

PubMed

Psotta, Rudolf; Abdollahipour, Reza

2017-12-01

The Movement Assessment Battery for Children-2nd Edition (MABC-2) is a test of motor development, widely used in clinical and research settings. To address which motor abilities are actually captured by the motor tasks in the two age versions of the MABC-2, the AB2 for 7- 10-year-olds and the AB3 for 11- 16-year-olds, we examined AB2 and AB3 factorial validity. We conducted confirmatory factor analysis (SPSS AMOS 22.0) on data from the test's standardization samples of children aged 7-10, n = 483, and 11-16, n = 674, in order to find the best fitting models. The covariance matrix of AB2 and AB3 fit a three-factor model that included tasks of manual dexterity, aiming and catching, and balance. However, factor analytic models fitting AB2 and AB3 did not involve the dynamic balance tasks of hopping with the better leg and hopping with the other leg; and the drawing trail showed very low factor validity. In sum, both AB2 and AB3 of the MABC-2 test are able to discriminate between the three specific motor abilities; but due to questionable psychometric quality, the drawing trail and hopping tasks should be modified to improve the construct validity for both age versions of the MABC-2.

Identification of Bacillus thuringiensis Cry1AbMod binding-proteins from Spodoptera frugiperda.

PubMed

Martínez de Castro, Diana L; García-Gómez, Blanca I; Gómez, Isabel; Bravo, Alejandra; Soberón, Mario

2017-12-01

Bacillus thuringiensis Cry toxins are currently used for pest control in transgenic crops but evolution of resistance by the insect pests threatens the use of this technology. The Cry1AbMod toxin was engineered to lack the alpha helix-1 of the parental Cry1Ab toxin and was shown to counter resistance to Cry1Ab and Cry1Ac toxins in different insect species including the fall armyworm Spodoptera frugiperda. In addition, Cry1AbMod showed enhanced toxicity to Cry1Ab-susceptible S. frugiperda populations. To gain insights into the mechanisms of this Cry1AbMod-enhanced toxicity, we isolated the Cry1AbMod toxin binding proteins from S. frugiperda brush border membrane vesicles (BBMV), which were identified by pull-down assay and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The LC-MS/MS results indicated that Cry1AbMod toxin could bind to four classes of aminopeptidase (N1, N3, N4 y N5) and actin, with the highest amino acid sequence coverage acquired for APN 1 and APN4. In addition to these proteins, we found other proteins not previously described as Cry toxin binding proteins. This is the first report that suggests the interaction between Cry1AbMod and APN in S. frugiperda. Copyright © 2017 Elsevier Inc. All rights reserved.

Dissecting linear and conformational epitopes on the native thyrotropin receptor.

PubMed

Ando, Takao; Latif, Rauf; Daniel, Samira; Eguchi, Katsumi; Davies, Terry F

2004-11-01

The TSH receptor (TSHR) is the primary antigen in Graves' disease. In this condition, autoantibodies to the TSHR that have intrinsic thyroid-stimulating activity develop. We studied the epitopes on the native TSHR using polyclonal antisera and monoclonal antibodies (mAbs) derived from an Armenian hamster model of Graves' disease. Of 14 hamster mAbs analyzed, five were shown to bind to conformational epitopes including one mAb with potent thyroid-stimulating activity. Overlapping conformational epitopes were determined by cell-binding competition assays using fluorescently labeled mAbs. We identified two distinct conformational epitopes: epitope A for both stimulating and blocking mAbs and epitope B for only blocking mAbs. Examination of an additional three mouse-derived stimulating TSHR-mAbs also showed exclusive binding to epitope A. The remaining nine hamster-derived mAbs were neutral or low-affinity blocking antibodies that recognized linear epitopes within the TSHR cleaved region (residues 316-366) (epitope C). Serum from the immunized hamsters also recognized conformational epitopes A and B but, in addition, also contained high levels of TSHR-Abs interacting within the linear epitope C region. In summary, these studies indicated that the natively conformed TSHR had a restricted set of epitopes recognized by TSHR-mAbs and that the binding site for stimulating TSHR-Abs was highly conserved. However, high-affinity TSHR-blocking antibodies recognized two conformational epitopes, one of which was indistinguishable from the thyroid-stimulating epitope. Hence, TSHR-stimulating and blocking antibodies cannot be distinguished purely on the basis of their conformational epitope recognition.

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.

Staphylococcus aureus Biofilms Induce Macrophage Dysfunction Through Leukocidin AB and Alpha-Toxin

PubMed Central

Scherr, Tyler D.; Hanke, Mark L.; Huang, Ouwen; James, David B. A.; Horswill, Alexander R.; Bayles, Kenneth W.; Fey, Paul D.; Torres, Victor J.

2015-01-01

ABSTRACT The macrophage response to planktonic Staphylococcus aureus involves the induction of proinflammatory microbicidal activity. However, S. aureus biofilms can interfere with these responses in part by polarizing macrophages toward an anti-inflammatory profibrotic phenotype. Here we demonstrate that conditioned medium from mature S. aureus biofilms inhibited macrophage phagocytosis and induced cytotoxicity, suggesting the involvement of a secreted factor(s). Iterative testing found the active factor(s) to be proteinaceous and partially agr-dependent. Quantitative mass spectrometry identified alpha-toxin (Hla) and leukocidin AB (LukAB) as critical molecules secreted by S. aureus biofilms that inhibit murine macrophage phagocytosis and promote cytotoxicity. A role for Hla and LukAB was confirmed by using hla and lukAB mutants, and synergy between the two toxins was demonstrated with a lukAB hla double mutant and verified by complementation. Independent confirmation of the effects of Hla and LukAB on macrophage dysfunction was demonstrated by using an isogenic strain in which Hla was constitutively expressed, an Hla antibody to block toxin activity, and purified LukAB peptide. The importance of Hla and LukAB during S. aureus biofilm formation in vivo was assessed by using a murine orthopedic implant biofilm infection model in which the lukAB hla double mutant displayed significantly lower bacterial burdens and more macrophage infiltrates than each single mutant. Collectively, these findings reveal a critical synergistic role for Hla and LukAB in promoting macrophage dysfunction and facilitating S. aureus biofilm development in vivo. PMID:26307164

Blockade of invariant TCR-CD1d interaction specifically inhibits antibody production against blood group A carbohydrates

PubMed Central

Tazawa, Hirofumi; Irei, Toshimitsu; Tanaka, Yuka; Igarashi, Yuka; Tashiro, Hirotaka

2013-01-01

Previously, we detected B cells expressing receptors for blood group A carbohydrates in the CD11b+CD5+ B-1a subpopulation in mice, similar to that in blood group O or B in humans. In the present study, we demonstrate that CD1d-restricted natural killer T (NKT) cells are required to produce anti-A antibodies (Abs), probably through collaboration with B-1a cells. After immunization of wild-type (WT) mice with human blood group A red blood cells (A-RBCs), interleukin (IL)-5 exclusively and transiently increased and the anti-A Abs were elevated in sera. However, these reactions were not observed in CD1d−/− mice, which lack NKT cells. Administration of anti-mouse CD1d blocking monoclonal Abs (mAb) prior to immunization abolished IL-5 production by NKT cells and anti-A Ab production in WT mice. Administration of anti-IL-5 neutralizing mAb also diminished anti-A Ab production in WT mice, suggesting that IL-5 secreted from NKT cells critically regulates anti-A Ab production by B-1a cells. In nonobese diabetic/severe combined immunodeficient (NOD/SCID/γcnull) mice, into which peripheral blood mononuclear cells from type O human volunteers were engrafted, administration of anti-human CD1d mAb prior to A-RBC immunization completely inhibited anti-A Ab production. Thus, anti-CD1d treatment might constitute a novel approach that could help in evading Ab-mediated rejection in ABO-incompatible transplant recipients. PMID:23943651

[Distribution of IgG subclasses of TgAb and TPOAb in sera from patients with Graves' disease, Graves' disease plus Hashimoto's thyroiditis and Hashimoto's thyrotoxicosis].

PubMed

Yuan, Shanshan; Yu, Nan; Gao, Ying; Huang, Wei; He, Yifan; Dong, Bin; Lu, Guizhi; Li, Maorong; Cai, Xiaopin; Peng, Dingqiong; Wang, Yunhong; Li, Ting; Huang, Youyuan; Gao, Yanming; Guo, Xiaohui; Shi, Bingyin

2014-01-14

To evaluate the distribution of IgG subclasses of TgAb and TPOAb in sera from patients with Graves' disease (GD), Graves' disease plus Hashimoto's thyroiditis (GH) and Hashimoto's thyrotoxicosis. Patients with GD (n = 33), GH (n = 31) or Hashimoto's thyrotoxicosis (n = 18) diagnosed by fine needle aspiration cytology at Department of Endocrinology of Peking University First Hospital, Beijing Haidian Hospital, China-Japan Friendship Hospital and Civil Aviation General Hospital during the period from January 2010 to May 2013 were enrolled. All of them had TgAb and TPOAb. The total serum IgG and IgG subclasses of TgAb and TPOAb were detected by antigen-specific enzyme-linked immunosorbent assay (ELISA). The prevalence and relative amount of IgG subclasses were calculated and compared among three groups. The levels of TRAb in GD group (21.80(7.53, 40) U/L) were significantly higher than those in GH (7.30(3.10, 25.40) U/L) (P = 0.000) and Hashimoto's thyrotoxicosis groups (4.90(1.69, 16.43) U/L) (P = 0.003). And no significant differences were found in the levels of TgAb and TPOAb. The prevalence of TgAb IgG3 subclass in Hashimoto's thyrotoxicosis group (66.7%) was higher than GD group (35.5%) and GH group (36.4%) and the difference was close to significance (P = 0.066). There were significant differences of relative amount of TgAb IgG2 and TgAb IgG4 among three groups (P = 0.039 and 0.013), and GD patients had higher relative amounts of TgAb IgG2 (0.59(0.34, 0.94)) and TgAb IgG4 (0.57(0.28, 0.97)) than GH patients (TgAb IgG2, 0.31(0.23, 0.34); TgAb IgG4, 0.26(0.09, 0.48)) or patients with Hashimoto's thyrotoxicosis (TgAb IgG2, 0.32(0.24, 0.83); TgAb IgG4, 0.33(0.10, 0.65)) (for TgAb IgG2, P = 0.009 and 0.167; for TgAb IgG4, P = 0.005 and 0.041 respectively). No significant difference was found in the prevalence of each TPOAb IgG subclass. The difference of relative amount of TPOAb IgG2 among three groups was close to significance (P = 0.069). And the relative amount was higher in sera from GD patients (0.39 ± 0.04) than that in GH patients (0.29 ± 0.13) or patients with Hashimoto's thyrotoxicosis (0.26 ± 0.02) (P = 0.104 and 0.002 respectively). The patients with high levels of TgAb IgG2, TgAb IgG4 and TPOAb IgG2 subclasses have a greater risk of GD. The IgG subclass distribution of TgAb and TPOAb might help to differentiate the causes of thyrotoxicosis in autoimmune thyroid diseases.

Pattern of cytokine receptors expressed by human dendritic cells migrated from dermal explants.

PubMed Central

Larregina, A T; Morelli, A E; Kolkowski, E; Sanjuan, N; Barboza, M E; Fainboim, L

1997-01-01

Different reasons account for the lack of information about the expression of cytokine receptors on human dendritic cells (DC): (a) DC are a trace population; (b) the proteolytic treatment used to isolate DC may alter enzyme-sensitive epitopes; and (c) low numbers of receptors per cell. In the present work the expression of cytokine receptors was analysed by flow cytometry on the population of dermal DC (DDC) that spontaneously migrate from short-term culture dermal explants. DDC obtained after dermal culture were CD1alow, CD1b+, CD1c+, human leucocyte antigen (HLA)-DR+, CD11chigh, CD11b+ and CD32+. The DC lineage was confirmed by ultrastructural analysis. DDC expressed interleukin (IL)-1R type 1 (monoclonal antibody (mAb) hIL-1R1-M1; and 6B5); IL-1R type 2 (mAb hIL-1R2-M22); IL-2R alpha chain (mAb anti-Tac; and hIL-2R-M1) and IL-2R gamma chain (mAb 3B5; and AG14C). DDC did not stain for IL-2R beta chain using four mAbs recognizing two different epitopes of IL-2R beta (mAb 2R-B; Mik-beta 1; and CF1; Mik-beta 3, respectively). DDC were also positive for the cytokine binding chains (alpha chains) of IL-3R (mAb 9F5); IL-4R (mAb hIL-4R-M57; and S456C9); and IL-7R (mAb hIL-7R-M20; and R3434). DDC showed low levels of IL-6R alpha chain (mAb B-F19; B-R6; and B-E23) and its signal transducer gp130 (mAb A2; and B1). DDC strongly expressed interferon-gamma receptor (IFN-gamma R) (mAb GIR-208) and were negative for IL-8R (mAb B-G20; and B-F25). All DDC were highly positive for granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) alpha chain (mAb hGM-CSFR-M1; SC06; SC04, and 8G6) and to a lesser extent for the common beta chain of GM-CSFR, IL-3R and IL-5R (mAb 3D7). On the other hand, reactivity was not found for granulocyte colony-stimulating factor receptor (G-CSFR) (mAb hGCSFR-M1) nor macrophage colony-stimulating factor receptor (M-CSFR) (mAb 7-7A3-17) confirming the DC lineage of DDC. As previously reported for lymphoid DC, DDC expressed tumour necrosis factor receptort (TNFR) 75000 MW (mAb utr-1; hTNFR-M1; and MR2-1) but lacked TNFR 55000 MW (mAb htr-9; MR1-1; and MR1-2). In summary, DDC express receptors for a broad panel of cytokines, even receptors for cytokines whose effects on DC are still unknown (i.e. IL-2R alpha gamma; IL-6R alpha/gp 130; IL-7R alpha gamma). Images Figure 1 PMID:9227332

[The management of asymptomatic bacteriuria in different patient population].

PubMed

Ivanov, M-L; Malinverni, R

2008-11-12

Who should be screened for asymptomatic bacteriuria (AB) and who should be treated? This review updates some aspects of the management of AB in different patient populations. A systematic screening for AB is recommended for pregnant women because of a significant risk of complications. In these cases as well as before any uro-gynecologic surgical procedure treatment of AB is strongly recommended. The management of AB in immunosuppressed or transplanted patients is more controversial. In other populations treating AB is not recommended and the outcome seems to be worse in case of treatment due to possible side effects and selection of resistant organisms. Recent studies have shown a considerable gap between clinical practice and recommendations.

Multicapillary SDS-gel electrophoresis for the analysis of fluorescently labeled mAb preparations: a high throughput quality control process for the production of QuantiPlasma and PlasmaScan mAb libraries.

PubMed

Székely, Andrea; Szekrényes, Akos; Kerékgyártó, Márta; Balogh, Attila; Kádas, János; Lázár, József; Guttman, András; Kurucz, István; Takács, László

2014-08-01

Molecular heterogeneity of mAb preparations is the result of various co- and post-translational modifications and to contaminants related to the production process. Changes in molecular composition results in alterations of functional performance, therefore quality control and validation of therapeutic or diagnostic protein products is essential. A special case is the consistent production of mAb libraries (QuantiPlasma™ and PlasmaScan™) for proteome profiling, quality control of which represents a challenge because of high number of mAbs (>1000). Here, we devise a generally applicable multicapillary SDS-gel electrophoresis process for the analysis of fluorescently labeled mAb preparations for the high throughput quality control of mAbs of the QuantiPlasma™ and PlasmaScan™ libraries. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Synthesis of an A'B' Precursor to Angelmicin B: Product Diversification in the Suárez Lactol Fragmentation.

PubMed

Li, Jialiang; Todaro, Louis; Mootoo, David R

2011-11-01

We describe a synthetic strategy for the angelimicin family of anthraquinoid natural products that involves converting a central highly oxygenated decalin intermediate to the AB and A'B' subunits. Herein, we report the synthesis of the bicyclic A'B' subunit that complements our earlier route to the tricyclic AB framework. The differentiating tact in the two syntheses focused on controlling the Suárez radical fragmentation of lactol precursors by modulating the substrate's structural rigidity. A more flexible lactol gave the tricyclic AB framework, whereas a more rigid substrate led to the bicyclic A'B' precursor, presumably through divergent pathways from the radical produced in the initial fragmentation step. These results establish a versatile advanced synthetic precursor for the angelimicins, and on a more general note, illustrate strategies for applying the Suárez fragmentation to diverse and complex molecular frameworks.

Effect of Graphene Oxide Mixed Epoxy on Mechanical Properties of Carbon Fiber/Acrylonitrile-Butadiene-Styrene Composites.

PubMed

Wang, Cuicui; Ge, Heyi; Ma, Xiaolong; Liu, Zhifang; Wang, Ting; Zhang, Jingyi

2018-04-01

In this study, the watersoluble epoxy resin was prepared via the ring-opening reaction between diethanolamine and epoxy resin. The modified resin mixed with graphene oxide (GO) as a sizing agent was coated onto carbon fiber (CF) and then the GO-CF reinforced acrylonitrile-butadienestyrene (ABS) composites were prepared. The influences of the different contents of GO on CF and CF/ABS composite were explored. The combination among epoxy, GO sheets and maleic anhydride grafted ABS (ABSMA) showed a synergistic effect on improving the properties of GO-CF and GO-CF/ABS composite. The GO-CF had higher single tensile strength than the commercial CF. The maximum ILSS of GO-CF/ABS composite obtained 19.2% improvement as compared with that of the commercial CF/ABS composite. Such multiscale enhancement method and the synergistic reinforced GO-CF/ABS composite show good prospective applications in many industry areas.

The effectiveness of antilock brake systems on motorcycles in reducing real-life crashes and injuries.

PubMed

Rizzi, Matteo; Strandroth, Johan; Tingvall, Claes

2009-10-01

This study set out to evaluate the effectiveness of antilock brake system (ABS) technology on motorcycles in reducing real-life injury crashes and to mitigate injury severity. The study comprised an analysis of in-depth fatal crash data in Sweden during 2005-2008 to investigate the potential of ABS as well an estimate of the effectiveness of ABS in crash reduction in Sweden between 2003 and 2008 using induced exposure methods. Findings show that head-on collisions were the least ABS-affected crash types and collisions at intersections the most influenced. Induced exposure analysis showed that the overall effectiveness of ABS was 38 percent on all crashes with injuries and 48 percent on all severe and fatal crashes, with a minimum effectiveness of 11 and 17 percent, respectively. The study recommends the fitment of ABS on all new motorcycles as soon as possible and that customers only purchase motorcycles with ABS.

Immune perturbations in HIV–1-infected individuals who make broadly reactive neutralizing antibodies

PubMed Central

Moody, M. Anthony; Pedroza-Pacheco, Isabela; Vandergrift, Nathan A.; Chui, Cecilia; Lloyd, Krissey E.; Parks, Robert; Soderberg, Kelly A.; Ogbe, Ane T.; Cohen, Myron S.; Liao, Hua-Xin; Gao, Feng; McMichael, Andrew J.; Montefiori, David C.; Verkoczy, Laurent; Kelsoe, Garnett; Huang, Jinghe; Shea, Patrick R.; Connors, Mark; Borrow, Persephone; Haynes, Barton F.

2017-01-01

Induction of broadly neutralizing antibodies (bnAbs) is a goal of HIV-1 vaccine development. BnAbs occur in some HIV-1-infected individuals and frequently have characteristics of autoantibodies. Here we have studied cohorts of HIV-1-infected individuals that made bnAbs and compared them to those who did not do so, and determined immune traits associated with the ability to produce bnAbs. HIV-1-infected individuals with bnAbs had a higher frequency of blood autoantibodies, a lower frequency of regulatory CD4+ T cells, a higher frequency of circulating memory T follicular helper CD4+ cells and a higher T regulatory cell level of programmed cell death-1 expression compared to HIV-1-infected individuals without bnAbs. Thus, induction of HIV-1 bnAbs may require vaccination regimens that transiently mimic immunologic perturbations in HIV-1-infected individuals. PMID:28783677

Executive functioning in preschool children: performance on A-not-B and other delayed response format tasks.

PubMed

Espy, K A; Kaufmann, P M; McDiarmid, M D; Glisky, M L

1999-11-01

The A-not-B (AB) task has been hypothesized to measure executive/frontal lobe function; however, the developmental and measurement characteristics of this task have not been investigated. Performances on AB and comparison tasks adapted from developmental and neuroscience literature was examined in 117 preschool children (ages 23-66 months). Age significantly predicted performance on AB, Delayed Alternation, Spatial Reversal, Color Reversal, and Self-Control tasks. A four-factor analytic model best fit task performance data. AB task indices loaded on two factors with measures from the Self-Control and Delayed Alternation tasks, respectively. AB indices did not load with those from the reversal tasks despite similarities in task administration and presumed cognitive demand (working memory). These results indicate that AB is sensitive to individual differences in age-related performance in preschool children and suggest that AB performance is related to both working memory and inhibition processes in this age range.

Can attenuation of attentional blink also evoke removal of repetition blindness?

PubMed

Choi, Hoon; Watanabe, Takeo

2014-06-01

A recent study showed that attentional blink (AB), which has been considered to reflect the capacity limitation of visual temporal attention, can be attenuated after a short period of the color-salient training, in which the second target (T2) within the AB period is given a salient color (Choi et al., 2012). The current study explored whether the effect of the color-salient training could be transferred to another phenomenon. In addition to AB, repetition blindness (RB) was employed, which is phenomenologically similar to, but fundamentally different from AB. After completion of the color-salient training with a nonrepeated T2 (corresponding to AB), RB was still observed, whereas AB was completely removed. However, the color-salient training with a repeated T2 (similar to RB) induced not only a significant reduction of RB but also an attenuation of AB. This result provides further evidence for dissociation between AB and RB. In addition, it implies that the color-salient training improves the attentional control mechanism related to target-distractor discrimination rather than to the perceptual system. Copyright © 2014 Elsevier Ltd. All rights reserved.

Bioaccessibility of cadmium in fresh and cooked Agaricus blazei Murill assessed by in vitro biomimetic digestion system.

PubMed

Sun, Liping; Liu, Gaoxiang; Yang, Meizhizi; Zhuang, Yongliang

2012-05-01

Bioaccessibility of cadmium (Cd) in fresh and cooked Agaricus blazei Murill (AbM) was studied by an in vitro biomimetic digestion system in this paper. The results showed that the Cd content in fresh AbM was 10.27 mg kg(-1) DM. The cooking treatments of boiling and microwaving with water significantly decreased Cd contents in fresh AbM by 36.4% and 30.2% (P<0.05), respectively. Cd in fresh AbM showed the highest bioaccessibility of 77.8% during the biomimetic digestion in stomach, followed by that of 69.4% from the gastrointestinal digestion. Cooking treatments also significantly lowered the bioaccessibility of Cd (P<0.05). Cd in boiled AbM showed 50.7% and 46.1% bioaccessibility during the gastric and gastrointestinal procedures. While, Cd in microwaved AbM showed 58.2% and 50.4% bioaccessibility. This study confirmed that the health risk assessment of AbM depending on the total Cd levels in fresh AbM was inaccurate, especially for the products domestically cooked. Copyright © 2012 Elsevier Ltd. All rights reserved.

What Are the Primary Limitations in B-Cell Affinity Maturation, and How Much Affinity Maturation Can We Drive with Vaccination? Breaking through Immunity's Glass Ceiling.

PubMed

Kelsoe, Garnett; Haynes, Barton F

2018-05-01

A key goal of HIV-1 vaccine development is the induction of broadly neutralizing antibodies (bnAbs) targeted to the vulnerable regions of the HIV envelope. BnAbs develop over time in ∼50% of HIV-1-infected individuals. However, to date, no vaccines have induced bnAbs and few or none of these vaccine-elicited HIV-1 antibodies carry the high frequencies of V(D)J mutations characteristic of bnAbs. Do the high frequencies of mutations characteristic of naturally induced bnAbs represent a fundamental barrier to the induction of bnAbs by vaccines? Recent studies suggest that high frequencies of V(D)J mutations can be achieved by serial vaccination strategies. Rather, it appears that, in the absence of HIV-1 infection, physiologic immune tolerance controls, including a germinal center process termed affinity reversion, may limit vaccine-driven bnAb development by clonal elimination or selecting for mutations incompatible with bnAb activity. Copyright © 2018 Cold Spring Harbor Laboratory Press; all rights reserved.

Synergistic anti-tumor therapy by a comb-like multifunctional antibody nanoarray with exceptionally potent activity

NASA Astrophysics Data System (ADS)

Li, Huafei; Sun, Yun; Chen, Di; Zhao, He; Zhao, Mengxin; Zhu, Xiandi; Ke, Changhong; Zhang, Ge; Jiang, Cheng; Zhang, Li; Zhang, Fulei; Wei, Huafeng; Li, Wei

2015-10-01

Simultaneously blocking multiple mediators offers new hope for the treatment of complex diseases. However, the curative potential of current combination therapy by chronological administration of separate monoclonal antibodies (mAbs) or multi-specific mAbs is still moderate due to inconvenient manipulation, low cooperative effectors, poor pharmacokinetics and insufficient tumor accumulation. Here, we describe a facile strategy that arms distinct mAbs with cooperative effectors onto a long chain to form a multicomponent comb-like nano mAb. Unlike dissociative parental mAbs, the multifunctional mAb nanoarray (PL-RB) constructed from type I/II anti-CD20 mAbs shows good pharmacokinetics. This PL-RB simultaneously targets distinct epitopes on a single antigen (Ag) and neighboring Ags on different lymphocytes. This unique intra- and intercellular Ag cross-linking endows the multifunctional mAb nanoarray with potent apoptosis activity. The exceptional apoptosis, complement-dependent cytotoxicity (CDC), antibody-dependent cellular cytotoxicity (ADCC) that are synchronously evoked by the nano PL-RB are further synergistically promoted via enhanced permeability and retention (EPR), which resulted in high intratumor accumulation and excellent anti-lymphoma efficiency.

Intramuscular Adeno-Associated Virus-Mediated Expression of Monoclonal Antibodies Provides 100% Protection Against Ebola Virus Infection in Mice.

PubMed

van Lieshout, Laura P; Soule, Geoff; Sorensen, Debra; Frost, Kathy L; He, Shihua; Tierney, Kevin; Safronetz, David; Booth, Stephanie A; Kobinger, Gary P; Qiu, Xiangguo; Wootton, Sarah K

2018-03-05

The 2013-2016 West Africa outbreak demonstrated the epidemic potential of Ebola virus and highlighted the need for counter strategies. Monoclonal antibody (mAb)-based therapies hold promise as treatment options for Ebola virus infections. However, production of clinical-grade mAbs is labor intensive, and immunity is short lived. Conversely, adeno-associated virus (AAV)-mediated mAb gene transfer provides the host with a genetic blueprint to manufacture mAbs in vivo, leading to steady release of antibody over many months. Here we demonstrate that AAV-mediated expression of nonneutralizing mAb 5D2 or 7C9 confers 100% protection against mouse-adapted Ebola virus infection, while neutralizing mAb 2G4 was 83% protective. A 2-component cocktail, AAV-2G4/AAV-5D2, provided complete protection when administered 7 days prior to challenge and was partially protective with a 3-day lead time. Finally, AAV-mAb therapies provided sustained protection from challenge 5 months following AAV administration. AAV-mAb may be a viable alternative strategy for vaccination against emerging infectious diseases.

Early Antibody Lineage Diversification and Independent Limb Maturation Lead to Broad HIV-1 Neutralization Targeting the Env High-Mannose Patch.

PubMed

MacLeod, Daniel T; Choi, Nancy M; Briney, Bryan; Garces, Fernando; Ver, Lorena S; Landais, Elise; Murrell, Ben; Wrin, Terri; Kilembe, William; Liang, Chi-Hui; Ramos, Alejandra; Bian, Chaoran B; Wickramasinghe, Lalinda; Kong, Leopold; Eren, Kemal; Wu, Chung-Yi; Wong, Chi-Huey; Kosakovsky Pond, Sergei L; Wilson, Ian A; Burton, Dennis R; Poignard, Pascal

2016-05-17

The high-mannose patch on HIV Env is a preferred target for broadly neutralizing antibodies (bnAbs), but to date, no vaccination regimen has elicited bnAbs against this region. Here, we present the development of a bnAb lineage targeting the high-mannose patch in an HIV-1 subtype-C-infected donor from sub-Saharan Africa. The Abs first acquired autologous neutralization, then gradually matured to achieve breadth. One Ab neutralized >47% of HIV-1 strains with only ∼11% somatic hypermutation and no insertions or deletions. By sequencing autologous env, we determined key residues that triggered the lineage and participated in Ab-Env coevolution. Next-generation sequencing of the Ab repertoire showed an early expansive diversification of the lineage followed by independent maturation of individual limbs, several of them developing notable breadth and potency. Overall, the findings are encouraging from a vaccine standpoint and suggest immunization strategies mimicking the evolution of the entire high-mannose patch and promoting maturation of multiple diverse Ab pathways. Copyright © 2016 Elsevier Inc. All rights reserved.

Synergistic anti-tumor therapy by a comb-like multifunctional antibody nanoarray with exceptionally potent activity.

PubMed

Li, Huafei; Sun, Yun; Chen, Di; Zhao, He; Zhao, Mengxin; Zhu, Xiandi; Ke, Changhong; Zhang, Ge; Jiang, Cheng; Zhang, Li; Zhang, Fulei; Wei, Huafeng; Li, Wei

2015-10-28

Simultaneously blocking multiple mediators offers new hope for the treatment of complex diseases. However, the curative potential of current combination therapy by chronological administration of separate monoclonal antibodies (mAbs) or multi-specific mAbs is still moderate due to inconvenient manipulation, low cooperative effectors, poor pharmacokinetics and insufficient tumor accumulation. Here, we describe a facile strategy that arms distinct mAbs with cooperative effectors onto a long chain to form a multicomponent comb-like nano mAb. Unlike dissociative parental mAbs, the multifunctional mAb nanoarray (PL-RB) constructed from type I/II anti-CD20 mAbs shows good pharmacokinetics. This PL-RB simultaneously targets distinct epitopes on a single antigen (Ag) and neighboring Ags on different lymphocytes. This unique intra- and intercellular Ag cross-linking endows the multifunctional mAb nanoarray with potent apoptosis activity. The exceptional apoptosis, complement-dependent cytotoxicity (CDC), antibody-dependent cellular cytotoxicity (ADCC) that are synchronously evoked by the nano PL-RB are further synergistically promoted via enhanced permeability and retention (EPR), which resulted in high intratumor accumulation and excellent anti-lymphoma efficiency.

Experimental determination of single CdSe nanowire absorption cross sections through photothermal imaging.

PubMed

Giblin, Jay; Syed, Muhammad; Banning, Michael T; Kuno, Masaru; Hartland, Greg

2010-01-26

Absorption cross sections ((sigma)abs) of single branched CdSe nanowires (NWs) have been measured by photothermal heterodyne imaging (PHI). Specifically, PHI signals from isolated gold nanoparticles (NPs) with known cross sections were compared to those of individual CdSe NWs excited at 532 nm. This allowed us to determine average NW absorption cross sections at 532 nm of (sigma)abs = (3.17 +/- 0.44) x 10(-11) cm2/microm (standard error reported). This agrees well with a theoretical value obtained using a classical electromagnetic analysis ((sigma)abs = 5.00 x 10(-11) cm2/microm) and also with prior ensemble estimates. Furthermore, NWs exhibit significant absorption polarization sensitivities consistent with prior NW excitation polarization anisotropy measurements. This has enabled additional estimates of the absorption cross section parallel ((sigma)abs) and perpendicular ((sigma)abs(perpendicular) to the NW growth axis, as well as the corresponding NW absorption anisotropy ((rho)abs). Resulting values of (sigma)abs = (5.6 +/- 1.1) x 10(-11) cm2/microm, (sigma)abs(perpendicular) = (1.26 +/- 0.21) x 10(-11) cm2/microm, and (rho)abs = 0.63+/- 0.04 (standard errors reported) are again in good agreement with theoretical predictions. These measurements all indicate sizable NW absorption cross sections and ultimately suggest the possibility of future direct single NW absorption studies.

Sensitivity-Enhancement of FRET Immunoassays by Multiple-Antibody Conjugation on Quantum Dots.

PubMed

Annio, Giacomo; Jennings, Travis; Tagit, Oya; Hildebrandt, Niko

2018-05-23

Quantum dots (QDs) are not only advantageous for color-tuning, improved brightness, and high stability, but their nanoparticle surfaces also allow for the attachment of many biomolecules. Because IgG antibodies (ABs) are in the same size range of biocompatible QDs and the AB orientation after conjugation to the QD is often random, it is difficult to predict if few or many ABs per QD will lead to an efficient AB-QD conjugate. This is particularly true for homogeneous Förster resonance energy transfer (FRET) sandwich immunoassays, for which the ABs on the QD must bind a biomarker that needs to bind a second AB-FRET-conjugate. Here, we investigate the performance of Tb-to-QD FRET immunoassays against total prostate specific antigen (TPSA) by changing the number of ABs per QD while leaving all the other assay components unchanged. We first characterize the AB-QD conjugation by various spectroscopic, microscopic, and chromatographic techniques and then quantify the TPSA immunoassay performance regarding sensitivity, limit of detection, and dynamic range. Our results show that an increasing conjugation ratio leads to significantly enhanced FRET immunoassays. These findings will be highly important for developing QD-based immunoassays in which the concentrations of both ABs and QDs can significantly influence the assay performance.

Antibodies with 'Original Antigenic Sin' Properties Are Valuable Components of Secondary Immune Responses to Influenza Viruses.

PubMed

Linderman, Susanne L; Hensley, Scott E

2016-08-01

Human antibodies (Abs) elicited by influenza viruses often bind with a high affinity to past influenza virus strains, but paradoxically, do not bind to the viral strain actually eliciting the response. This phenomena is called 'original antigenic sin' (OAS) since this can occur at the expense of generating new de novo Abs. Here, we characterized the specificity and functionality of Abs elicited in mice that were sequentially exposed to two antigenically distinct H1N1 influenza virus strains. Many Abs elicited under these conditions had an OAS phenotype, in that they bound strongly to the viral strain used for the first exposure and very weakly to the viral strain used for the second exposure. We found that OAS and non-OAS Abs target the same general region of the influenza hemagglutinin protein and that B cells expressing these two types of Abs can be clonally-related. Surprisingly, although OAS Abs bound with very low affinities, some were able to effectively protect against an antigenically drifted viral strain following passive transfer in vivo. Taken together, our data indicate that OAS Abs share some level of cross-reactivity between priming and recall viral strains and that B cells producing these Abs can be protective when recalled into secondary immune responses.

Antithyroid Antibodies Are Implicated in Epileptogenesis of Adult Patients With Epilepsy.

PubMed

Tsai, Meng-Han; Fu, Ting-Ying; Chen, Nai-Ching; Shih, Fu-Yuan; Lu, Yan-Ting; Cheng, Mei-Yun; Chuang, Hung-Yi; Chuang, Yao-Chung

2015-07-01

Antithyroid antibodies (Abs) are associated with epilepsy in steroid-responsive encephalopathy, but have been rarely studied in unselected epilepsy patients. This study aimed to characterize the prevalence and associated factors of antithyroid Abs and other auto-Abs in adult patients with epilepsy.Epilepsy patients without autoimmune disorders were surveyed for antinuclear antibody (ANA), anti-β2 glycoprotein 1 antibody (aβ2GP1), anticardiolipin IgG Ab, antimicrosomal antibody (AMA), antithyroglobulin antibody (ATA), and thyroid function test.Of 319 patients, 75 (23.5%) were positive for at least 1 Ab. The most common Ab was anticardiolipin antibody (aCL) (30/319, 9.4%), followed by AMA (24/319, 7.5%), ANA (18/319, 5.6%), aβ2GP1 (18/319, 6.5%), and ATA (6/319, 3.25%). Antimicrosomal Abs were significantly more frequent in patients who were female, older at disease onset, older at the time of study, and had unknown seizure etiology. The presence of aCL was significantly associated with more frequent seizures. Most patients with antithyroid Ab were female and had focal seizures with unknown etiology.The association of different auto-Abs with different factors suggests that they may have different roles in adult patients with epilepsy. Recurrent seizures and certain antiepileptic medications may cause the production of aCL. The role of antithyroid Abs in adult focal epilepsy with unknown cause, especially in females, warrants further evaluation because of the potential implications on treatment.

Antithyroid Antibodies Are Implicated in Epileptogenesis of Adult Patients With Epilepsy

PubMed Central

Tsai, Meng-Han; Fu, Ting-Ying; Chen, Nai-Ching; Shih, Fu-Yuan; Lu, Yan-Ting; Cheng, Mei-Yun; Chuang, Hung-Yi; Chuang, Yao-Chung

2015-01-01

Abstract Antithyroid antibodies (Abs) are associated with epilepsy in steroid-responsive encephalopathy, but have been rarely studied in unselected epilepsy patients. This study aimed to characterize the prevalence and associated factors of antithyroid Abs and other auto-Abs in adult patients with epilepsy. Epilepsy patients without autoimmune disorders were surveyed for antinuclear antibody (ANA), anti-β2 glycoprotein 1 antibody (aβ2GP1), anticardiolipin IgG Ab, antimicrosomal antibody (AMA), antithyroglobulin antibody (ATA), and thyroid function test. Of 319 patients, 75 (23.5%) were positive for at least 1 Ab. The most common Ab was anticardiolipin antibody (aCL) (30/319, 9.4%), followed by AMA (24/319, 7.5%), ANA (18/319, 5.6%), aβ2GP1 (18/319, 6.5%), and ATA (6/319, 3.25%). Antimicrosomal Abs were significantly more frequent in patients who were female, older at disease onset, older at the time of study, and had unknown seizure etiology. The presence of aCL was significantly associated with more frequent seizures. Most patients with antithyroid Ab were female and had focal seizures with unknown etiology. The association of different auto-Abs with different factors suggests that they may have different roles in adult patients with epilepsy. Recurrent seizures and certain antiepileptic medications may cause the production of aCL. The role of antithyroid Abs in adult focal epilepsy with unknown cause, especially in females, warrants further evaluation because of the potential implications on treatment. PMID:26131823

High titers of autoantibodies to glutamate decarboxylase in Type 1 Diabetes Patients: Epitope Analysis and Inhibition of Enzyme Activity

PubMed Central

Hampe, Christiane S.; Maitland, Murray E.; Gilliam, Lisa K.; Thi Phan, Thanh-H.; Sweet, Ian R.; Radtke, Jared R.; Bota, Vasile; Ransom, Bruce R.; Hirsch, Irl B.

2014-01-01

Objective Autoantibodies to glutamate decarboxylase (GAD65Ab) are found in patients with autoimmune neurological disorders and patients with type 1 diabetes. The correct diagnosis of GAD65Ab-associated neurological disorders is often delayed by the variability of symptoms and a lack of diagnostic markers. We hypothesize that the frequency of neurological disorders with high GAD65Ab titers is significantly higher than currently recognized. Methods We analyzed GAD65Ab titer, inhibition of GAD65 enzyme activity, and pattern of GAD65Ab epitopes in a cohort of type 1 diabetes patients (n=100) and correlated our findings with neurological symptoms and diseases. Results Fourty-three percent (43/100) of the patients had detectable GAD65Ab titers (median=400 U/ml, range: 142–250,000U/ml). The GAD65Ab titers in 10 type 1 diabetes patients exceeded the 90th percentile of the cohort (2,000–250,000 U/ml). Sera of these 10 patients were analyzed for their GAD65Ab epitope specificity and their ability to inhibit GAD65 enzyme activity in vitro. GAD65Ab of five patients inhibited the enzyme activity significantly (by 34–55%). Three of these patients complained of muscle stiffness and pain, which was documented in two of these patients. Conclusions Based on our findings we suggest that neurological disorders with high GAD65Ab titers are more frequent in type 1 diabetes patients than currently recognized. PMID:23512385

Impact of Pre-exposure History and Host Genetics on Antibody Avidity Following Norovirus Vaccination.

PubMed

Lindesmith, Lisa C; Mallory, Michael L; Jones, Taylor A; Richardson, Charles; Goodwin, Robert R; Baehner, Frank; Mendelman, Paul M; Bargatze, Robert F; Baric, Ralph S

2017-03-15

Development of high avidity, broadly neutralizing antibodies (Abs) is a priority after vaccination against rapidly evolving, widely disseminated viruses like human norovirus. After vaccination with a multivalent GI.1 and GII.4c norovirus virus-like particle (VLP) vaccine candidate adjuvanted with alum and monophosphoryl lipid A (MPL), blockade Ab titers peaked early, with no increase in titer following a second vaccine dose. Blockade Ab relative avidity was evaluated by measuring the slope of blockade Ab neutralization curves. Blockade Ab avidity to the GI.1 vaccine component peaked at day 35 (7 days after dose 2). Avidities to heterotypic genogroup I VLPs were not sustained at day 35 after vaccination or GI.1 infection, as measured from archived sera. Only secretor-positive participants maintained high avidity blockade Ab to GI.1 at day 180. Avidity to the GII.4c vaccine component peaked at day 7, remained elevated through day 180, and was not secretor dependent. Avidity to an immunologically novel GII.4 strain VLP correlated with preexisting Ab titer to an ancestral strain Epitope A. Host genetics and pre-exposure history shape norovirus vaccine Ab responses, including blockade Ab avidity. Avidity of potentially neutralizing Ab may be an important metric for evaluating vaccine responses to highly penetrant viruses with cross-reactive serotypes. © 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.

Mechanisms Mediating Enhanced Neutralization Efficacy of Staphylococcal Enterotoxin B by Combinations of Monoclonal Antibodies*

PubMed Central

Dutta, Kaushik; Varshney, Avanish K.; Franklin, Matthew C.; Goger, Michael; Wang, Xiaobo; Fries, Bettina C.

2015-01-01

Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-β chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used to validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Finally structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations. PMID:25572397

The structure and unconventional dihydrogen bonding of a pressure-stabilized hydrogen-rich (NH 3BH 3)(H 2) x(x = 1.5) compound

DOE PAGES

Lin, Yu; Welchman, Evan; Thonhauser, Timo; ...

2017-03-15

Combining X-ray diffraction, Raman spectroscopy, and ab initio simulations we characterize an extremely hydrogen-rich phase with the chemical formula (NH 3BH 3)(H 2) x (x = 1.5). This phase was formed by compressing ammonia borane (AB, NH 3BH 3) in an environment with an excess of molecular hydrogen (H 2). This compound can store a total of 26.8 wt% hydrogen, both as molecular hydrogen and chemically bonded hydrogen in AB, making it one of the most hydrogen-rich solids currently known. The new compound possesses a layered AB structure where additional H 2 molecules reside in channels created through the weavingmore » of AB layers. The unconventional dihydrogen bonding network of the new compound is significantly modified from its parent AB phase and contains H•••H contacts between adjacent AB molecules and between AB and H 2 molecules. H–H can be either a proton donor or a proton acceptor that forms new types of dihydrogen bonding with the host AB molecules, which are depicted as H–H•••H–B or H–H•••H–N, respectively. Furthermore, this study not only demonstrates the strategy and the promise of using pressure for new material synthesis, but also unleashes the power of combining experiments and ab initio calculations for elucidating novel structures and unusual bonding configurations in dense low-Z materials.« less

The structure and unconventional dihydrogen bonding of a pressure-stabilized hydrogen-rich (NH 3BH 3)(H 2) x(x = 1.5) compound

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

Lin, Yu; Welchman, Evan; Thonhauser, Timo

Combining X-ray diffraction, Raman spectroscopy, and ab initio simulations we characterize an extremely hydrogen-rich phase with the chemical formula (NH 3BH 3)(H 2) x (x = 1.5). This phase was formed by compressing ammonia borane (AB, NH 3BH 3) in an environment with an excess of molecular hydrogen (H 2). This compound can store a total of 26.8 wt% hydrogen, both as molecular hydrogen and chemically bonded hydrogen in AB, making it one of the most hydrogen-rich solids currently known. The new compound possesses a layered AB structure where additional H 2 molecules reside in channels created through the weavingmore » of AB layers. The unconventional dihydrogen bonding network of the new compound is significantly modified from its parent AB phase and contains H•••H contacts between adjacent AB molecules and between AB and H 2 molecules. H–H can be either a proton donor or a proton acceptor that forms new types of dihydrogen bonding with the host AB molecules, which are depicted as H–H•••H–B or H–H•••H–N, respectively. Furthermore, this study not only demonstrates the strategy and the promise of using pressure for new material synthesis, but also unleashes the power of combining experiments and ab initio calculations for elucidating novel structures and unusual bonding configurations in dense low-Z materials.« less

Mechanisms mediating enhanced neutralization efficacy of Staphylococcal enterotoxin B by combinations of monoclonal antibodies

DOE PAGES

Dutta, Kaushik; Varshney, Avanish K.; Franklin, Matthew C.; ...

2015-01-08

Staphylococcal enterotoxin B (SEB) is a superantigen that cross-links the major histocompatibility complex class II and specific V-β chains of the T-cell receptor, thus forming a ternary complex. Developing neutralizing mAb to disrupt the ternary complex and abrogate the resulting toxicity is a major therapeutic challenge because SEB is effective at very low concentrations. We show that combining two SEB-specific mAbs enhances their efficacy, even though one of the two mAbs by itself has no effect on neutralization. Crystallography was employed for fine-mapping conformational epitopes in binary and ternary complexes between SEB and Fab fragments. NMR spectroscopy was used tomore » validate and identify subtle allosteric changes induced by mAbs binding to SEB. The mapping of epitopes established that a combination of different mAbs can enhance efficacy of mAb-mediated protection from SEB induced lethal shock by two different mechanisms: one mAb mixture promoted clearance of the toxin both in vitro and in vivo by FcR-mediated cross-linking and clearance, whereas the other mAb mixture induced subtle allosteric conformational changes in SEB that perturbed formation of the SEB·T-cell receptor·major histocompatibility complex class II trimer. Lastly structural information accurately predicted mAb binding to other superantigens that share conformational epitopes with SEB. Fine mapping of conformational epitopes is a powerful tool to establish the mechanism and optimize the action of synergistic mAb combinations.« less

mAb C19 targets a novel surface marker for the isolation of human cardiac progenitor cells from human heart tissue and differentiated hESCs.

PubMed

Leung, Hau Wan; Moerkamp, Asja T; Padmanabhan, Jayanthi; Ng, Sze-Wai; Goumans, Marie-José; Choo, Andre

2015-05-01

Cardiac progenitor cells (CPCs) have been isolated from adult and developing hearts using an anti-mouse Sca-1 antibody. However, the absence of a human Sca-1 homologue has hampered the clinical application of the CPCs. Therefore, we generated novel monoclonal antibodies (mAbs) specifically raised against surface markers expressed by resident human CPCs. Here, we explored the suitability of one of these mAbs, mAb C19, for the identification, isolation and characterization of CPCs from fetal heart tissue and differentiating cultures of human embryonic stem cells (hESCs). Using whole-cell immunization, mAbs were raised against Sca-1+ CPCs and screened for reactivity to various CPC lines by flow cytometry. mAb C19 was found to be specific for Sca-1+ CPCs, with high cell surface binding capabilities. mAb C19 stained small stem-like cells in cardiac tissue sections. Moreover, during differentiation of hESCs towards cardiomyocytes, a transient population of cells with mAb C19 reactivity was identified and isolated using magnetic-activated cell sorting. Their cell fate was tracked and found to improve cardiomyocyte purity from hESC-derived cultures. mAb C19+ CPCs, from both hESC differentiation and fetal heart tissues, were maintained and expanded in culture, while retaining their CPC-like characteristics and their ability to further differentiate into cardiomyocytes by stimulation with TGFβ1. Finally, gene expression profiling of these mAb C19+ CPCs suggested a highly angiogenic nature, which was further validated by cell-based angiogenesis assays. mAb C19 is a new surface marker for the isolation of multipotent CPCs from both human heart tissues and differentiating hESCs. Copyright © 2015 Elsevier Ltd. All rights reserved.

Combination of Weight-Bearing Training and Anti-MSTN Polyclonal Antibody Improve Bone Quality In Rats.

PubMed

Tang, Liang; Gao, Xiaohang; Yang, Xiaoying; Zhang, Didi; Zhang, Xiaojun; Du, Haiping; Han, Yanqi; Sun, Lijun

2016-12-01

Weight-bearing exercise is beneficial to bone health. Myostatin (MSTN) deficiency has a positive effect on bone formation. We wondered if a combination of weight-bearing training and polyclonal antibody for MSTN (MsAb) would augment bone formation to a greater degree than single treatment. In this study, rats were randomly assigned to four groups: Control, weight-bearing training (WT), MsAb, and WT+MsAb. The trained rats ran at 15 m/min bearing with 35% of their body weight, 40 min/day (2 min of running followed by 2 min of rest), 6 days/week, for 8 weeks. The rats with MsAb were injected once a week with MsAb for 8 weeks. MicroCT analysis showed that compared with the MsAb group, WT+MsAb significantly enhanced cortical bone mineral density (BMD) (p < .01), bone volume over total volume (BV/TV) (p < .01), trabecular thickness (p < .05), and reduced trabecular separation (Tb.Sp) (p < .01). Compared with the WT group, WT+MsAb significantly increased trabecular BMD (p < .05), BV/TV (p < .05), and decreased Tb.Sp (p < .05). Three-point bending test demonstrated that MsAb failed to improve bone biomechanical properties (p > .05), weight-bearing training significantly increased energy absorption (p < .05) and elastic modulus (p < .05). However, when they combined, biomechanical properties including maximum load (p < .05), stiffness (p < .05), elastic modulus (p < .01) and energy absorption (p < .01) were all significantly enhanced. In conclusion, the combination of weight-bearing training and MsAb have a greater positive effect on bone than treatment with either MsAb or weight-bearing training alone, suggesting that resistance training in combination with MSTN antagonists could be an effective approach for improving bone health and reducing osteoporosis risk.

Modulation of immune response to Lol p I by pretreatment with anti-idiotypic antibody is not restricted to the idiotypic expression.

PubMed Central

Boutin, Y; Hébert, J

1994-01-01

To study the role of anti-idiotypic antibodies in the regulation of the immune response to Lol p I (the major allergenic component of rye grass pollen), we have recently generated a panel of three MoAbs directed against distinct epitopes of Lolp I and an anti-idiotypic MoAb directed against the idiotype borne by one of the anti-Lol p I MoAbs (290A-167). The effects of pretreatment with this anti-idiotypic MoAb in BALB/c mice before immunization with the antigen have been examined. The anti-idiotypic MoAb or unrelated MoAb were given weekly for 8 weeks intraperitoneally. Mice then received the antigen (2 micrograms) adsorbed with alum (2 mg) at weeks 9, 11 and 13. Serum anti-Lol p I antibodies (IgG or IgE) and specific idiotypic responses were measured. Anti-Lol p I IgG antibodies could be detected before immunization with Lol p I only in mice pretreated with anti-idiotypic MoAb. Immunization with Lol p I induced an anti-Lol p I IgG response in both groups, but this response was higher in mice that received anti-idiotypic MoAb. Similar profiles were seen for specific IgE antibodies and idiotypic responses. Surprisingly, idiotypes borne by other anti-Lol p I MoAbs (539A-6 and 348A-6) had also been enhanced after pretreatment with the anti-290A-167 MoAb. These observations suggested that the pretreatment with this anti-idiotypic MoAb modulates not only the expression of the respective idiotype, but also affects other idiotype responses. PMID:7514517

Self-Reported bruxism and associated factors in Israeli adolescents.

PubMed

Emodi Perlman, A; Lobbezoo, F; Zar, A; Friedman Rubin, P; van Selms, M K A; Winocur, E

2016-06-01

Little is known about the epidemiological characteristics of sleep and awake bruxism (SB and AB) in adolescents. The aims of the study were: to assess the prevalence rates of self-reported SB and AB in Israeli adolescents; to determine the associations between SB/AB and several demographical, exogenous and psychosocial factors in Israeli adolescents; and to investigate the possible concordance between SB and AB. The study made use of a questionnaire. The study population included 1000 students from different high schools in the centre of Israel. Prevalence of self-reported SB and AB in the Israeli adolescents studied was 9·2% and 19·2%, respectively. No gender difference was found regarding the prevalence of SB and AB. Multiple variable regression analysis revealed that the following predicting variables were related to SB: temporomandibular joint sounds (P = 0·002) and feeling stressed (P = 0·001). The following predicting variables were related to AB: age (P = 0·018), temporomandibular joint sounds (P = 0·002), oro-facial pain (P = 0·006), and feeling stressed (P = 0·002) or sad (P = 0·006). A significant association was found between SB and AB; that is, an individual reporting SB had a higher probability of reporting AB compared with an individual who did not report SB (odds ratio = 5·099). Chewing gum was the most common parafunction reported by adolescents. The results of this study demonstrate that self-reports of AB and SB are common in the Israeli adolescents population studied and are not related to gender. The significant correlation found between SB and AB may be a confounding bias that affects proper diagnosis of bruxism through self-reported questionnaires only. © 2016 John Wiley & Sons Ltd.

Detection of a novel specificity (CTLA-4) in ATG/TMG globulins and sera from ATG-treated leukemic patients.

PubMed

Pistillo, Maria Pia; Tazzari, Pier Luigi; Bonifazi, Francesca; Bandini, Giuseppe; Kato, Tomohiro; Matsui, Toshihiro; Nishioka, Kusuki; Conte, Roberto; Ferrara, Giovanni Battista

2002-04-27

T-cell costimulation has been shown to provide positive signals for T-cell activation and generation of effector activity. In this study, we analyzed the presence of antibodies (Abs) against the T-lymphocyte costimulatory molecules CD28, CTLA-4, CD80, and CD86 in anti-T-lymphocyte (ATG) and antithymocyte (TMG) globulin preparations to address their mechanism of action. We focused our attention on the role of CTLA-4-specific Abs in the immunosuppressive effect of ATG/TMG, because anti-CTLA-4 agonistic Abs may suppress T-cell proliferation and nonagonistic Abs may lead to T-cell depletion through an Ab-dependent cell cytotoxicity mechanism. ATG/TMG and patients' sera were tested for binding to recombinant human costimulatory molecules by ELISA techniques. CTLA-4 specificity was also analyzed by cytoplasmic immunofluorescence staining of a CTLA-4 transfectant by competitive inhibition immunofluorescence and by cell proliferation assay in allogeneic mixed lymphocyte reaction (MLR). Either ATG or TMG predominantly contained anti-CTLA-4 Abs, with higher reactivity in ATG followed by anti-CD86 and -CD28 Abs, whereas anti-CD80 Abs were found only in ATG. Anti-CTLA-4 Abs present in ATG/TMG recognized the native form of CTLA-4 molecule, and their removal reduced the effect of ATG in an allogeneic MLR. Kinetic studies indicated that such Abs were present in the sera of 12 ATG-treated leukemic patients up to 21 days after ATG administration. These data suggest that the novel anti-CTLA-4 Abs found in ATG may greatly contribute to its immunosuppressive effect, thus accounting for the absence of rejection and exceptionally low incidence of graft-versus-host disease in the group of patients analyzed.

Specificity and biologic activities of novel anti-membrane IgM antibodies

PubMed Central

Welt, Rachel S.; Welt, Jonathan A.; Kostyal, David; Gangadharan, Yamuna D; Raymond, Virginia; Welt, Sydney

2016-01-01

The concept that the B-cell Receptor (BCR) initiates a driver pathway in lymphoma-leukemia has been clinically validated. Previously described unique BCR Ig-class-specific sequences (proximal domains (PDs)), are not expressed in serum Ig (sIg). As a consequence of sequence and structural differences in the membrane IgM (mIgM) μ-Constant Domain 4, additional epitopes distinguish mIgM from sIgM. mAbs generated to linear and conformational epitopes, restricted to mIgM and not reacting with sIgM, were generated despite the relative hydrophobicity of the PDm sequence. Anti-PD mAbs (mAb1, mAb2, and mAb3) internalize mIgM. Anti-mIgM mAb4, which recognizes a distinct non-ligand binding site epitope, mediates mIgM internalization, and in low-density cultures, growth inhibition, anti-clonogenic activity, and apoptosis. We show that mAb-mediated mIgM internalization generally does not interrupt BCR-directed cell growth, however, mAb4 binding to a non-ligand binding site in the mIgM PDm-μC4 domain induces both mIgM internalization and anti-tumor effects. BCR micro-clustering in many B-cell leukemia and lymphoma lines is demonstrated by SEM micrographs using these new mAb reagents. mAb4 is a clinical candidate as a mediator of inhibition of the BCR signaling pathway. As these agents do not bind to non-mIgM B-cells, nor cross-react to non-lymphatic tissues, they may spare B-cell/normal tissue destruction as mAb-drug conjugates. PMID:27732950

Localization of key amino acid residues in the dominant conformational epitopes on thyroid peroxidase recognized by mouse monoclonal antibodies.

PubMed

Godlewska, Marlena; Czarnocka, Barbara; Gora, Monika

2012-09-01

Autoantibodies to thyroid peroxidase (TPO), the major target autoantigen in autoimmune thyroid diseases, recognize conformational epitopes limited to two immunodominant regions (IDRs) termed IDR-A and -B. The apparent restricted heterogeneity of TPO autoantibodies was discovered using TPO-specific mouse monoclonal antibodies (mAbs) and later confirmed by human recombinant Fabs. In earlier studies we identified key amino acids crucial for the interaction of human autoantibodies with TPO. Here we show the critical residues that participate in binding of five mAbs to the conformational epitopes on the TPO surface. Using ELISA we tested the reactivity of single and multiple TPO mutants expressed in CHO cells with a panel of mAbs specifically recognizing IDR-A (mAb 2 and 9) and IDR-B (mAb 15, 18, 64). We show that antibodies recognizing very similar regions on the TPO surface may interact with different sets of residues. We found that residues K713 and E716 contribute to the interaction between mAb 2 and TPO. The epitope for mAb 9 is critically dependent on residues R646 and E716. Moreover, we demonstrate that amino acids E604 and D630 are part of the functional epitope for mAb 15, and amino acids D624 and K627 for mAb 18. Finally, residues E604, D620, D624, K627, and D630 constitute the epitope for mAb 64. This is the first detailed study identifying the key resides for binding of mAbs 2, 9, 15, 18, and 64. Better understanding of those antibodies' specificity will be helpful in elucidating the properties of TPO as an antigen in autoimmune disorders.

Evaluation of the hemostatic and histopathological effects of Ankaferd Blood Stopper in experimental liver injury in rats.

PubMed

Satar, Nihal Y Gul; Akkoc, Ahmet; Oktay, Ayberk; Topal, Ayse; Inan, Kivanc

2013-07-01

We studied the hemostatic and histopathological effects, and intra-abdominal adhesion scores of a new hemostatic agent, Ankaferd Blood Stopper (ABS), in an experimental liver injury model and compared it with regenerated oxidized cellulose. Thirty-six rats were randomly assigned to ABS, oxidized cellulose (Surgicel), and control groups (n=12, each). A wedge resection was performed on the left medial lobe of the liver. In the ABS group the liver surface was sprayed with ABS, whereas in the Surgicel group the liver was covered with double-layered oxidized cellulose. In the control group, saline solution was sprayed on the cut surface. The mean bleeding time was shorter in the ABS (23.08±6.99s) and Surgicel groups (47.91±8.21s) than in the control group (223.42±57.83s). No significant difference was found in the ABS and Surgicel groups in terms of preoperative and postoperative hematocrit (hct) values (P>0.05). Whereas there was no significant difference on day 7 (P>0.05), total adhesion score of ABS group was lower than both Surgicel (P<0.05) and control groups (P<0.01) on day 14. Liver sections from ABS group displayed more favorable histopathological changes when compared with Surgicel group on day 7 and day 14. All livers in the ABS group completed their regeneration process with minimal signs of inflammation. Our findings suggest that ABS is more effective than Surgicel and control groups in achieving hemostasis and in reducing blood loss. Apart from this, ABS causes more encouraging histopathological changes and better intra-abdominal adhesion scores in rat experimental liver trauma model.

Characterization of human monoclonal antibodies that neutralize multiple poliovirus serotypes.

PubMed

Puligedda, Rama Devudu; Kouiavskaia, Diana; Al-Saleem, Fetweh H; Kattala, Chandana Devi; Nabi, Usman; Yaqoob, Hamid; Bhagavathula, V Sandeep; Sharma, Rashmi; Chumakov, Konstantin; Dessain, Scott K

2017-10-04

Following the eradication of wild poliovirus (PV), achieving and maintaining a polio-free status will require eliminating potentially pathogenic PV strains derived from the oral attenuated vaccine. For this purpose, a combination of non-cross-resistant drugs, such as small molecules and neutralizing monoclonal antibodies (mAbs), may be ideal. We previously isolated chimpanzee and human mAbs capable of neutralizing multiple PV types (cross-neutralization). Here, we describe three additional human mAbs that neutralize types 1 and 2 PV and one mAb that neutralizes all three types. Most bind conformational epitopes and have unusually long heavy chain complementarity determining 3 domains (HC CDR3). We assessed the ability of the mAbs to neutralize A12 escape mutant PV strains, and found that the neutralizing activities of the mAbs were disrupted by different amino acid substitutions. Competitive binding studies further suggested that the specific mAb:PV interactions that enable cross-neutralization differ among mAbs and serotypes. All of the cloned mAbs bind PV in the vicinity of the "canyon", a circular depression around the 5-fold axis of symmetry through which PV recognizes its cellular receptor. We were unable to generate escape mutants to two of the mAbs, suggesting that their epitopes are important for the PV life cycle. These data indicate that PV cross-neutralization involves binding to highly conserved structures within the canyon that binds to the cellular receptor. These may be facilitated by the long HC CDR3 domains, which may adopt alternative binding configurations. We propose that the human and chimpanzee mAbs described here could have potential as anti-PV therapeutics. Copyright © 2017 Elsevier Ltd. All rights reserved.

Pharmacokinetics and pharmacokinetic-pharmacodynamic relationships of monoclonal antibodies in children.

PubMed

Edlund, Helena; Melin, Johanna; Parra-Guillen, Zinnia P; Kloft, Charlotte

2015-01-01

Monoclonal antibodies (mAbs) constitute a therapeutically and economically important drug class with increasing use in both adult and paediatric patients. The rather complex pharmacokinetic and pharmacodynamic properties of mAbs have been extensively reviewed in adults. In children, however, limited information is currently available. This paper aims to comprehensively review published pharmacokinetic and pharmacokinetic-pharmacodynamic studies of mAbs in children. The current status of mAbs in the USA and in Europe is outlined, including a critical discussion of the dosing strategies of approved mAbs. The pharmacokinetic properties of mAbs in children are exhaustively summarised along with comparisons to reports in adults: for each pharmacokinetic process, we discuss the general principles and mechanisms of the pharmacokinetic/pharmacodynamic characteristics of mAbs, as well as key growth and maturational processes in children that might impact these characteristics. Throughout this review, considerable knowledge gaps are identified, especially regarding children-specific properties that influence pharmacokinetics, pharmacodynamics and immunogenicity. Furthermore, the large heterogeneity in the presentation of pharmacokinetic/pharmacodynamic data limited clinical inferences in many aspects of paediatric mAb therapy. Overall, further studies are needed to fully understand the impact of body size and maturational changes on drug exposure and response. To maximise future knowledge gain, we propose a 'Guideline for Best Practice' on how to report pharmacokinetic and pharmacokinetic-pharmacodynamic results from mAb studies in children which also facilitates comparisons. Finally, we advocate the use of more sophisticated modelling strategies (population analysis, physiology-based approaches) to appropriately characterise pharmacokinetic-pharmacodynamic relationships of mAbs and, thus, allow for a more rational use of mAb in the paediatric population.

Prevalence of migraine and Raynaud phenomenon in women with apical ballooning syndrome (Takotsubo or stress cardiomyopathy).

PubMed

Scantlebury, Dawn C; Prasad, Abhiram; Rabinstein, Alejandro A; Best, Patricia J M

2013-05-01

Apical ballooning syndrome (ABS), migraine, and Raynaud phenomenon are characterized by female predominance, identifiable triggers, and, likely, vascular dysfunction. Estrogen deficiency, such as in menopause, is considered to be a predisposing factor for ABS. We investigated the association of ABS with migraine, Raynaud phenomenon, and hormonal factors. We compared 25 consecutive residents (all women) of Olmsted County, Minnesota, presenting with ABS, to 2 age-matched control groups from the same community: 25 women presenting with ST-segment elevation myocardial infarction (STEMI), matched for the index ABS event date, and 50 women with neither diagnosis. The patients with ABS were more likely to have a migraine history (11 [44%] vs 4 [16%] STEMI controls, p = 0.031, and vs 6 [12%] population controls, p = 0.003), and "possible migraine" (including other headache syndromes suggestive of migraine; 15 [60%] vs 6 [24%] STEMI controls, p = 0.012; and vs 12 [24%] population controls, p = 0.003). Of the patients with ABS, 4 (16%) had Raynaud phenomenon compared to no STEMI controls and 1 (2%) population control (p = 0.038). No differences were present in parity, menopausal status, years since the onset of menopause, and frequency of oophorectomy. Current hormonal replacement therapy use was greater in those with ABS than in the population controls: 4 (16%) versus none (p = 0.002). In conclusion, the association of ABS with migraine and Raynaud phenomenon supports a role of vasomotor dysfunction in the pathogenesis of ABS. The absence of an association with reproductive characteristics and the finding that ABS occurred despite exogenous hormonal use in some patients suggests that estrogen deficiency per se is not the primary factor in the pathophysiology. Copyright © 2013 Elsevier Inc. All rights reserved.

Trehalose metabolism genes render rice white tip nematode Aphelenchoides besseyi (Nematoda: Aphelenchoididae) resistant to an anaerobic environment

PubMed Central

Chen, Qiaoli; Zhang, Ruizhi; Ling, Yaming

2018-01-01

ABSTRACT After experiencing anaerobic environments, Aphelenchoides besseyi will enter a state of suspended animation known as anoxybiosis, during which it may use trehalose as an energy supply to survive. To explore the function of trehalose metabolism, two trehalose-6-phosphate synthase (TPS) genes (Ab-tps1 and Ab-tps2) encoding enzymes catalysing trehalose synthesis, and three trehalase (TRE) genes (Ab-ntre1, Ab-ntre2 and Ab-atre) encoding enzymes catalysing the hydrolysis of trehalose, were identified and investigated. Ab-tps1 and Ab-tps2 were active during certain periods of anoxybiosis for A. besseyi, and Ab-tps2, Ab-ntre1, Ab-ntre2 and Ab-atre were active during certain periods of recovery. The results of RNA interference experiments suggested that TRE genes regulated each other and both TPS genes, while a single TPS gene only regulated the other TPS gene. However, two TPS genes together could regulate TRE genes, which indicated a feedback mechanism between these genes. All these genes also positively regulated the survival and resumption of active metabolism of the nematode. Genes functioning at re-aeration have a greater impact on nematode survival, suggesting that these genes could play roles in anoxybiosis regulation, but may function within restricted time frames. Changes in trehalose levels matched changes in TRE activity during the anoxybiosis–re-aeration process, suggesting that trehalose may act as an energy supply source. The observation of up-regulation of TPS genes during anoxybiosis suggested a possible signal role of trehalose. Trehalose metabolism genes could also work together to control trehalose levels at a certain level when the nematode is under anaerobic conditions. PMID:29158222

Differentiation of Strains of Infectious Bovine Rhinotracheitis Virus by Neutralization Kinetics with Late 19S Rabbit Antibodies

PubMed Central

Potgieter, Leon N. D.; Maré, C. John

1974-01-01

Two vaccine, two respiratory (infectious bovine rhinotracheitis [IBR]), and two genital (infectious pustular vulvovaginitis [IPV]) strains of infectious bovine rhinotracheitis virus were compared by neutralization kinetics using late 19S antibody (AB). The two vaccine strains were indistinguishable from one another, but were neutralized far more rapidly than the other four strains when either anti-IBR or anti-IPV 19S AB was used. The two IPV strains were indistinguishable from one another, but were neutralized significantly more rapidly than the two IBR strains when anti-IBR 19S AB was used. The 2 IBR strains were neutralized at a similar rate with the latter globulin preparation. Almost identical results were obtained with anti-IPV 19S AB, except that one IPV strain was neutralized at a rate similar to the IBR strains. However, when early and late rabbit 7S AB were used, IBR strains could not be distinguished from IPV strains by neutralization kinetics. Preliminary experiments indicated that both early and late 19S rabbit antibodies neutralized the homologous strain more rapidly than the heterologous strain, but the difference was more noticeable with late 19S AB. It was also determined that neutralization of IBR-IPV virus by specific early and late 19S rabbit AB and early 7S rabbit AB was markedly enhanced by guinea pig complement. Neutralization of this virus by late 7S AB, however, was only slightly enhanced by complement. These results suggest that vaccine strains of IBR-IPV virus may be distinguished by neutralization kinetics with late 19S rabbit AB, and that genital and respiratory strains may possibly also be distinguishable with some 19S AB preparations. Images PMID:4372173

Specificity and biologic activities of novel anti-membrane IgM antibodies.

PubMed

Welt, Rachel S; Welt, Jonathan A; Kostyal, David; Gangadharan, Yamuna D; Raymond, Virginia; Welt, Sydney

2016-11-15

The concept that the B-cell Receptor (BCR) initiates a driver pathway in lymphoma-leukemia has been clinically validated. Previously described unique BCR Ig-class-specific sequences (proximal domains (PDs)), are not expressed in serum Ig (sIg). As a consequence of sequence and structural differences in the membrane IgM (mIgM) µ-Constant Domain 4, additional epitopes distinguish mIgM from sIgM. mAbs generated to linear and conformational epitopes, restricted to mIgM and not reacting with sIgM, were generated despite the relative hydrophobicity of the PDm sequence. Anti-PD mAbs (mAb1, mAb2, and mAb3) internalize mIgM. Anti-mIgM mAb4, which recognizes a distinct non-ligand binding site epitope, mediates mIgM internalization, and in low-density cultures, growth inhibition, anti-clonogenic activity, and apoptosis. We show that mAb-mediated mIgM internalization generally does not interrupt BCR-directed cell growth, however, mAb4 binding to a non-ligand binding site in the mIgM PDm-μC4 domain induces both mIgM internalization and anti-tumor effects. BCR micro-clustering in many B-cell leukemia and lymphoma lines is demonstrated by SEM micrographs using these new mAb reagents. mAb4 is a clinical candidate as a mediator of inhibition of the BCR signaling pathway. As these agents do not bind to non-mIgM B-cells, nor cross-react to non-lymphatic tissues, they may spare B-cell/normal tissue destruction as mAb-drug conjugates.

The AB Doradus system revisited: The dynamical mass of AB Dor A/C

NASA Astrophysics Data System (ADS)

Azulay, R.; Guirado, J. C.; Marcaide, J. M.; Martí-Vidal, I.; Ros, E.; Tognelli, E.; Jauncey, D. L.; Lestrade, J.-F.; Reynolds, J. E.

2017-10-01

Context. The study of pre-main-sequence (PMS) stars with model-independent measurements of their masses is essential to check the validity of theoretical models of stellar evolution. The well-known PMS binary AB Dor A/C is an important benchmark for this task, since it displays intense and compact radio emission, which makes possible the application of high-precision astrometric techniques to this system. Aims: We aim to revisit the dynamical masses of the components of AB Dor A/C to refine earlier comparisons between the measurements of stellar parameters and the predictions of stellar models. Methods: We observed in phase-reference mode the binary AB Dor A/C, 0.2'' separation, with the Australian Long Baseline Array at 8.4 GHz. The astrometric information resulting from our observations was analyzed along with previously reported VLBI, optical (Hipparcos), and infrared measurements. Results: The main star AB Dor A is clearly detected in all the VLBI observations, which allowed us to analyze the orbital motion of the system and to obtain model-independent dynamical masses of 0.90 ± 0.08 M⊙ and 0.090 ± 0.008 M⊙, for AB Dor A and AB Dor C, respectively. Comparisons with PMS stellar evolution models favor and age of 40-50 Myr for AB Dor A and of 25-120 Myr for AB Dor C. Conclusions: We show that the orbital motion of the AB Dor A/C system is remarkably well determined, leading to precise estimates of the dynamical masses. Comparison of our results with the prediction of evolutionary models support the observational evidence that theoretical models tend to slightly underestimate the mass of the low-mass stars.

Production and characterization of monoclonal antibodies against conserved epitopes of P-selectin (CD62P).

PubMed

Massaguer, A; Engel, P; Pérez-del-Pulgar, S; Bosch, J; Pizcueta, P

2000-08-01

P-selectin (CD62P) is an adhesion molecule expressed on the activated endothelium and activated platelets that is involved in the initial attachment of leukocytes to inflamed vascular endothelium. Blocking monoclonal antibodies (mAbs) and P-selectin-deficient mice have shown that P-selectin is a potential target in anti-inflammatory therapy. Most mAbs against P-selectin do not bind to conserved epitopes, including the ligand-binding region, since P-selectin from mammalian species shares high amino acid sequence homology. The aim of this study was to generate a novel panel of anti-P-selectin mAbs against the conserved epitopes present in several animal species. To produce these mAbs, P-selectin-deficient mice were immunized with a pre-B-cell line transfected with human P-selectin cDNA. Twelve mouse mAbs that recognize human P-selectin were obtained. Individual mAbs that bound to human, rat, mouse, rabbit and pig activated platelets were characterized by flow-cytometry, immunohistochemistry, adhesion assays and immunoprecipitation. Four of these mAbs (P-sel.KO.2.3, P-sel.KO.2.4, P-sel.KO.2.7 and P-sel.KO.2.12) cross-reacted with human, rat and mouse P-selectin. Another three mAbs (P-sel.KO.2.2, P-sel.KO.2.11 and P-sel.KO.2.12) blocked the attachment of HL60 cells to P-selectin-transfected COS cells, demonstrating that these mAbs inhibit P-selectin-mediated adhesion. MAb cross-blocking experiments showed that these three mAbs bind to very close and overlapping epitopes. An ELISA assay using mAbs P-sel.KO.2.3 and P-sel.KO.2.12 was designed to measure soluble rat, mouse and human P-selectin. These anti-P-selectin mAbs are unique since they recognize common epitopes conserved during mammalian evolution and they may be useful for studying P-selectin function in inflammatory models in various species.

Fc functional antibodies in humans with severe H7N9 and seasonal influenza

PubMed Central

Vanderven, Hillary A.; Liu, Lu; Ana-Sosa-Batiz, Fernanda; Nguyen, Thi H.O.; Wan, Yanmin; Hogarth, P. Mark; Tilmanis, Danielle; Parsons, Matthew S.; Hurt, Aeron C.; Davenport, Miles P.; Kotsimbos, Tom; Cheng, Allen C.; Kedzierska, Katherine; Zhang, Xiaoyan; Xu, Jianqing; Kent, Stephen J.

2017-01-01

BACKGROUND. Both seasonal and novel avian influenza viruses can result in severe infections requiring hospitalization. Anti-influenza antibodies (Abs) with Fc-mediated effector functions, such as Ab-dependent cellular cytotoxicity (ADCC), are of growing interest in control of influenza but have not previously been studied during severe human infections. As such, the objective of this study was to examine Fc-mediated Ab functions in humans hospitalized with influenza infection. METHODS. Serum Ab response was studied in subjects hospitalized with either pandemic H7N9 avian influenza virus in China (n = 18) or circulating seasonal influenza viruses in Melbourne, Australia (n = 16). Recombinant soluble Fc receptor dimer ELISAs, natural killer (NK) cell activation assays, and Ab-dependent killing assays with influenza-infected target cells were used to assess the Fc functionality of anti-influenza hemagglutinin (HA) Abs during severe human influenza infection. RESULTS. We found that the peak generation of Fc functional HA Abs preceded that of neutralizing Abs for both severe H7N9 and seasonal influenza infections. Subjects who succumbed to complications of H7N9 infection demonstrated reduced HA-specific Fc receptor–binding Abs (in magnitude and breadth) immediately prior to death compared with those who survived. Subjects who recovered from H7N9 and severe seasonal influenza infections demonstrated increased Fc receptor–binding Abs not only against the homologous infecting strain but against HAs from different influenza A subtypes. CONCLUSION. Collectively, survivors of severe influenza infection rapidly generate a functional Ab response capable of mediating ADCC against divergent influenza viruses. Broadly binding HA Abs with Fc-mediated functions may be a useful component of protective immunity to severe influenza infection. FUNDING. The National Health and Medical Research Council ([NHMRC] grants 1023294, 1041832, and 1071916), the Australian Department of Health, and the joint University of Melbourne/Fudan University International Research and Research Training Fund provided funding for this study. PMID:28679958

Structural and Biophysical Characterization of Bacillus thuringiensis Insecticidal Proteins Cry34Ab1 and Cry35Ab1

PubMed Central

Kelker, Matthew S.; Berry, Colin; Evans, Steven L.; Pai, Reetal; McCaskill, David G.; Wang, Nick X.; Russell, Joshua C.; Baker, Matthew D.; Yang, Cheng; Pflugrath, J. W.; Wade, Matthew; Wess, Tim J.; Narva, Kenneth E.

2014-01-01

Bacillus thuringiensis strains are well known for the production of insecticidal proteins upon sporulation and these proteins are deposited in parasporal crystalline inclusions. The majority of these insect-specific toxins exhibit three domains in the mature toxin sequence. However, other Cry toxins are structurally and evolutionarily unrelated to this three-domain family and little is known of their three dimensional structures, limiting our understanding of their mechanisms of action and our ability to engineer the proteins to enhance their function. Among the non-three domain Cry toxins, the Cry34Ab1 and Cry35Ab1 proteins from B. thuringiensis strain PS149B1 are required to act together to produce toxicity to the western corn rootworm (WCR) Diabrotica virgifera virgifera Le Conte via a pore forming mechanism of action. Cry34Ab1 is a protein of ∼14 kDa with features of the aegerolysin family (Pfam06355) of proteins that have known membrane disrupting activity, while Cry35Ab1 is a ∼44 kDa member of the toxin_10 family (Pfam05431) that includes other insecticidal proteins such as the binary toxin BinA/BinB. The Cry34Ab1/Cry35Ab1 proteins represent an important seed trait technology having been developed as insect resistance traits in commercialized corn hybrids for control of WCR. The structures of Cry34Ab1 and Cry35Ab1 have been elucidated to 2.15 Å and 1.80 Å resolution, respectively. The solution structures of the toxins were further studied by small angle X-ray scattering and native electrospray ion mobility mass spectrometry. We present here the first published structure from the aegerolysin protein domain family and the structural comparisons of Cry34Ab1 and Cry35Ab1 with other pore forming toxins. PMID:25390338

The relationship between clinical characteristics, radiographic osteoarthritis and 3D bone area: data from the osteoarthritis initiative.

PubMed

Barr, A J; Dube, B; Hensor, E M A; Kingsbury, S R; Peat, G; Bowes, M A; Conaghan, P G

2014-10-01

Radiographic measures of osteoarthritis (OA) are based upon two dimensional projection images. Active appearance modelling (AAM) of knee magnetic resonance imaging (MRI) enables accurate, 3D quantification of joint structures in large cohorts. This cross-sectional study explored the relationship between clinical characteristics, radiographic measures of OA and 3D bone area (tAB). Clinical data and baseline paired radiographic and MRI data, from the medial compartment of one knee of 2588 participants were obtained from the NIH Osteoarthritis Initiative (OAI). The medial femur (MF) and tibia (MT) tAB were calculated using AAM. 'OA-attributable' tAB (OA-tAB) was calculated using data from regression models of tAB of knees without OA. Associations between OA-tAB and radiographic measures of OA were investigated using linear regression. In univariable analyses, height, weight, and age in female knees without OA explained 43.1%, 32.1% and 0.1% of the MF tAB variance individually and 54.4% when included simultaneously in a multivariable model. Joint space width (JSW), osteophytes and sclerosis explained just 5.3%, 14.9% and 10.1% of the variance of MF OA-tAB individually and 17.4% when combined. Kellgren Lawrence (KL) grade explained approximately 20% of MF OA-tAB individually. Similar results were seen for MT OA-tAB. Height explained the majority of variance in tAB, confirming an allometric relationship between body and joint size. Radiographic measures of OA, derived from a single radiographic projection, accounted for only a small amount of variation in 3D knee OA-tAB. The additional structural information provided by 3D bone area may explain the lack of a substantive relationship with these radiographic OA measures. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

GB Virus Type C Envelope Protein E2 Elicits Antibodies That React with a Cellular Antigen on HIV-1 Particles and Neutralize Diverse HIV-1 Isolates

PubMed Central

Mohr, Emma L.; Xiang, Jinhua; McLinden, James H.; Kaufman, Thomas M.; Chang, Qing; Montefiori, David C.; Klinzman, Donna; Stapleton, Jack T.

2012-01-01

Broadly neutralizing Abs to HIV-1 are well described; however, identification of Ags that elicit these Abs has proven difficult. Persistent infection with GB virus type C (GBV-C) is associated with prolonged survival in HIV-1–infected individuals, and among those without HIV-1 viremia, the presence of Ab to GBV-C glycoprotein E2 is also associated with survival. GBV-C E2 protein inhibits HIV-1 entry, and an antigenic peptide within E2 interferes with gp41-induced membrane perturbations in vitro, suggesting the possibility of structural mimicry between GBV-C E2 protein and HIV-1 particles. Naturally occurring human and experimentally induced GBV-C E2 Abs were examined for their ability to neutralize infectious HIV-1 particles and HIV-1–enveloped pseudovirus particles. All GBV-C E2 Abs neutralized diverse isolates of HIV-1 with the exception of rabbit anti-peptide Abs raised against a synthetic GBV-C E2 peptide. Rabbit anti–GBV-C E2 Abs neutralized HIV-1–pseudotyped retrovirus particles but not HIV-1–pseudotyped vesicular stomatitis virus particles, and E2 Abs immune-precipitated HIV-1 gag particles containing the vesicular stomatitis virus type G envelope, HIV-1 envelope, GBV-C envelope, or no viral envelope. The Abs did not neutralize or immune-precipitate mumps or yellow fever viruses. Rabbit GBV-C E2 Abs inhibited HIV attachment to cells but did not inhibit entry following attachment. Taken together, these data indicate that the GBV-C E2 protein has a structural motif that elicits Abs that cross-react with a cellular Ag present on retrovirus particles, independent of HIV-1 envelope glycoproteins. The data provide evidence that a heterologous viral protein can induce HIV-1–neutralizing Abs. PMID:20826757

Biochip-based instruments development for space exploration: influence of the antibody immobilization process on the biochip resistance to freeze-drying, temperature shifts and cosmic radiations

NASA Astrophysics Data System (ADS)

Coussot, G.; Moreau, T.; Faye, C.; Vigier, F.; Baqué, M.; Le Postollec, A.; Incerti, S.; Dobrijevic, M.; Vandenabeele-Trambouze, O.

2017-04-01

Due to the diversity of antibody (Ab)-based biochips chemistries available and the little knowledge about biochips resistance to space constraints, immobilization of Abs on the surface of the biochips dedicated to Solar System exploration is challenging. In the present paper, we have developed ten different biochip models including covalent or affinity immobilization with full-length Abs or Ab fragments. Ab immobilizations were carried out in oriented/non-oriented manner using commercial activated surfaces with N-hydroxysuccinic ester (NHS-surfaces) or homemade surfaces using three generations of dendrimers (dendrigraft of poly L-lysine (DGL) surfaces). The performances of the Ab -based surfaces were cross-compared on the following criteria: (i) analytical performances (expressed by both the surface density of immobilized Abs and the amount of antigens initially captured by the surface) and (ii) resistance of surfaces to preparation procedure (freeze-drying, storage) or spatial constraints (irradiation and temperature shifts) encountered during a space mission. The latter results have been expressed as percentage of surface binding capacity losses (or percentage of remaining active Abs). The highest amount of captured antigen was achieved with Ab surfaces having full-length Abs and DGL-surfaces that have much higher surface densities than commercial NHS-surface. After freeze-drying process, thermal shift and storage sample exposition, we found that more than 80% of surface binding sites remained active in this case. In addition, the resistance of Ab surfaces to irradiation with particles such as electron, carbon ions or protons depends not only on the chemistries (covalent/affinity linkages) and strategies (oriented/non-oriented) used to construct the biochip, but also on the type, energy and fluence of incident particles. Our results clearly indicate that full-length Ab immobilization on NHS-surfaces and DGL-surfaces should be preferred for potential use in instruments for planetary exploration.

Capacity of Broadly Neutralizing Antibodies to Inhibit HIV-1 Cell-Cell Transmission Is Strain- and Epitope-Dependent

PubMed Central

Reh, Lucia; Magnus, Carsten; Schanz, Merle; Weber, Jacqueline; Uhr, Therese; Rusert, Peter; Trkola, Alexandra

2015-01-01

An increasing number of broadly neutralizing antibodies (bnAbs) are considered leads for HIV-1 vaccine development and novel therapeutics. Here, we systematically explored the capacity of bnAbs to neutralize HIV-1 prior to and post-CD4 engagement and to block HIV-1 cell-cell transmission. Cell-cell spread is known to promote a highly efficient infection with HIV-1 which can inflict dramatic losses in neutralization potency compared to free virus infection. Selection of bnAbs that are capable of suppressing HIV irrespective of the transmission mode therefore needs to be considered to ascertain their in vivo activity in therapeutic use and vaccines. Employing assay systems that allow for unambiguous discrimination between free virus and cell-cell transmission to T cells, we probed a panel of 16 bnAbs for their activity against 11 viruses from subtypes A, B and C during both transmission modes. Over a wide range of bnAb-virus combinations tested, inhibitory activity against HIV-1 cell-cell transmission was strongly decreased compared to free virus transmission. Activity loss varied considerably between virus strains and was inversely associated with neutralization of free virus spread for V1V2- and V3-directed bnAbs. In rare bnAb-virus combinations, inhibition for both transmission modes was comparable but no bnAb potently blocked cell-cell transmission across all probed virus strains. Mathematical analysis indicated an increased probability of bnAb resistance mutations to arise in cell-cell rather than free virus spread, further highlighting the need to block this pathway. Importantly, the capacity to efficiently neutralize prior to CD4 engagement correlated with the inhibition efficacy against free virus but not cell-cell transmitted virus. Pre-CD4 attachment activity proved strongest amongst CD4bs bnAbs and varied substantially for V3 and V1V2 loop bnAbs in a strain-dependent manner. In summary, bnAb activity against divergent viruses varied depending on the transmission mode and differed depending on the window of action during the entry process, underscoring that powerful combinations of bnAbs are needed for in vivo application. PMID:26158270

Life-History Traits of Spodoptera frugiperda Populations Exposed to Low-Dose Bt Maize

PubMed Central

Sousa, Fernanda F.; Mendes, Simone M.; Santos-Amaya, Oscar F.; Araújo, Octávio G.; Oliveira, Eugenio E.

2016-01-01

Exposure to Bacillus thuringiensis (Bt) toxins in low- and moderate-dose transgenic crops may induce sublethal effects and increase the rate of Bt resistance evolution, potentially compromising control efficacy against target pests. We tested this hypothesis using the fall armyworm Spodoptera frugiperda, a major polyphagous lepidopteran pest relatively tolerant to Bt notorious for evolving field-relevant resistance to single-gene Bt maize. Late-instar larvae were collected from Bt Cry1Ab and non-Bt maize fields in five locations in Brazil, and their offspring was compared for survival, development, and population growth in rearing environment without and with Cry1Ab throughout larval development. Larval survival on Cry1Ab maize leaves varied from 20 to 80% among the populations. Larvae reared on Cry1Ab maize had seven-day delay in development time in relation to control larvae, and such delay was shorter in offspring of armyworms from Cry1Ab maize. Population growth rates were 50–70% lower for insects continuously exposed to Cry1Ab maize relative to controls, showing the population-level effect of Cry1Ab, which varied among the populations and prior exposure to Cry1Ab maize in the field. In three out of five populations, armyworms derived from Bt maize reared on Cry1Ab maize showed higher larval weight, faster larval development and better reproductive performance than the armyworms derived from non-Bt maize, and one of these populations showed better performance on both Cry1Ab and control diets, indicating no fitness cost of the resistance trait. Altogether, these results indicate that offspring of armyworms that developed on field-grown, single-gene Bt Cry1Ab maize had reduced performance on Cry1Ab maize foliage in two populations studied, but in other three populations, these offspring had better overall performance on the Bt maize foliage than that of the armyworms from non-Bt maize fields, possibly because of Cry1Ab resistance alleles in these populations. Implications of these findings for resistance management of S. frugiperda in Bt crops are discussed. PMID:27243977

Life-History Traits of Spodoptera frugiperda Populations Exposed to Low-Dose Bt Maize.

PubMed

Sousa, Fernanda F; Mendes, Simone M; Santos-Amaya, Oscar F; Araújo, Octávio G; Oliveira, Eugenio E; Pereira, Eliseu J G

2016-01-01

Exposure to Bacillus thuringiensis (Bt) toxins in low- and moderate-dose transgenic crops may induce sublethal effects and increase the rate of Bt resistance evolution, potentially compromising control efficacy against target pests. We tested this hypothesis using the fall armyworm Spodoptera frugiperda, a major polyphagous lepidopteran pest relatively tolerant to Bt notorious for evolving field-relevant resistance to single-gene Bt maize. Late-instar larvae were collected from Bt Cry1Ab and non-Bt maize fields in five locations in Brazil, and their offspring was compared for survival, development, and population growth in rearing environment without and with Cry1Ab throughout larval development. Larval survival on Cry1Ab maize leaves varied from 20 to 80% among the populations. Larvae reared on Cry1Ab maize had seven-day delay in development time in relation to control larvae, and such delay was shorter in offspring of armyworms from Cry1Ab maize. Population growth rates were 50-70% lower for insects continuously exposed to Cry1Ab maize relative to controls, showing the population-level effect of Cry1Ab, which varied among the populations and prior exposure to Cry1Ab maize in the field. In three out of five populations, armyworms derived from Bt maize reared on Cry1Ab maize showed higher larval weight, faster larval development and better reproductive performance than the armyworms derived from non-Bt maize, and one of these populations showed better performance on both Cry1Ab and control diets, indicating no fitness cost of the resistance trait. Altogether, these results indicate that offspring of armyworms that developed on field-grown, single-gene Bt Cry1Ab maize had reduced performance on Cry1Ab maize foliage in two populations studied, but in other three populations, these offspring had better overall performance on the Bt maize foliage than that of the armyworms from non-Bt maize fields, possibly because of Cry1Ab resistance alleles in these populations. Implications of these findings for resistance management of S. frugiperda in Bt crops are discussed.

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.

Field Trial Performance of Herculex XTRA (Cry34Ab1/Cry35Ab1) and SmartStax (Cry34Ab1/Cry35Ab1 + Cry3Bb1) Hybrids and Soil Insecticides Against Western and Northern Corn Rootworms (Coleoptera: Chrysomelidae).

PubMed

Johnson, K D; Campbell, L A; Lepping, M D; Rule, D M

2017-06-01

Western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), and northern corn rootworm, Diabrotica barberi Smith and Lawrence (Coleoptera: Chrysomelidae), are important insect pests in corn, Zea mays L. For more than a decade, growers have been using transgenic plants expressing proteins from the bacterium Bacillus thuringiensis (Bt) to protect corn roots from feeding. In 2011, western corn rootworm populations were reported to have developed resistance to Bt hybrids expressing Cry3Bb1 and later found to be cross-resistant to hybrids expressing mCry3A and eCry3.1Ab. The identification of resistance to Cry3 (Cry3Bb1, mCry3A, and eCry3.1Ab) hybrids led to concerns about durability and efficacy of products with single traits and of products containing a pyramid of a Cry3 protein and the binary Bt proteins Cry34Ab1 and Cry35Ab1. From 2012 to 2014, 43 field trials were conducted across the central United States to estimate root protection provided by plants expressing Cry34Ab1/Cry35Ab1 alone (Herculex RW) or pyramided with Cry3Bb1 (SmartStax). These technologies were evaluated with and without soil-applied insecticides to determine if additional management measures provided benefit where Cry3 performance was reduced. Trials were categorized for analysis based on rootworm damage levels on Cry3-expressing hybrids and rootworm feeding pressure within each trial. Across scenarios, Cry34Ab1/Cry35Ab1 hybrids provided excellent root protection. Pyramided traits provided greater root and yield protection than non-Bt plus a soil-applied insecticide, and only in trials where larval feeding pressure exceeded two nodes of damage did Cry34Ab1/Cry35Ab1 single-trait hybrids and pyramided hybrids show greater root protection from the addition of soil-applied insecticides. © The Authors 2017. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Glutamate receptor antibodies directed against AMPA receptors subunit 3 peptide B (GluR3B) can be produced in DBA/2J mice, lower seizure threshold and induce abnormal behavior.

PubMed

Ganor, Yonatan; Goldberg-Stern, Hadassa; Cohen, Ran; Teichberg, Vivian; Levite, Mia

2014-04-01

Anti-GluR3B antibodies (GluR3B Ab's), directed against peptide B/aa372-395 of GluR3 subunit of glutamate/AMPA receptors, are found in ∼35% of epilepsy patients, activate glutamate/AMPA receptors, evoke ion currents, kill neurons and damage the brain. We recently found that GluR3B Ab's also associate with neurological/psychiatric/behavioral abnormalities in epilepsy patients. Here we asked if GluR3B Ab's could be produced in DBA/2J mice, and also modulate seizure threshold and/or cause behavioral/motor impairments in these mice. DBA/2J mice were immunized with the GluR3B peptide in Complete Freund's Adjuvant (CFA), or with controls: ovalbumin (OVA), CFA, or phosphate-buffer saline (PBS). GluR3B Ab's and OVA Ab's were tested. Seizures were induced in all mice by the chemoconvulsant pentylenetetrazole (PTZ) at three time points, each time with less PTZ to avoid non-specific death. Behavior was examined in Open-Field, RotaRod and Grip tests. GluR3B Ab's were produced only in GluR3B-immunized mice, while OVA Ab's were produced only in OVA-immunized mice, showing high Ab's specificity. In GluR3B Ab's negative mice, seizure severity scores and percentages of animals developing generalized seizures declined in response to decreasing PTZ doses. In contrast, both parameters remained unchanged/high in the GluR3B Ab's positive mice, showing that these mice were more susceptible to seizures. The seizure scores associated significantly with the GluR3B Ab's levels. GluR3B Ab's positive mice were also more anxious in Open-Field test, fell faster in RotaRod test, and fell more in Grip test, compared to all the control mice. GluR3B Ab's are produced in DBA/2J mice, facilitate seizures and induce behavioral/motor impairments. This animal model can therefore serve for studying autoimmune epilepsy and abnormal behavior mediated by pathogenic anti-GluR3B Ab's. Copyright © 2014 Elsevier Ltd. All rights reserved.

Production and characterization of specific monoclonal antibodies binding the Plasmodium falciparum diagnostic biomarker, histidine-rich protein 2.

PubMed

Leow, Chiuan Herng; Jones, Martina; Cheng, Qin; Mahler, Stephen; McCarthy, James

2014-07-18

Early and accurate diagnosis of Plasmodium falciparum infection is important for providing appropriate treatment to patients with malaria. However, technical limitations of currently available diagnostic tests limit their use in control programs. One possible explanation for the vulnerability of current antibodies used in RDTs is their propensity to degrade at high ambient temperatures. Isolation of new antibodies with better thermal stability represents an appealing approach to improve the performance of RDTs. In this study, phage display technology was deployed to isolate novel binders by screening a human naïve scFv antibody library against recombinant Plasmodium falciparum histidine rich protein 2 (rPfHRP2). The isolated scFv clones were reformatted to whole IgG and the recombinant mAbs were produced in a mammalian CHO cell expression system. To verify the biological activity of these purified recombinant mAbs, range of functional assays were characterized. Two unique clones (D2 and F9) were isolated after five rounds of biopanning. The reformatted and expressed antibodies demonstrated high binding specificity to malaria recombinant PfHRP2 and native proteins. When 5 μg/mL of mAbs applied, mAb C1-13 had the highest sensitivity, with an OD value of 1, the detection achieved 5 ng/mL of rPfHRP2, followed by mAbs D2 and F9 at 10 ng/mL and 100 ng/mL of rPfHRP2, respectively. Although the sensitivity of mAbs D2 and F9 was lower than the control, these recombinant human mAbs have shown better stability compared to mouse mAb C1-13 at various temperatures in DSC and blot assays. In view of epitope mapping, the predominant motif of rPfHRP2 recognized by mAb D2 was AHHAADAHHA, whereas mAb F9 was one amino acid shorter, resulting in AHHAADAHH. mAb F9 had the strongest binding affinity to rPfHRP2 protein, with a KD value of 4.27 × 10(-11) M, followed by control mAb C1-13 at 1.03 × 10(-10) M and mAb D2 at 3.05 × 10(-10) M. Overall, the performance of these mAbs showed comparability to currently available PfHRP2-specific mouse mAb C1-13. The stability of these novel binders indicate that they merit further work to evaluate their utility in the development of new generation point of care diagnosis of malaria.

Epitope mapping of PfCP-2.9, an asexual blood-stage vaccine candidate of Plasmodium falciparum.

PubMed

Li, Changling; Wang, Rui; Wu, Yuan; Zhang, Dongmei; He, Zhicheng; Pan, Weiqing

2010-04-12

Apical membrane antigen 1 (AMA-1) and merozoite surface protein 1 (MSP1) of Plasmodium falciparum are two leading blood-stage malaria vaccine candidates. A P. falciparum chimeric protein 2.9 (PfCP-2.9) has been constructed as a vaccine candidate, by fusing AMA-1 domain III (AMA-1 (III)) with a C-terminal 19 kDa fragment of MSP1 (MSP1-19) via a 28-mer peptide hinge. PfCP-2.9 was highly immunogenic in animal studies, and antibodies elicited by the PfCP-2.9 highly inhibited parasite growth in vitro. This study focused on locating the distribution of epitopes on PfCP-2.9. A panel of anti-PfCP-2.9 monoclonal antibodies (mAbs) were produced and their properties were examined by Western blot as well as in vitro growth inhibition assay (GIA). In addition, a series of PfCP-2.9 mutants containing single amino acid substitution were produced in Pichia pastoris. Interaction of the mAbs with the PfCP-2.9 mutants was measured by both Western blot and enzyme-linked immunosorbent assay (ELISA). Twelve mAbs recognizing PfCP-2.9 chimeric protein were produced. Of them, eight mAbs recognized conformational epitopes and six mAbs showed various levels of inhibitory activities on parasite growth in vitro. In addition, seventeen PfCP-2.9 mutants with single amino acid substitution were produced in Pichia pastoris for interaction with mAbs. Reduced binding of an inhibitory mAb (mAb7G), was observed in three mutants including M62 (Phe491-->Ala), M82 (Glu511-->Gln) and M84 (Arg513-->Lys), suggesting that these amino acid substitutions are critical to the epitope corresponding to mAb7G. The binding of two non-inhibitory mAbs (mAbG11.12 and mAbW9.10) was also reduced in the mutants of either M62 or M82. The substitution of Leu31 to Arg resulted in completely abolishing the binding of mAb1E1 (a blocking antibody) to M176 mutant, suggesting that the Leu residue at this position plays a crucial role in the formation of the epitope. In addition, the Asn15 residue may also play an important role in the global folding of PfCP-2.9, as its substitution by Arg lead to reduced binding of most mAbs and abolishing the binding of mAb6G and mAbP5-W12. This study provided valuable information on epitopes of PfCP-2.9 vaccine candidate through generation of a panel of mAbs and a series of PfCP-2.9 mutants. The information may prove to be useful for designing more effective malaria vaccines against blood-stage parasites.

78 FR 15790 - Self-Regulatory Organizations; Financial Industry Regulatory Authority, Inc.; Order Granting...

Federal Register 2010, 2011, 2012, 2013, 2014

2013-03-12

... Security ``(ABS'') (except ABS traded To Be Announced (``TBA'')), in the limited instances when members... Factor in the limited instances when the member effects a transaction in an ABS (except a TBA transaction...-conforming Factor'').\\13\\ As a result of the proposed rule change, when an ABS transaction (except for a TBA...

Associative Interference and Recognition Memory.

ERIC Educational Resources Information Center

Underwood, Benton J.; And Others

Three experiments tested the generality of the conclusion that associative unlearning is minimal in the A-B, A-D paradigm. In Experiment 1, single-trial study of A-D, following single-trial study of A-B, did not produce retroactive inhibition in the recognition of A-B. In Experiment 2, A-B was acquired by associative matching. The interpolated…

A Phase Field Model of Deformation Twinning: Nonlinear Theory and Numerical Simulations

DTIC Science & Technology

2011-03-01

the system. Concepts for mod- eling multiphase systems were advanced by Steinbach et al. [3] and Steinbach and Apel [4]. Fried and Gurtin [5] and...a3b3 in a three- dimensional vector space. The outer product is (a ⊗ b) AB = aAbB. Juxtaposition implies summation over one set of adjacent indices...e.g., ( AB ) AB = AACBCB. The colon denotes summation over two sets of indices; e.g., A : B = AABBAB and (C : E) AB = CABCDECD. The transpose of amatrix is

Development of Anti-lock Braking System (ABS) for Vehicles Braking

NASA Astrophysics Data System (ADS)

Minh, Vu Trieu; Oamen, Godwin; Vassiljeva, Kristina; Teder, Leo

2016-11-01

This paper develops a real laboratory of anti-lock braking system (ABS) for vehicle and conducts real experiments to verify the ability of this ABS to prevent the vehicle wheel from being locked while braking. Two controllers of PID and fuzzy logic are tested for analysis and comparison. This ABS laboratory is designed for bachelor and master students to simulate and analyze performances of ABS with different control techniques on various roads and load conditions. This paper provides educational theories and practices on the design of control for system dynamics.

Commercial nickel-metal hydride (Ni-MH) technology evaluation

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

Erbacher, J.K.; Vukson, S.P.

1997-12-01

Available cylindrical and prismatic commercial Ni-MH batteries using AB{sub 5} and AB{sub 2} cathodes were evaluated for possible application to military aircraft batteries. Commercial AB{sub 5} technology is further advanced than AB{sub 2} technology and would require less alloy, electrolyte and single cell/battery development for near term (3--5 years) applications. Tested AB{sub 2} technology appears inadequate to meet the near term military requirements and would require a major development in the alloy to overcome the irreversible capacity loss at temperatures above 49 C.

The Effect of Fuel Composition, Compression Pressure, and Fuel-Air Ratio on the Compression-Ignition Characteristics of Several Fuels

DTIC Science & Technology

1948-03-01

to this arbitrary value as ntandard. The compression time was maintained unifcum by uElng a ccnstent . driving ~ essure of 500 pounds per squ=e inch...ir ratio,0.066 (chemlo~ correct); initial~ essure , 14.7 lb/sq in. abs.; intt Ml temperature, 609° F abs~ Ignitiondelay Compres- Compres- sion...chemically correct); initial ~ essure , 14.7 lb/sq in. abso; fimal pressures 379 lb/sq in. abs.; initial temperatume~ 6090 F abs.; final temperature, 1340° F abs

Passive Multistatic Detection of Maritime Targets using Opportunistic Radars

DTIC Science & Technology

2014-03-01

coordinate position for aa =1:1:length(Err_time1) Err_Total1( aa ) = max(abs(Err_time1( aa )),abs(Err_L1( aa ))+abs(Err_thetaR1( aa ))); end for aa =1...1:length(Err_time1) Err_Total1( aa ) = max(abs(Err_Total1( aa )),abs(Err_thetaR1( aa ))); end 93 %%%%%%%%%%%%%%%%%%%%% Tx 2...Err_Total2=zeros(size(Err_time2)); % Find th emax of the three source of error and use that for that % coordinate position for aa =1:1:length

Epitopes in α8β1 and other RGD-binding integrins delineate classes of integrin-blocking antibodies and major binding loops in α subunits

PubMed Central

Nishimichi, Norihisa; Kawashima, Nagako; Yokosaki, Yasuyuki

2015-01-01

Identification of epitopes for integrin-blocking monoclonal antibodies (mAbs) has aided our understanding of structure-function relationship of integrins. We mapped epitopes of chicken anti-integrin-α8-subunit-blocking mAbs by mutational analyses, examining regions that harboured all mapped epitopes recognized by mAbs against other α-subunits in the RGD-binding-integrin subfamily. Six mAbs exhibited blocking function, and these mAbs recognized residues on the same W2:41-loop on the top-face of the β-propeller. Loop-tips sufficiently close to W2:41 (<25 Å) contained within a footprint of the mAbs were mutated, and the loop W3:34 on the bottom face was identified as an additional component of the epitope of one antibody, clone YZ5. Binding sequences on the two loops were conserved in virtually all mammals, and that on W3:34 was also conserved in chickens. These indicate 1) YZ5 binds both top and bottom loops, and the binding to W3:34 is by interactions to conserved residues between immunogen and host species, 2) five other blocking mAbs solely bind to W2:41 and 3) the α8 mAbs would cross-react with most mammals. Comparing with the mAbs against the other α-subunits of RGD-integrins, two classes were delineated; those binding to “W3:34 and an top-loop”, and “solely W2:41”, accounting for 82% of published RGD-integrin-mAbs. PMID:26349930

Epitopes in α8β1 and other RGD-binding integrins delineate classes of integrin-blocking antibodies and major binding loops in α subunits.

PubMed

Nishimichi, Norihisa; Kawashima, Nagako; Yokosaki, Yasuyuki

2015-09-09

Identification of epitopes for integrin-blocking monoclonal antibodies (mAbs) has aided our understanding of structure-function relationship of integrins. We mapped epitopes of chicken anti-integrin-α8-subunit-blocking mAbs by mutational analyses, examining regions that harboured all mapped epitopes recognized by mAbs against other α-subunits in the RGD-binding-integrin subfamily. Six mAbs exhibited blocking function, and these mAbs recognized residues on the same W2:41-loop on the top-face of the β-propeller. Loop-tips sufficiently close to W2:41 (<25 Å) contained within a footprint of the mAbs were mutated, and the loop W3:34 on the bottom face was identified as an additional component of the epitope of one antibody, clone YZ5. Binding sequences on the two loops were conserved in virtually all mammals, and that on W3:34 was also conserved in chickens. These indicate 1) YZ5 binds both top and bottom loops, and the binding to W3:34 is by interactions to conserved residues between immunogen and host species, 2) five other blocking mAbs solely bind to W2:41 and 3) the α8 mAbs would cross-react with most mammals. Comparing with the mAbs against the other α-subunits of RGD-integrins, two classes were delineated; those binding to "W3:34 and an top-loop", and "solely W2:41", accounting for 82% of published RGD-integrin-mAbs.

Immunohistochemistry for the detection of neural and inflammatory cells in equine brain tissue

PubMed Central

Liu, Junjie; Herrington, Jenna M.; Vallario, Kelsey

2016-01-01

Phenotypic characterization of cellular responses in equine infectious encephalitides has had limited description of both peripheral and resident cell populations in central nervous system (CNS) tissues due to limited species-specific reagents that react with formalin-fixed, paraffin embedded tissue (FFPE). This study identified a set of antibodies for investigating the immunopathology of infectious CNS diseases in horses. Multiple commercially available staining reagents and antibodies derived from antigens of various species for manual immunohistochemistry (IHC) were screened. Several techniques and reagents for heat-induced antigen retrieval, non-specific protein blocking, endogenous peroxidase blocking, and visualization-detection systems were tested during IHC protocol development. Boiling of slides in a low pH, citrate-based buffer solution in a double-boiler system was most consistent for epitope retrieval. Pressure-cooking, microwaving, high pH buffers, and proteinase K solutions often resulted in tissue disruption or no reactivity. Optimal blocking reagents and concentrations of each working antibody were determined. Ultimately, a set of monoclonal (mAb) and polyclonal antibodies (pAb) were identified for CD3+ (pAb A0452, Dako) T-lymphocytes, CD79αcy+ B-lymphocytes (mAb HM57, Dako), macrophages (mAb MAC387, Leica), NF-H+ neurons (mAb NAP4, EnCor Biotechnology), microglia/macrophage (pAb Iba-1, Wako), and GFAP+ astrocytes (mAb 5C10, EnCor Biotechnology). In paraffin embedded tissues, mAbs and pAbs derived from human and swine antigens were very successful at binding equine tissue targets. Individual, optimized protocols are provided for each positively reactive antibody for analyzing equine neuroinflammatory disease histopathology. PMID:26855862

Effect of autogenous and fresh-frozen bone grafts on osteoblast differentiation.

PubMed

Ferraz, E P; Xavier, S P; Azevedo, F G; de Oliveira, F S; Beloti, M M; Rosa, A L

2015-01-01

Fresh-frozen bone allograft (FFBA) is an alternative to autogenous bone (AB) for reconstructing maxillary bone. Despite the promising clinical results, cell responses to FFBA and AB were not evaluated. Thus, our aim was to compare cells harvested from maxillary reconstructed sites with either AB or FFBA in terms of osteoblast differentiation and to evaluate the effect of culturing cells in contact with FFBA. Cells harvested from three patients submitted to bilateral maxillary reconstruction with AB and FFBA were cultured to evaluate: proliferation, alkaline phosphatase activity, extracellular matrix mineralization and gene expression of osteoblastic markers. The effect of FFBA on osteoblast differentiation was studied by culturing cells harvested from AB in contact with FFBA and evaluating the same parameters. Data were compared using either two-way ANOVA followed by Tukey-b test or Student's t test (p≤0.05). Cell proliferation was higher in cultures from AB grafted sites and extracellular matrix mineralization was higher in cultures derived from FFBA grafted sites. The gene expression of alkaline phosphatase, RUNX2, bone sialoprotein and osteocalcin was higher in cells derived from FFBA compared with cells from AB grafted sites. However, the exposure of cells derived from AB to FFBA particles did not have any remarkable effect on osteoblast differentiation. These results indicate the higher osteogenic activity of cells derived from FFBA compared with AB reconstructed sites, offering an explanation at cellular level of why FFBA could be a suitable alternative to AB for reconstructing maxillary bone defects. Copyright © 2014 Elsevier Ltd. All rights reserved.

Progress in the development of immunoanalytical methods incorporating recombinant antibodies to small molecular weight biotoxins.

PubMed

Kavanagh, Owen; Elliott, Christopher T; Campbell, Katrina

2015-04-01

Rapid immunoanalytical screening of food and environmental samples for small molecular weight (hapten) biotoxin contaminations requires the production of antibody reagents that possess the requisite sensitivity and specificity. To date animal-derived polyclonal (pAb) and monoclonal (mAb) antibodies have provided the binding element of the majority of these assays but recombinant antibodies (rAb) isolated from in vitro combinatorial phage display libraries are an exciting alternative due to (1) circumventing the need for experimental animals, (2) speed of production in commonly used in vitro expression systems and (3) subsequent molecular enhancement of binder performance. Short chain variable fragments (scFv) have been the most commonly employed rAb reagents for hapten biotoxin detection over the last two decades but antibody binding fragments (Fab) and single domain antibodies (sdAb) are increasing in popularity due to increased expression efficiency of functional binders and superior resistance to solvents. rAb-based immunochromatographic assays and surface plasmon resonance (SPR) biosensors have been reported to detect sub-regulatory levels of fungal (mycotoxins), marine (phycotoxins) and aquatic biotoxins in a wide range of food and environmental matrices, however this technology has yet to surpass the performances of the equivalent mAb- and pAb-based formats. As such the full potential of rAb technology in hapten biotoxin detection has yet to be achieved, but in time the inherent advantages of engineered rAb are set to provide the next generation of ultra-high performing binder reagents for the rapid and specific detection of hapten biotoxins.

The effect of arginine glutamate on the stability of monoclonal antibodies in solution.

PubMed

Kheddo, Priscilla; Tracka, Malgorzata; Armer, Jonathan; Dearman, Rebecca J; Uddin, Shahid; van der Walle, Christopher F; Golovanov, Alexander P

2014-10-01

Finding excipients which mitigate protein self-association and aggregation is an important task during formulation. Here, the effect of an equimolar mixture of l-Arg and l-Glu (Arg·Glu) on colloidal and conformational stability of four monoclonal antibodies (mAb1-mAb4) at different pH is explored, with the temperatures of the on-set of aggregation (Tagg) and unfolding (Tm1) measured by static light scattering and intrinsic fluorescence, respectively. Arg·Glu increased the Tagg of all four mAbs in concentration-dependent manner, especially as pH increased to neutral. Arg·Glu also increased Tm1 of the least thermally stable mAb3, but without similar direct effect on the Tm1 of other mAbs. Raising pH itself from 5 to 7 increased Tm1 for all four mAbs. Selected mAb formulations were assessed under accelerated stability conditions for the monomer fraction remaining in solution after storage. The aggregation of mAb3 was suppressed to a greater extent by Arg·Glu than by Arg·HCl. Furthermore, Arg·Glu suppressed the aggregation of mAb1 at neutral pH such that the fraction monomer was near to that at the more typical formulation pH of 5.5. We conclude that Arg·Glu can suppress mAb aggregation with increasing temperature/pH and, importantly, under accelerated stability conditions at weakly acidic to neutral pH. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Phage-Mediated Competitive Chemiluminescent Immunoassay for Detecting Cry1Ab Toxin by Using an Anti-Idiotypic Camel Nanobody.

PubMed

Qiu, Yulou; Li, Pan; Dong, Sa; Zhang, Xiaoshuai; Yang, Qianru; Wang, Yulong; Ge, Jing; Hammock, Bruce D; Zhang, Cunzheng; Liu, Xianjin

2018-01-31

Cry toxins have been widely used in genetically modified organisms for pest control, raising public concern regarding their effects on the natural environment and food safety. In this work, a phage-mediated competitive chemiluminescent immunoassay (c-CLIA) was developed for determination of Cry1Ab toxin using anti-idiotypic camel nanobodies. By extracting RNA from camels' peripheral blood lymphocytes, a naive phage-displayed nanobody library was established. Using anti-Cry1Ab toxin monoclonal antibodies (mAbs) against the library for anti-idiotypic antibody screening, four anti-idiotypic nanobodies were selected and confirmed to be specific for anti-Cry1Ab mAb binding. Thereafter, a c-CLIA was developed for detection of Cry1Ab toxin based on anti-idiotypic camel nanobodies and employed for sample testing. The results revealed a half-inhibition concentration of developed assay to be 42.68 ± 2.54 ng/mL, in the linear range of 10.49-307.1 ng/mL. The established method is highly specific for Cry1Ab recognition, with negligible cross-reactivity for other Cry toxins. For spiked cereal samples, the recoveries of Cry1Ab toxin ranged from 77.4% to 127%, with coefficient of variation of less than 9%. This study demonstrated that the competitive format based on phage-displayed anti-idiotypic nanobodies can provide an alternative strategy for Cry toxin detection.

Enhanced production of L-sorbose from D-sorbitol by improving the mRNA abundance of sorbitol dehydrogenase in Gluconobacter oxydans WSH-003.

PubMed

Xu, Sha; Wang, Xiaobei; Du, Guocheng; Zhou, Jingwen; Chen, Jian

2014-10-18

Production of L-sorbose from D-sorbitol by Gluconobacter oxydans is the first step to produce L-ascorbic acid on industrial scale. The sldhAB gene, which encodes the sorbitol dehydrogenase (SLDH), was overexpressed in an industrial strain G. oxydans WSH-003 with a strong promoter, P tufB . To enhance the mRNA abundance, a series of artificial poly(A/T) tails were added to the 3'-terminal of sldhAB gene. Besides, their role in sldhAB overexpression and their subsequent effects on L-sorbose production were investigated. The mRNA abundance of the sldhAB gene could be enhanced in G. oxydans by suitable poly(A/T) tails. By self-overexpressing the sldhAB gene in G. oxydans WSH-003 with an optimal poly(A/T) tail under the constitutive promoter P tufB , the titer and the productivity of L-sorbose were enhanced by 36.3% and 25.0%, respectively, in a 1-L fermenter. Immobilization of G. oxydans-sldhAB6 cells further improved the L-sorbose titer by 33.7% after 20 days of semi-continuous fed-batch fermentation. The artificial poly(A/T) tails could significantly enhance the mRNA abundance of the sldhAB. Immobilized G. oxydans-sldhAB6 cells could further enlarge the positive effect caused by enhanced mRNA abundance of the sldhAB.

Anti-p53 antibodies in sera from patients with chronic obstructive pulmonary disease can predate a diagnosis of cancer.

PubMed

Trivers, G E; De Benedetti, V M; Cawley, H L; Caron, G; Harrington, A M; Bennett, W P; Jett, J R; Colby, T V; Tazelaar, H; Pairolero, P; Miller, R D; Harris, C C

1996-10-01

Serum anti-p53 antibodies (p53-Abs) may be surrogate markers for both p53 alterations and preclinical cancer. Ancillary to a prospective trial to abate progressive development of clinical stages of chronic obstructive pulmonary disease, we conducted a retrospective, nested case-control study. Twenty-three cases were diagnosed with cancer during the trial. Enzyme immunoassay, immunoblotting, and immunoprecipitation were used to detect p53-Abs in serum, immunohistochemistry (IHC) to detect p53 accumulation, and single-strand conformation polymorphism and DNA sequencing to detect p53 mutations in tumor samples. p53-Abs were detected by three types of assays in five (23%) of the cancer patients, 80% of whom had detectable p53-Abs before diagnosis: 2 lung cancers (7 and 6 months before), 1 prostate cancer (11 months), and 1 breast cancer (5 months). Four Ab-positive patients had IHC-positive tumors. Two of 4 Ab-positive patients and 2 of 14 Ab-negative had p53 missense mutations or base pair deletion and IHC-positive tumors. The 44 noncancer COPD controls, matched with the cancer cases for age, gender, and smoking habits, were negative for p53-Abs. These results indicate that p53-Abs may facilitate the early diagnosis of cancer in a subset of smokers with chronic obstructive pulmonary disease who are at an increased cancer risk.

Generation and characterization of a human-mouse chimeric high-affinity antibody that detects the DYKDDDDK FLAG peptide.

PubMed

Ikeda, Koki; Koga, Tomoaki; Sasaki, Fumiyuki; Ueno, Ayumi; Saeki, Kazuko; Okuno, Toshiaki; Yokomizo, Takehiko

2017-05-13

DYKDDDDK peptide (FLAG) is a useful tool for investigating the function and localization of proteins whose antibodies (Abs) are not available. We recently established a high-affinity monoclonal antibody (mAb) for FLAG (clone 2H8). The 2H8 Ab is highly sensitive for detecting FLAG-tagged proteins by flowcytometry and immunoprecipitation, but it can yield nonspecific signals in immunohistochemistry of mouse tissues because it is of mouse origin. In this study, we reduced nonspecific signals by generating a chimeric 2H8 Ab with Fc fragments derived from human immunoglobulin. We fused a 5' terminal cDNA fragments for the Fab region of 2H8 mAb with 3' terminal cDNA fragments for Fc region of human IgG1. We transfected both chimeric plasmids and purified the resulting human-mouse chimeric 2H8. The chimeric 2H8 Ab successfully detected FLAG-tagged proteins in flowcytometry with anti-human IgG secondary Ab with comparable sensitivity to 2H8 mAb. Importantly, chimeric 2H8 detected specific FLAG peptide signals without nonspecific signals in immunohistochemical analysis with mouse tissues. This human-mouse chimeric high-affinity anti-FLAG Ab will prove useful for future immunohistochemical analysis of mouse tissues. Copyright © 2017 Elsevier Inc. All rights reserved.

Undoing Gender through Legislation and Schooling: The Case of AB 537 and AB 394 in California, USA

ERIC Educational Resources Information Center

Knotts, Greg

2009-01-01

This article investigates California laws AB 537: The Student Safety and Violence Prevention Act of 2000, and the recently enacted AB 394: Safe Place to Learn Act. Both demand that gender identity and sexual orientation be added to the lexicon of anti-harassment protection in public education. However, despite these progressive measures, schools…

Effects of goat manure liquid fertilizer combined with AB-MIX on foliage vegetables growth in hydroponic

NASA Astrophysics Data System (ADS)

Sunaryo, Y.; Purnomo, D.; Darini, M. T.; Cahyani, V. R.

2018-03-01

Hydroponic as one of the protected cultivation practices is very important to be developed in Indonesia due to not only the reduction of arable agricultural lands in lines with increasing of residential demand and other public facilities but also due to the negative influences of climate change as well global warming to plant growth. The effects of liquid fertilizer made from goat manure (LFGM) in combination with AB-Mix on three kinds of foliage vegetable growth was examined in hydroponics. The research was conducted by 3 x 4 factorial experiment and arranged in Completely Randomized Design with 3 replications. The first factor was foliage vegetable consisting of 3 levels: Mustard Green, Lettuce, and Red Spinach. The second factor was the mixture composition of nutrient solution consisting of 4 levels: LFGM + AB-Mix (v/v: 1:1), LFGM + AB-Mix (v/v: 1:3), LFGM + AB-Mix (v/v: 3:1), and A/B mix as control. Results indicated that the application of LFGM + AB-Mix (v/v: 1:3) resulted in similar plant growth as control (AB-Mix application), and also resulted in the highest chlorophyll content of Mustard green.

Prenatal diagnosis of amniotic band syndrome - risk factors and ultrasonic signs.

PubMed

Barzilay, Eran; Harel, Yael; Haas, Jigal; Berkenstadt, Michal; Katorza, Eldad; Achiron, Reuven; Gilboa, Yinon

2015-02-01

The aim of this study was to describe our experience with amniotic band syndrome (ABS), define specific sonographic characteristics and common features. Patients diagnosed with ABS underwent detailed ultrasound evaluation at the time of diagnosis and during follow-up. Their ultrasound examinations and medical records concerning the current pregnancy and past medical records were analyzed. Ten pregnancies were diagnosed with ABS. Most pregnancies were diagnosed at the beginning of the second trimester. Two cases were bichorionic twin pregnancies involving one of the fetuses and these were the only women who continued their pregnancies to term. The other eight cases with ABS chose to terminate their pregnancies. One pregnancy was conceived following trachelectomy. We found a significantly higher rate of prior uterine surgeries (p = 0.008) in patient with ABS compared to control. In three cases, all above 15 weeks of gestation, a small vestige at the distal part of the amputated limb was observed. ABS diagnosed in early pregnancy can be a sporadic event. However, there is a higher risk of ABS in pregnancies preceded by uterine procedures. The ultrasonic vestige sign at the amputated limb may contribute to the diagnosis of ABS.

Frontiers of monoclonal antibodies: Applications in medical practices.

PubMed

Ghagane, Shridhar C; Puranik, Sridevi I; Gan, Siew Hua; Hiremath, Murigendra B; Nerli, R B; Ravishankar, M V

2017-01-01

With the flourishing of innovation in drug discovery into a new era of personalized therapy, the use of monoclonal antibodies (mAbs) in the treatment of various ailments lies at the forefront. Major improvements in genetic sequencing and biomedical techniques as well as research into mAbs emphasize on determining new targets for advanced therapy while maximizing efficacy for clinical application. However, a balance has to be achieved concerning developing a target with low toxicity combined with high specificity and versatility, to allow a specific antibody to facilitate several biotic effects, ranging from neutralization of virus mechanisms to modulation of immune response and maintaining low global economic cost. Presently, there are approximately 30 mAbs' permitted for therapeutic use with many more being tested in clinical trials. Nevertheless, the heavy cost of mAbs' production, stowage and management as well as the subsequent hindrances to their development are outweighed by mAbs' clinical advantages. Compared to conventional drugs, since mAbs use as pharmacologic iotas have specific physical features and modes of action, they should be considered as a discrete therapeutic category. In this review, the history of mAb generation and the innovative technological applications of mAbs that has advanced in clinical practices is reviewed.

Separation of polycarbonate and acrylonitrile-butadiene-styrene waste plastics by froth flotation combined with ammonia pretreatment.

PubMed

Wang, Chong-Qing; Wang, Hui; Liu, Qun; Fu, Jian-Gang; Liu, You-Nian

2014-12-01

The objective of this research is flotation separation of polycarbonate (PC) and acrylonitrile-butadiene-styrene (ABS) waste plastics combined with ammonia pretreatment. The PC and ABS plastics show similar hydrophobicity, and ammonia treatment changes selectively floatability of PC plastic while ABS is insensitive to ammonia treatment. The contact angle measurement indicates the dropping of flotation recovery of PC is ascribed to a decline of contact angle. X-ray photoelectron spectroscopy demonstrates reactions occur on PC surface, which makes PC surface more hydrophilic. Separation of PC and ABS waste plastics was conducted based on the flotation behavior of single plastic. At different temperatures, PC and ABS mixtures were separated efficiently through froth flotation with ammonia pretreatment for different time (13 min at 23 °C, 18 min at 18 °C and 30 min at 23 °C). For both PC and ABS, the purity and recovery is more than 95.31% and 95.35%, respectively; the purity of PC and ABS is up to 99.72% and 99.23%, respectively. PC and ABS mixtures with different particle sizes were separated effectively, implying that ammonia treatment possesses superior applicability. Copyright © 2014 Elsevier Ltd. All rights reserved.

Observation of small cluster formation in concentrated monoclonal antibody solutions and its implications to solution viscosity.

PubMed

Yearley, Eric J; Godfrin, Paul D; Perevozchikova, Tatiana; Zhang, Hailiang; Falus, Peter; Porcar, Lionel; Nagao, Michihiro; Curtis, Joseph E; Gawande, Pradad; Taing, Rosalynn; Zarraga, Isidro E; Wagner, Norman J; Liu, Yun

2014-04-15

Monoclonal antibodies (mAbs) are a major class of biopharmaceuticals. It is hypothesized that some concentrated mAb solutions exhibit formation of a solution phase consisting of reversibly self-associated aggregates (or reversible clusters), which is speculated to be responsible for their distinct solution properties. Here, we report direct observation of reversible clusters in concentrated solutions of mAbs using neutron spin echo. Specifically, a stable mAb solution is studied across a transition from dispersed monomers in dilute solution to clustered states at more concentrated conditions, where clusters of a preferred size are observed. Once mAb clusters have formed, their size, in contrast to that observed in typical globular protein solutions, is observed to remain nearly constant over a wide range of concentrations. Our results not only conclusively establish a clear relationship between the undesirable high viscosity of some mAb solutions and the formation of reversible clusters with extended open structures, but also directly observe self-assembled mAb protein clusters of preferred small finite size similar to that in micelle formation that dominate the properties of concentrated mAb solutions. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

SdAb heterodimer formation using leucine zippers

NASA Astrophysics Data System (ADS)

Goldman, Ellen R.; Anderson, George P.; Brozozog-Lee, P. Audrey; Zabetakis, Dan

2013-05-01

Single domain antibodies (sdAb) are variable domains cloned from camel, llama, or shark heavy chain only antibodies, and are among the smallest known naturally derived antigen binding fragments. SdAb derived from immunized llamas are able to bind antigens with high affinity, and most are capable of refolding after heat or chemical denaturation to bind antigen again. We hypothesized that the ability to produce heterodimeric sdAb would enable reagents with the robust characteristics of component sdAb, but with dramatically improved overall affinity through increased avidity. Previously we had constructed multimeric sdAb by genetically linking sdAb that bind non-overlapping epitopes on the toxin, ricin. In this work we explored a more flexible approach; the construction of multivalent binding reagents using multimerization domains. We expressed anti-ricin sdAb that recognize different epitopes on the toxin as fusions with differently charged leucine zippers. When the initially produced homodimers are mixed the leucine zipper domains will pair to produce heterodimers. We used fluorescence resonance energy transfer to confirm heterodimer formation. Surface plasmon resonance, circular dichroism, enzyme linked immunosorbent assays, and fluid array assays were used to characterize the multimer constructs, and evaluate their utility in toxin detection.

What happens when you want to talk to the animals: lessons of a career in animal bioacoustics

NASA Astrophysics Data System (ADS)

Bowles, Ann E.

2004-05-01

Animal bioacoustics (AB) is the study of sound in nonhuman animal biology. I entered the field because I was interested in the evolution of language, and I wanted to study the acoustic communication of whales and dolphins. Topics like this within the scope of AB make the discipline accessible to students and laypeople. Although career opportunities are limited (professionals declaring AB as their primary area represent only 3% of ASA membership [http://www.acoustics.org/WIA, statistics for 2000]), an interest in AB can foster entry into more marketable disciplines. It has been a particularly important avenue for bringing women into careers in traditionally male-dominated subject areas. For example, women represent 14% of the ASA membership and 12% or fewer of those declaring Underwater Acoustics, Engineering Acoustics, and Noise as their primary interest. However, 25% of those declaring AB as their primary interest are women, and AB includes all three topic areas within its scope. Unfortunately, AB is still fairly inaccessible to interested lay professionals such as educators, science writers, and environmental planners. By helping them to develop a deeper understanding of topics in AB, ASA can help them foster careers in acoustics.

Molecular bases of protective immune responses against botulinum neurotoxin A--how antitoxin antibodies block its action.

PubMed

Atassi, M Zouhair; Dolimbek, Behzod Z; Steward, Lance E; Aoki, K Roger

2007-01-01

In studies from this laboratory, we localized the regions on the H chain of botulinum neurotoxin A (BoNT/A) that are recognized by anti-BoNT/A antibodies (Abs) and block the activity of the toxin in vivo. These Abs were obtained from cervical dystonia patients who had been treated with BoNT/A and had become unresponsive to the treatment, as well as blocking Abs raised in mouse, horse, and chicken. We also localized the regions involved in BoNT/A binding to mouse brain synaptosomes (snp). Comparison of spatial proximities in the three-dimensional structure of the Ab-binding regions and the snp binding showed that except for one, the Ab-binding regions either coincide or overlap with the snp regions. It should be folly expected that protective Abs when bound to the toxin at sites that coincide or overlap with snp binding would prevent the toxin from binding to nerve synapse and therefore block toxin entry into the neuron. Thus, analysis of the locations of the Ab-binding and the snp-binding regions provides a molecular rationale for the ability of protecting Abs to block BoNT/A action in vivo.

Diversion of HIV-1 Vaccine-induced Immunity by gp41-Microbiota Cross-reactive Antibodies

PubMed Central

Williams, Wilton B; Liao, Hua-Xin; Moody, M. Anthony; Kepler, Thomas B.; Alam, S Munir; Gao, Feng; Wiehe, Kevin; Trama, Ashley M.; Jones, Kathryn; Zhang, Ruijun; Song, Hongshuo; Marshall, Dawn J; Whitesides, John F; Sawatzki, Kaitlin; Hua, Axin; Liu, Pinghuang; Tay, Matthew Z; Seaton, Kelly; Shen, Xiaoying; Foulger, Andrew; Lloyd, Krissey E.; Parks, Robert; Pollara, Justin; Ferrari, Guido; Yu, Jae-Sung; Vandergrift, Nathan; Montefiori, David C.; Sobieszczyk, Magdalena E; Hammer, Scott; Karuna, Shelly; Gilbert, Peter; Grove, Doug; Grunenberg, Nicole; McElrath, Julie; Mascola, John R.; Koup, Richard A; Corey, Lawrence; Nabel, Gary J.; Morgan, Cecilia; Churchyard, Gavin; Maenza, Janine; Keefer, Michael; Graham, Barney S.; Baden, Lindsey R.; Tomaras, Georgia D.; Haynes, Barton F.

2015-01-01

A HIV-1 DNA prime-recombinant Adenovirus Type 5 (rAd5) boost vaccine failed to protect from HIV-1 acquisition. We studied the nature of the vaccine-induced antibody (Ab) response to HIV-1 envelope (Env). HIV-1-reactive plasma Ab titers were higher to Env gp41 than gp120, and repertoire analysis demonstrated that 93% of HIV-1-reactive Abs from memory B cells was to Env gp41. Vaccine-induced gp41-reactive monoclonal antibodies (mAbs) were non-neutralizing, and frequently polyreactive with host and environmental antigens including intestinal microbiota (IM). Next generation sequencing of an IGHV repertoire prior to vaccination revealed an Env-IM cross-reactive Ab that was clonally-related to a subsequent vaccine-induced gp41-reactive Ab. Thus, HIV-1 Env DNA-rAd5 vaccine induced a dominant IM-polyreactive, non-neutralizing gp41-reactive Ab repertoire response that was associated with no vaccine efficacy. PMID:26229114

Oriented conjugation of single-domain antibodies and quantum dots.

PubMed

Brazhnik, Kristina; Nabiev, Igor; Sukhanova, Alyona

2014-01-01

Nanoparticle-based biodetection routinely employs monoclonal antibodies (mAbs) for targeting. However, the large size of mAbs limits the number of ligands per nanoparticle and severely restricts the bioavailability and distribution of these probes in a sample. Furthermore, conventional conjugation techniques provide nanoprobes with irregular orientation of mAbs on the nanoparticle surface and often provoke mAb unfolding. Here, we describe a protocol for engineering a new generation of ultrasmall diagnostic nanoprobes through oriented conjugation of semiconductor quantum dots (QDs) with 13 kDa single-domain antibodies (sdAbs) derived from llama immunoglobulin G (IgG). The sdAbs are conjugated with QDs in a highly oriented manner via an additional cysteine residue specifically integrated into the sdAb C-terminus. The resultant nanoprobes are <12 nm in diameter, ten times smaller in volume compared to the known alternatives. They have been proved highly efficient in flow cytometry and immunuhistochemical diagnostics. This approach can be easily extended to other semiconductor and plasmonic nanoparticles.

Global remote sensing of water-chlorophyll ratio in terrestrial plant leaves.

PubMed

Kushida, Keiji

2012-10-01

I evaluated the use of global remote sensing techniques for estimating plant leaf chlorophyll a + b (C(ab); μg cm(-2)) and water (C(w); mg cm(-2)) concentrations as well as the ratio of C(w)/C(ab) with the PROSAIL model under possible distributions for leaf and soil spectra, leaf area index (LAI), canopy geometric structure, and leaf size. First, I estimated LAI from the normalized difference vegetation index. I found that, at LAI values <2, C(ab), C(w), and C(w)/C(ab) could not be reliably estimated. At LAI values >2, C(ab) and C(w) could be estimated for only restricted ranges of the canopy structure; however, the ratio of C(w)/C(ab) could be reliably estimated for a variety of possible canopy structures with coefficients of determination (R(2)) ranging from 0.56 to 0.90. The remote estimation of the C(w)/C(ab) ratio from satellites offers information on plant condition at a global scale.

Crystallin-αB Regulates Skeletal Muscle Homeostasis via Modulation of Argonaute2 Activity*

PubMed Central

Neppl, Ronald L.; Kataoka, Masaharu; Wang, Da-Zhi

2014-01-01

The core functional machinery of the RNAi pathway is the RNA-induced silencing complex (RISC), wherein Argonaute2 (Ago2) is essential for siRNA-directed endonuclease activity and RNAi/microRNA-mediated gene silencing. Crystallin-αB (CryAB) is a small heat shock protein involved in preventing protein aggregation. We demonstrate that CryAB interacts with the N and C termini of Ago2, not the catalytic site defined by the convergence of the PAZ, MID, and PIWI domains. We further demonstrate significantly reduced Ago2 activity in the absence of CryAB, highlighting a novel role of CryAB in the mammalian RNAi/microRNA pathway. In skeletal muscle of CryAB null mice, we observe a shift in the hypertrophy-atrophy signaling axis toward atrophy under basal conditions. Moreover, loss of CryAB altered the capability of satellite cells to regenerate skeletal muscle. These studies establish that CryAB is necessary for normal Ago2/RISC activity and cellular homeostasis in skeletal muscle. PMID:24782307

Development of Isaacs' syndrome following complete recovery of voltage-gated potassium channel antibody-associated limbic encephalitis.

PubMed

Takahashi, Hirokatsu; Mori, Masahiro; Sekiguchi, Yukari; Misawa, Sonoko; Sawai, Setsu; Hattori, Takamichi; Kuwabara, Satoshi

2008-12-15

Autoantibodies against voltage-gated potassium channels (VGKC-Abs) are associated with acquired neuromyotonia (Isaacs' syndrome) and related disorders such as Morvan's syndrome and some cases of limbic encephalitis. The mechanisms underlying the various phenotypes induced by VGKC-Abs are not fully understood. Recently, we reported a case of LE with VGKC-Abs accompanied by severe intestinal pseudo-obstruction and thymoma. Thymectomy and immunosuppressive therapy induced dramatic clinical improvement of LE symptoms, and VGKC-Abs titers decreased from 1254 pM to 549 pM (normal>100 pM). Seventeen months later, the patient developed progressive generalized muscle cramping, paresthesias in his lower extremities, excessive sweating, and severe constipation. There was no recurrence of the LE. Electromyography showed fasciculation potentials and myokymic discharges, and the plasma VGKC-Abs titer was again elevated to 879 pM. Here we report a case of Isaacs' syndrome after complete remission of LE with VGKC-Abs that may provide an insight into a possible link among VGKC-Abs associated syndromes.

HIV-1 broadly neutralizing antibody precursor B cells revealed by germline-targeting immunogen

DOE PAGES

Jardine, Joseph G.; Kulp, Daniel W.; Havenar-Daughton, Colin; ...

2016-03-25

Induction of broadly neutralizing antibodies (bnAbs) is a major HIV vaccine goal. Germline-targeting immunogens aim to initiate bnAb induction by activating bnAb germline precursor B cells. Critical unmet challenges are to determine whether bnAb precursor naïve B cells bind germline-targeting immunogens and occur at sufficient frequency in humans for reliable vaccine responses. We employed deep mutational scanning and multi-target optimization to develop a germline-targeting immunogen (eOD-GT8) for diverse VRC01-class bnAbs. We then used the immunogen to isolate VRC01-class precursor naïve B cells from HIV-uninfected donors. Frequencies of true VRC01-class precursors, their structures, and their eOD-GT8 affinities support this immunogen asmore » a candidate human vaccine prime. Lastly, these methods could be applied to germline targeting for other classes of HIV bnAbs and for Abs to other pathogens.« less

Voltage-Gated Potassium Channel Antibodies in Slow-Progression Motor Neuron Disease.

PubMed

Godani, Massimiliano; Zoccarato, Marco; Beronio, Alessandro; Zuliani, Luigi; Benedetti, Luana; Giometto, Bruno; Del Sette, Massimo; Raggio, Elisa; Baldi, Roberta; Vincent, Angela

2017-01-01

The spectrum of autoimmune neurological diseases associated with voltage-gated potassium channel (VGKC)-complex antibodies (Abs) ranges from peripheral nerve disorders to limbic encephalitis. Recently, low titers of VGKC-complex Abs have also been reported in neurodegenerative disorders, but their clinical relevance is unknown. The aim of the study was to explore the prevalence of VGKC-complex Abs in slow-progression motor neuron disease (MND). We compared 11 patients affected by slow-progression MND with 9 patients presenting typical progression illness. Sera were tested for VGKC-complex Abs by radioimmunoassay. The distribution of VGKC-complex Abs was analyzed with the Mann-Whitney U test. The statistical analysis showed a significant difference between the mean values in the study and control groups. A case with long-survival MND harboring VGKC-complex Abs and treated with intravenous immunoglobulins is described. Although VGKC-complex Abs are not likely to be pathogenic, these results could reflect the coexistence of an immunological activation in patients with slow disease progression. © 2016 S. Karger AG, Basel.

HIV-1 broadly neutralizing antibody precursor B cells revealed by germline-targeting immunogen

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

Jardine, Joseph G.; Kulp, Daniel W.; Havenar-Daughton, Colin

Induction of broadly neutralizing antibodies (bnAbs) is a major HIV vaccine goal. Germline-targeting immunogens aim to initiate bnAb induction by activating bnAb germline precursor B cells. Critical unmet challenges are to determine whether bnAb precursor naïve B cells bind germline-targeting immunogens and occur at sufficient frequency in humans for reliable vaccine responses. We employed deep mutational scanning and multi-target optimization to develop a germline-targeting immunogen (eOD-GT8) for diverse VRC01-class bnAbs. We then used the immunogen to isolate VRC01-class precursor naïve B cells from HIV-uninfected donors. Frequencies of true VRC01-class precursors, their structures, and their eOD-GT8 affinities support this immunogen asmore » a candidate human vaccine prime. Lastly, these methods could be applied to germline targeting for other classes of HIV bnAbs and for Abs to other pathogens.« less

Data and Analysis of the Double Stars STFA 10AB and STFA 1744AB

NASA Astrophysics Data System (ADS)

Arcilla, Marisa; Bowden, Sam; DeBlase, Jacqueline; Hall, Anthony; Hall, Corielyn; Hernandez, Alyssa; Renna, Danielle; Rodriguez, Fatima; Salazar, Cassandra; Sanchez, Andres; Teeter, Dayton; Brewer, Mark; Funk, Benjamin; Gillette, Travis; Sharpe, Scott

2017-04-01

Eighth grade students at Vanguard Preparatory School measured the double stars STFA 10AB and STFA 1744AB. A 22-inch Newtonian Alt/Az telescope and a 14-inch Celestron Schmidt Cassegrain telescope were used. The star Bellatrix was used as the calibration star to determine the scale constant of the 22-inch telescope to be 7.8 “/tick marks. The double star STFA 1744AB was used as the calibration star to determine the scale constant of the 14-inch telescope to be 5.1 “/tick marks. The separation and position angle of STFA 10AB was determined by the 22-inch telescope to be 347.9” and 339.3°. The separation and position angle of STFA 1744AB was determined by the 14-inch telescope to be 3.6” and 158.1°. The measurements that were calculated were compared to the most recent measurements listed in the Washington Double Star Catalog.

The growth and potential of human antiviral monoclonal antibody therapeutics.

PubMed

Marasco, Wayne A; Sui, Jianhua

2007-12-01

Monoclonal antibodies (mAbs) have long provided powerful research tools for virologists to understand the mechanisms of virus entry into host cells and of antiviral immunity. Even so, commercial development of human (or humanized) mAbs for the prophylaxis, preemptive and acute treatment of viral infections has been slow. This is surprising, as new antibody discovery tools have increased the speed and precision with which potent neutralizing human antiviral mAbs can be identified. As longstanding barriers to antiviral mAb development, such as antigenic variability of circulating viral strains and the ability of viruses to undergo neutralization escape, are being overcome, deeper insight into the mechanisms of mAb action and engineering of effector functions are also improving the efficacy of antiviral mAbs. These successes, in both industrial and academic laboratories, coupled with ongoing changes in the biomedical and regulatory environments, herald an era when the commercial development of human antiviral mAb therapies will likely surge.

Structural Constraints of Vaccine-Induced Tier-2 Autologous HIV Neutralizing Antibodies Targeting the Receptor-Binding Site.

PubMed

Bradley, Todd; Fera, Daniela; Bhiman, Jinal; Eslamizar, Leila; Lu, Xiaozhi; Anasti, Kara; Zhang, Ruijung; Sutherland, Laura L; Scearce, Richard M; Bowman, Cindy M; Stolarchuk, Christina; Lloyd, Krissey E; Parks, Robert; Eaton, Amanda; Foulger, Andrew; Nie, Xiaoyan; Karim, Salim S Abdool; Barnett, Susan; Kelsoe, Garnett; Kepler, Thomas B; Alam, S Munir; Montefiori, David C; Moody, M Anthony; Liao, Hua-Xin; Morris, Lynn; Santra, Sampa; Harrison, Stephen C; Haynes, Barton F

2016-01-05

Antibodies that neutralize autologous transmitted/founder (TF) HIV occur in most HIV-infected individuals and can evolve to neutralization breadth. Autologous neutralizing antibodies (nAbs) against neutralization-resistant (Tier-2) viruses are rarely induced by vaccination. Whereas broadly neutralizing antibody (bnAb)-HIV-Envelope structures have been defined, the structures of autologous nAbs have not. Here, we show that immunization with TF mutant Envs gp140 oligomers induced high-titer, V5-dependent plasma neutralization for a Tier-2 autologous TF evolved mutant virus. Structural analysis of autologous nAb DH427 revealed binding to V5, demonstrating the source of narrow nAb specificity and explaining the failure to acquire breadth. Thus, oligomeric TF Envs can elicit autologous nAbs to Tier-2 HIVs, but induction of bnAbs will require targeting of precursors of B cell lineages that can mature to heterologous neutralization. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

HIV-1 broadly neutralizing antibody precursor B cells revealed by germline-targeting immunogen.

PubMed

Jardine, Joseph G; Kulp, Daniel W; Havenar-Daughton, Colin; Sarkar, Anita; Briney, Bryan; Sok, Devin; Sesterhenn, Fabian; Ereño-Orbea, June; Kalyuzhniy, Oleksandr; Deresa, Isaiah; Hu, Xiaozhen; Spencer, Skye; Jones, Meaghan; Georgeson, Erik; Adachi, Yumiko; Kubitz, Michael; deCamp, Allan C; Julien, Jean-Philippe; Wilson, Ian A; Burton, Dennis R; Crotty, Shane; Schief, William R

2016-03-25

Induction of broadly neutralizing antibodies (bnAbs) is a major HIV vaccine goal. Germline-targeting immunogens aim to initiate bnAb induction by activating bnAb germline precursor B cells. Critical unmet challenges are to determine whether bnAb precursor naïve B cells bind germline-targeting immunogens and occur at sufficient frequency in humans for reliable vaccine responses. Using deep mutational scanning and multitarget optimization, we developed a germline-targeting immunogen (eOD-GT8) for diverse VRC01-class bnAbs. We then used the immunogen to isolate VRC01-class precursor naïve B cells from HIV-uninfected donors. Frequencies of true VRC01-class precursors, their structures, and their eOD-GT8 affinities support this immunogen as a candidate human vaccine prime. These methods could be applied to germline targeting for other classes of HIV bnAbs and for Abs to other pathogens. Copyright © 2016, American Association for the Advancement of Science.

Therapeutic antibodies against cancer

PubMed Central

Adler, Mark J.; Dimitrov, Dimiter S.

2012-01-01

Antibody-based therapeutics against cancer are highly successful in clinic and currently enjoy unprecedented recognition of their potential; 13 monoclonal antibodies (mAbs) have been approved for clinical use in the European Union and in the United States (one, mylotarg, was withdrawn from market in 2010). Three of the mAbs (bevacizumab, rituximab, trastuzumab) are in the top six selling protein therapeutics with sales in 2010 of more than $5 bln each. Hundreds of mAbs including bispecific mAbs and multispecific fusion proteins, mAbs conjugated with small molecule drugs and mAbs with optimized pharmacokinetics are in clinical trials. However, challenges remain and it appears that deeper understanding of mechanisms is needed to overcome major problems including resistance to therapy, access to targets, complexity of biological systems and individual variations. PMID:22520975

Authorization basis requirements comparison report

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

Brantley, W.M.

The TWRS Authorization Basis (AB) consists of a set of documents identified by TWRS management with the concurrence of DOE-RL. Upon implementation of the TWRS Basis for Interim Operation (BIO) and Technical Safety Requirements (TSRs), the AB list will be revised to include the BIO and TSRs. Some documents that currently form part of the AB will be removed from the list. This SD identifies each - requirement from those documents, and recommends a disposition for each to ensure that necessary requirements are retained when the AB is revised to incorporate the BIO and TSRs. This SD also identifies documentsmore » that will remain part of the AB after the BIO and TSRs are implemented. This document does not change the AB, but provides guidance for the preparation of change documentation.« less

Surface Segregation Energies of BCC Binaries from Ab Initio and Quantum Approximate Calculations

NASA Technical Reports Server (NTRS)

Good, Brian S.

2003-01-01

We compare dilute-limit segregation energies for selected BCC transition metal binaries computed using ab initio and quantum approximate energy method. Ab initio calculations are carried out using the CASTEP plane-wave pseudopotential computer code, while quantum approximate results are computed using the Bozzolo-Ferrante-Smith (BFS) method with the most recent parameterization. Quantum approximate segregation energies are computed with and without atomistic relaxation. The ab initio calculations are performed without relaxation for the most part, but predicted relaxations from quantum approximate calculations are used in selected cases to compute approximate relaxed ab initio segregation energies. Results are discussed within the context of segregation models driven by strain and bond-breaking effects. We compare our results with other quantum approximate and ab initio theoretical work, and available experimental results.

Comparison of DOTA and NODAGA as chelators for (64)Cu-labeled immunoconjugates.

PubMed

Ghosh, Sukhen C; Pinkston, Kenneth L; Robinson, Holly; Harvey, Barrett R; Wilganowski, Nathaniel; Gore, Karen; Sevick-Muraca, Eva M; Azhdarinia, Ali

2015-02-01

Bifunctional chelators have been shown to impact the biodistribution of monoclonal antibody (mAb)-based imaging agents. Recently, radiolabeled 1,4,7-triazacyclononane,1-glutaric acid-4,7-acetic acid (NODAGA)-peptide complexes have demonstrated improved in vivo stability and performance compared to their 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) counterparts. Here, we investigated if similar utility could be achieved with mAbs and compared (64)Cu-labeled DOTA and NODAGA-immunoconjugates for the detection of epithelial cell adhesion molecule (EpCAM) in a prostate cancer model. DOTA and NODAGA-immunoconjugates of an EpCAM targeting mAb (mAb7) were synthesized and radiolabeled with (64)Cu (DOTA: 40°C for 1hr; NODAGA: 25°C for 1hr). The average number of chelators per mAb was quantified by isotopic dilution, and the biological activity of the immunoconjugates was evaluated by flow cytometry and ELISA. Radioligand assays were performed to compare cellular uptake and determine the dissociation constant (Kd) and maximum number of binding sites (Bmax) for the immunoconjugates using DsRed-transfected PC3-cells. A PC3-DsRed xenograft tumor model was established in nude mice and used to perform biodistribution studies to compare organ uptake and pharmacokinetics. (64)Cu-DOTA-mAb7 and (64)Cu-NODAGA-mAb7 were prepared with chelator/protein ratios of 2-3 and obtained in comparable radiochemical yields ranging from 59 to 71%. Similar immunoreactivity was observed with both agents, and mock labeling studies indicated that incubation at room temperature or 40°C did not affect potency. (64)Cu-NODAGA-mAb7 demonstrated higher in vitro cellular uptake while (64)Cu-DOTA-mAb7 had higher Kd and Bmax values. From the biodistribution data, we found similar tumor uptake (13.44±1.21%ID/g and 13.24±4.86%ID/g for (64)Cu-DOTA-mAb7 and (64)Cu-NODAGA-mAb7, respectively) for both agents at 24hr, although normal prostate tissue was significantly lower for (64)Cu-NODAGA-mAb7. (64)Cu-NODAGA-mAb7 also had less accumulation in the liver, suggesting excellent retention of the chelation complex in vivo. This was further confirmed by the higher blood activity of (64)Cu-NODAGA-mAb7, which corresponds to increased bioavailability afforded by the enhanced in vivo stability of the agent. Although tumor/muscle ratios were comparable, tumor/prostate ratios were >2-fold and 1.5-fold higher for (64)Cu-NODAGA-mAb7 at 24 and 48hr, respectively, and suggest better ability to discriminate tumor tissue with (64)Cu-NODAGA-mAb7 in our prostate cancer model. To the best of our knowledge, this study represents the first comparison of (64)Cu-labeled DOTA and NODAGA immunoconjugates in vivo. Our results show favorable in vivo performance for (64)Cu-NODAGA-mAb7 which builds upon previous data on our hybrid mAb7 imaging agent by increasing the detection sensitivity for metastatic prostate tumors, as well as for other types of cancer that express EpCAM. Copyright © 2014 Elsevier Inc. All rights reserved.

Site-Specific Recombination at XerC/D Sites Mediates the Formation and Resolution of Plasmid Co-integrates Carrying a blaOXA-58- and TnaphA6-Resistance Module in Acinetobacter baumannii

PubMed Central

Cameranesi, María M.; Morán-Barrio, Jorgelina; Limansky, Adriana S.; Repizo, Guillermo D.; Viale, Alejandro M.

2018-01-01

Members of the genus Acinetobacter possess distinct plasmid types which provide effective platforms for the acquisition, evolution, and dissemination of antimicrobial resistance structures. Many plasmid-borne resistance structures are bordered by short DNA sequences providing potential recognition sites for the host XerC and XerD site-specific tyrosine recombinases (XerC/D-like sites). However, whether these sites are active in recombination and how they assist the mobilization of associated resistance structures is still poorly understood. Here we characterized the plasmids carried by Acinetobacter baumannii Ab242, a multidrug-resistant clinical strain belonging to the ST104 (Oxford scheme) which produces an OXA-58 carbapenem-hydrolyzing class-D β-lactamase (CHDL). Plasmid sequencing and characterization of replication, stability, and adaptive modules revealed the presence in Ab242 of three novel plasmids lacking self-transferability functions which were designated pAb242_9, pAb242_12, and pAb242_25, respectively. Among them, only pAb242_25 was found to carry an adaptive module encompassing an ISAba825-blaOXA-58 arrangement accompanied by a TnaphA6 transposon, the whole structure conferring simultaneous resistance to carbapenems and aminoglycosides. Ab242 plasmids harbor several XerC/D-like sites, with most sites found in pAb242_25 located in the vicinity or within the adaptive module described above. Electrotransformation of susceptible A. nosocomialis cells with Ab242 plasmids followed by imipenem selection indicated that the transforming plasmid form was a co-integrate resulting from the fusion of pAb242_25 and pAb242_12. Further characterization by cloning and sequencing studies indicated that a XerC/D site in pAb242_25 and another in pAb242_12 provided the active sister pair for the inter-molecular site-specific recombination reaction mediating the fusion of these two plasmids. Moreover, the resulting co-integrate was found also to undergo intra-molecular resolution at the new pair of XerC/D sites generated during fusion thus regenerating the original pAb242_25 and pAb242_12 plasmids. These observations provide the first evidence indicating that XerC/D-like sites in A. baumannii plasmids can provide active pairs for site-specific recombination mediating inter-molecular fusions and intra-molecular resolutions. The overall results shed light on the evolutionary dynamics of A. baumannii plasmids and the underlying mechanisms of dissemination of genetic structures responsible for carbapenem and other antibiotics resistance among the Acinetobacter clinical population. PMID:29434581

Did the Bering Sea Form as a Cenozoic Backarc Basin?

NASA Astrophysics Data System (ADS)

Stern, R. J.; Barth, G. A.; Scheirer, D. S.; Scholl, D. W.

2012-12-01

Understanding the origins of Bering Sea marginal basins (Aleutian, Bowers, and Komandorsky basins; AB, BB, KB) is key for reconstructing N. Pacific tectonic and magmatic evolution. New acquisitions and recompilations of MCS, OBS, and potential field data (Barth et al. poster. this session) for USGS Extended Continental Shelf project and selection of Aleutians as GeoPrisms Subduction Cycles and Deformation focus site stimulate reconsideration of BB, KB, and especially AB origins. AB has long been regarded as N. Pacific crust trapped when the Aleutian subduction began ~45-50 Ma. BB and KB probably formed together as Miocene backarc basins. Presence of Oligo-Miocene arc volcanics on Bowers and Shirshov ridges suggests that these are remnant arcs, orphaned by AB and KB opening. Seven lines of evidence suggest that AB formed as a Paleogene backarc basin: 1) AB heatflow suggests an age of about 44 Ma (Langseth et al 1980 JGR). 2) Formation of NNW-trending rift basins on Bering shelf (Navarin, Pribilof, and St. George basins) in Paleogene time indicate extension at this time. 3) The early Paleogene "red unconformity" of the Beringian margin could indicate uplift, erosion, and subsidence associated with AB opening. 4) ~N-S magnetic anomalies in AB contrasts with E-W Kula anomalies on N. Pacific, indicating that the two tracts of oceanic crust formed at different spreading ridges. 5) Thicker sediment in AB (2-4 km) vs. BB and KB (< 2km) indicates AB is older and is consistent with episodic and short-lived (~20 m.y. duration) opening expected for backarc basins. 6) Aleutian arc magmatic activity began ~50 Ma, about the same time that the Beringian arc shut down. This could also be reconciled by rifting of the Beringian arc to form the AB as backarc basin, accompanied by the displacement of arc magmatic activity to near the present Aleutian arc. 7) Formation of the Aleutian arc as ~3900 km long, nearly perfect small circle is easiest to reconcile with an easily deformed backarc region, as might result from backarc basin opening. If the Aleutian arc formed by rifting of the Beringian margin then it is likely to contain some pre-50 Ma crust. The possibility that AB is a Paleogene backarc basin requires testing via IODP drilling through AB sediments and into oceanic crust.ectonic scenario for formation of Aleutian Arc and Bering Sea basins. Green = present land; yellow = shelf; AB = Aleutian Basin; KB = Komandorsky Basin; BB = Bowers Basin; SR = Shirshov Ridge, BR = Bowers Ridge; Red = active volcanism and spreading ; Blue = extinct volcanism and spreading

Investigations into the development of catalytic activity in anti-acetylcholinesterase idiotypic and anti-idiotypic antibodies.

PubMed

Johnson, Glynis; Moore, Samuel W

2009-01-01

We have previously described anti-acetylcholinesterase antibodies that display acetylcholinesterase-like catalytic activity. No evidence of contaminating enzymes was found, and the antibodies are kinetically and apparently structurally distinct from both acetylcholinesterase (AChE) and butyrylcholinesterase. We have also mimicked the antibody catalytic sites in anti-anti-idiotypic (Ab3) antibodies. Independently from us, similar acetylcholinesterase-like antibodies have been raised as anti-idiotypic (Ab2) antibodies against a non-catalytic anti-acetylcholinesterase antibody, AE-2. In this paper, we describe an epitope analysis, using synthetic peptides in ELISA and competition ELISA, and a peptide array, of five catalytic anti-acetylcholinesterase antibodies (Ab1s), three catalytic Ab3s, as well as antibody AE-2 and a non-catalytic Ab2. The catalytic Ab1s and Ab3s recognized three Pro- and Gly-containing sequences ((40)PPMGPRRFL, (78)PGFEGTE, and (258)PPGGTGGNDTELVAC) on the AChE surface. As these sequences do not adjoin in the AChE structure, recognition would appear to be due to cross-reaction. This was confirmed by the observation that the sequences superimpose structurally. The non-catalytic antibodies, AE-2 and the Ab2, recognized AChE's peripheral anionic site (PAS), in particular, the sequence (70)YQYVD, which contains two of the site's residues. The crystal structure of the AChE tetramer (Bourne et al., 1999) shows direct interaction and high complementarity between the (257)CPPGGTGGNDTELVAC sequence and the PAS. Antibodies recognizing the sequence and the PAS may, in turn, be complementary; this may account for the apparent paradox of catalytic development in both Ab1s and Ab2s. The PAS binds, but does not hydrolyze, substrate. The catalytic Ab1s, therefore, recognize a site that may function as a substrate analog, and this, together with the presence of an Arg-Glu salt bridge in the epitope, suggests mechanisms whereby catalytic activity may have developed. In conclusion, the development of AChE-like catalytic activity in anti-AChE Ab1s and Ab2s appears to be the result of a combination of structural complementarity to a substrate-binding site, charge complementarity to a salt bridge, and specific structural peculiarities of the AChE molecule. Copyright 2008 John Wiley & Sons, Ltd.

Adhesion between liposomes mediated by the chlorophyll a/b light- harvesting complex isolated from chloroplast membranes

PubMed Central

1980-01-01

A highly purified chlorophyll a/b light-harvesting complex (chl a/b LHC; chl a/b ratio 1.2) was obtained from Triton-solubilized chloroplast membranes of pea and barley according to the method of Burke et al. (1978, Arch. Biochem. Biophys. 187: 252--263). Gel electrophoresis of the cation-precipitated chl a/b LHC from peas reveals the presence of four polypeptides in the 23- to 28-kdalton size range. Three of these peptides appear to be identical to those derived from re-electrophoresed CPII and CPII* bands. In freeze-fracture replicas, the cation-precipitated chl a/b LHC appears as a semicrystalline aggregate of membranous sheets containing closely spaced granules. Upon removal of the cations by dialysis, the aggregates break up into their constituent membranous sheets without changing their granular substructure. These membranous sheets can be resolubilized in 1.5% Triton X-100, and the chl a/b LHC particles then reconstituted into soybean lecithin liposomes. Freeze-fracture micrographs of the reconstituted chl a/b LHC vesicles suspended in a low salt medium reveal randomly dispersed approximately 80-A particles on both concave and convex fracture faces as well as some crystalline particle arrays, presumably resulting from incompletely solubilized fragments of the membranous sheets. Based on the approximately 80-A diameter of the particles, and on the assumption that one freeze- fracture particle represents the structural unit of one chl a/b LHC aggregate, a theoretical mol wt of approximately 200 kdalton has been calculated for the chl a/b LHC. Deep-etching and negative-staining techniques reveal that the chl a/b LHC particles are also exposed on the surface of the bilayer membranes. Addition of greater than or equal to 2 mM MgCl2 or greater than or equal to 60 mM NaCl to the reconstituted vesicles leads to their aggregation and, with divalent cations, to the formation of extensive membrane stacks. At the same time, the chl a/b LHC particles become clustered into the adhering membrane regions. Under these conditions the particles in adjacent membranes usually become precisely aligned. Evidence is presented to aupport the hypothesis that adhesion between the chl a/b LHC particles is mediated by hydrophobic interactions, and that the cations are needed to neutralize surface charges on the particles. PMID:7350170

40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

Code of Federal Regulations, 2010 CFR

2010-07-01

... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as plant...

40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 40 Protection of Environment 24 2011-07-01 2011-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as plant...

40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 40 Protection of Environment 25 2012-07-01 2012-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as plant...

40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 40 Protection of Environment 25 2013-07-01 2013-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as plant...

40 CFR 174.511 - Bacillus thuringiensis Cry1Ab protein in all plants; exemption from the requirement of a tolerance.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 40 Protection of Environment 24 2014-07-01 2014-07-01 false Bacillus thuringiensis Cry1Ab protein... Cry1Ab protein in all plants; exemption from the requirement of a tolerance. Residues of Bacillus thuringiensis Cry1Ab protein in all plants are exempt from the requirement of a tolerance when used as plant...

Ab Initio-Based Predictions of Hydrocarbon Combustion Chemistry

DTIC Science & Technology

2015-07-15

There are two prime objectives of the research. One is to develop and apply efficient methods for using ab initio potential energy surfaces (PESs...31-Mar-2015 Approved for Public Release; Distribution Unlimited Final Report: Ab Initio -Based Predictions of Hydrocarbon Combustion Chemistry The...Office P.O. Box 12211 Research Triangle Park, NC 27709-2211 hydrocarbon combustion, ab initio quantum chemistry, potential energy surfaces, chemical

Molecular evidence for the existence of lipopolysaccharide-induced TNF-alpha factor (LITAF) and Rel/NF-kB pathways in disk abalone (Haliotis discus discus).

PubMed

De Zoysa, Mahanama; Nikapitiya, Chamilani; Oh, Chulhong; Whang, Ilson; Lee, Jae-Seong; Jung, Sung-Ju; Choi, Cheol Young; Lee, Jehee

2010-01-01

The lipopolysaccharide-induced TNF-alpha factor (LITAF) and Rel family nuclear factor kappaB (Rel/NF-kB) are two important transcription factors which play major roles in the regulating inflammatory cytokine, apoptosis and immune related genes. Here, we report the discovery of disk abalone LITAF (AbLITAF) and Rel/NF-kB (AbRel/NF-kB) homologues and their immune responses. Full-length cDNA of AbLITAF consists of 441 bp open reading frame (ORF) that translates into putative peptide of 147 aa. Analysis of AbLITAF sequence showed it has characteristic LITAF (Zn(+2)) binding domain with two CXXC motifs. Phylogenetic analysis results further revealed that AbLITAF is a member of LITAF family. AbRel/NF-kB is 584 aa protein that contains several characteristic motifs including Rel homology domain (RHD), Rel protein signature, DNA binding motif, nuclear localization signal (NLS) and transcription factor immunoglobulin - like fold (TIG) similar to their invertebrate and vertebrate counterparts. Tissue specific analysis results showed that both AbLITAF and AbRel/NF-kB mRNA was expressed ubiquitously in all selected tissues in constitutive manner. However, constitutive expression of AbLITAF was higher than AbRel/NF-kB in all tissues except mantle. Upon immune challenge by bacteria (Vibrio alginolyticus, Vibrio parahemolyticus and Lysteria monocytogenes) and viral hemoragic septicemia virus (VHSV), AbLITAF showed the significant up-regulation in gills while AbRel/NF-kB transcription was not change significantly. Based on transcriptional response against immune challenge, we could suggest that regulation of TNF-alpha expression may have occurred mainly by LITAF activation rather than NF-kB in disk abalone. The cumulative data from other molluscs and our data with reference to TNF-alpha, LITAF and Rel/NF-kB from disk abalone provide strong evidence that LITAF and NF-kB are independent pathways likely to occur throughout the Phylum mollusca. 2010 Elsevier Ltd. All rights reserved.

Monoclonal antibodies to meningococcal factor H binding protein with overlapping epitopes and discordant functional activity.

PubMed

Giuntini, Serena; Beernink, Peter T; Reason, Donald C; Granoff, Dan M

2012-01-01

Meningococcal factor H binding protein (fHbp) is a promising vaccine candidate. Anti-fHbp antibodies can bind to meningococci and elicit complement-mediated bactericidal activity directly. The antibodies also can block binding of the human complement down-regulator, factor H (fH). Without bound fH, the organism would be expected to have increased susceptibility to bacteriolysis. Here we describe bactericidal activity of two anti-fHbp mAbs with overlapping epitopes in relation to their different effects on fH binding and bactericidal activity. Both mAbs recognized prevalent fHbp sequence variants in variant group 1. Using yeast display and site-specific mutagenesis, binding of one of the mAbs (JAR 1, IgG3) to fHbp was eliminated by a single amino acid substitution, R204A, and was decreased by K143A but not by R204H or D142A. The JAR 1 epitope overlapped that of previously described mAb (mAb502, IgG2a) whose binding to fHbp was eliminated by R204A or R204H substitutions, and was decreased by D142A but not by K143A. Although JAR 1 and mAb502 appeared to have overlapping epitopes, only JAR 1 inhibited binding of fH to fHbp and had human complement-mediated bactericidal activity. mAb502 enhanced fH binding and lacked human complement-mediated bactericidal activity. To control for confounding effects of different mouse IgG subclasses on complement activation, we created chimeric mAbs in which the mouse mAb502 or JAR 1 paratopes were paired with human IgG1 constant regions. While both chimeric mAbs showed similar binding to fHbp, only JAR 1, which inhibited fH binding, had human complement-mediated bactericidal activity. The lack of human complement-mediated bactericidal activity by anti-fHbp mAb502 appeared to result from an inability to inhibit binding of fH. These results underscore the importance of inhibition of fH binding for anti-fHbp mAb bactericidal activity.

CD28 homodimer interface mimetic peptide acts as a preventive and therapeutic agent in models of severe bacterial sepsis and gram-negative bacterial peritonitis.

PubMed

Ramachandran, Girish; Kaempfer, Raymond; Chung, Chun-Shiang; Shirvan, Anat; Chahin, Abdullah B; Palardy, John E; Parejo, Nicolas A; Chen, Yaping; Whitford, Melissa; Arad, Gila; Hillman, Dalia; Shemesh, Ronen; Blackwelder, William; Ayala, Alfred; Cross, Alan S; Opal, Steven M

2015-03-15

Severe gram-negative bacterial infections and sepsis are major causes of morbidity and mortality. Dysregulated, excessive proinflammatory cytokine expression contributes to the pathogenesis of sepsis. A CD28 mimetic peptide (AB103; previously known as p2TA) that attenuates CD28 signaling and T-helper type 1 cytokine responses was tested for its ability to increase survival in models of polymicrobial infection and gram-negative sepsis. Mice received AB103, followed by an injection of Escherichia coli 0111:B4 lipopolysaccharide (LPS); underwent induction E. coli 018:K1 peritonitis induction, followed by treatment with AB103; or underwent cecal ligation and puncture (CLP), followed by treatment with AB103. The effects of AB103 on factors associated with and the lethality of challenge infections were analyzed. AB103 strongly attenuated induction of tumor necrosis factor α and interleukin 6 (IL-6) by LPS in human peripheral blood mononuclear cells. Receipt of AB103 following intraperitoneal injection of LPS resulted in survival among 73% of CD1 mice (11 of 15), compared with 20% of controls (3 of 15). Suboptimal doses of antibiotic alone protected 20% of mice (1 of 5) from E. coli peritonitis, whereas 100% (15 of 15) survived when AB103 was added 4 hours following infection. Survival among mice treated with AB103 12 hours after CLP was 100% (8 of 8), compared with 17% among untreated mice (1 of 6). In addition, receipt of AB103 12 hours after CLP attenuated inflammatory cytokine responses and neutrophil influx into tissues and promoted bacterial clearance. Receipt of AB103 24 hours after CLP still protected 63% of mice (5 of 8). Single-dose AB103 reduces mortality in experimental models of polymicrobial and gram-negative bacterial infection and sepsis, warranting further studies of this agent in clinical trials. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Alisol B 23-acetate protects against ANIT-induced hepatotoxity and cholestasis, due to FXR-mediated regulation of transporters and enzymes involved in bile acid homeostasis

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

Meng, Qiang; Chen, Xin-li; Wang, Chang-yuan

2015-03-15

Intrahepatic cholestasis is a clinical syndrome with systemic and intrahepatic accumulation of excessive toxic bile acids that ultimately cause hepatobiliary injury. Appropriate regulation of bile acids in hepatocytes is critically important for protection against liver injury. In the present study, we characterized the protective effect of alisol B 23-acetate (AB23A), a natural triterpenoid, on alpha-naphthylisothiocyanate (ANIT)-induced liver injury and intrahepatic cholestasis in mice and further elucidated the mechanisms in vivo and in vitro. AB23A treatment dose-dependently protected against liver injury induced by ANIT through reducing hepatic uptake and increasing efflux of bile acid via down-regulation of hepatic uptake transporters (Ntcp)more » and up-regulation of efflux transporter (Bsep, Mrp2 and Mdr2) expression. Furthermore, AB23A reduced bile acid synthesis through repressing Cyp7a1 and Cyp8b1, increased bile acid conjugation through inducing Bal, Baat and bile acid metabolism through an induction in gene expression of Sult2a1. We further demonstrate the involvement of farnesoid X receptor (FXR) in the hepatoprotective effect of AB23A. The changes in transporters and enzymes, as well as ameliorative liver histology in AB23A-treated mice were abrogated by FXR antagonist guggulsterone in vivo. In vitro evidences also directly demonstrated the effect of AB23A on FXR activation in a dose-dependent manner using luciferase reporter assay in HepG2 cells. In conclusion, AB23A produces protective effect against ANIT-induced hepatotoxity and cholestasis, due to FXR-mediated regulation of transporters and enzymes. - Highlights: • AB23A has at least three roles in protection against ANIT-induced liver injury. • AB23A decreases Ntcp, and increases Bsep, Mrp2 and Mdr2 expression. • AB23A represses Cyp7a1 and Cyp8b1 through inducing Shp and Fgf15 expression. • AB23A increases bile acid metabolism through inducing Sult2a1 expression. • FXR activation is involved in the hepatoprotective effect of AB23A.« less

Predominant antitumor effects by fully human anti-TRAIL-receptor2 (DR5) monoclonal antibodies in human glioma cells in vitro and in vivo

PubMed Central

Nagane, Motoo; Shimizu, Saki; Mori, Eiji; Kataoka, Shiro; Shiokawa, Yoshiaki

2010-01-01

Tumor necrosis factor–related apoptosis-inducing ligand (TRAIL/Apo2 L) preferentially induces apoptosis in human tumor cells through its cognate death receptors DR4 or DR5, thereby being investigated as a potential agent for cancer therapy. Here, we applied fully human anti-human TRAIL receptor monoclonal antibodies (mAbs) to specifically target one of death receptors for TRAIL in human glioma cells, which could also reduce potential TRAIL-induced toxicity in humans. Twelve human glioma cell lines treated with several fully human anti-human TRAIL receptor mAbs were sensitive to only anti-DR5 mAbs, whereas they were totally insensitive to anti-DR4 mAb. Treatment with anti-DR5 mAbs exerted rapid cytotoxicity and lead to apoptosis induction. The cellular sensitivity was closely associated with cell-surface expression of DR5. Expression of c-FLIPL, Akt, and Cyclin D1 significantly correlated with sensitivity to anti-DR5 mAbs. Primary cultures of glioma cells were also relatively resistant to anti-DR5 mAbs, exhibiting both lower DR5 and higher c-FLIPL expression. Downregulation of c-FLIPL expression resulted in the sensitization of human glioma cells to anti-DR5 mAbs, whereas overexpression of c-FLIPL conferred resistance to anti-DR5 mAb. Treatment of tumor-burden nude mice with the direct agonist anti-DR5 mAb KMTR2 significantly suppressed growth of subcutaneous glioma xenografts leading to complete regression. Similarly, treatment of nude mice bearing intracerebral glioma xenografts with KMTR2 significantly elongated lifespan without tumor recurrence. These results suggest that DR5 is the predominant TRAIL receptor mediating apoptotic signals in human glioma cells, and sensitivity to anti-DR5 mAbs was determined at least in part by the expression level of c-FLIPL and Akt. Specific targeting of death receptor pathway through DR5 using fully human mAbs might provide a novel therapeutic strategy for intractable malignant gliomas. PMID:20511188

Adsorption behavior of a human monoclonal antibody at hydrophilic and hydrophobic surfaces

PubMed Central

Couston, Ruairidh G.; Skoda, Maximilian W.; Uddin, Shahid; van der Walle, Christopher F.

2013-01-01

One aspiration for the formulation of human monoclonal antibodies (mAb) is to reach high solution concentrations without compromising stability. Protein surface activity leading to instability is well known, but our understanding of mAb adsorption to the solid-liquid interface in relevant pH and surfactant conditions is incomplete. To investigate these conditions, we used total internal reflection fluorescence (TIRF) and neutron reflectometry (NR). The mAb tested (“mAb-1”) showed highest surface loading to silica at pH 7.4 (~12 mg/m2), with lower surface loading at pH 5.5 (~5.5 mg/m2, further from its pI of 8.99) and to hydrophobized silica (~2 mg/m2). The extent of desorption of mAb-1 from silica or hydrophobized silica was related to the relative affinity of polysorbate 20 or 80 for the same surface. mAb-1 adsorbed to silica on co-injection with polysorbate (above its critical micelle concentration) and also to silica pre-coated with polysorbate. A bilayer model was developed from NR data for mAb-1 at concentrations of 50–5000 mg/L, pH 5.5, and 50–2000 mg/L, pH 7.4. The inner mAb-1 layer was adsorbed to the SiO2 surface at near saturation with an end-on” orientation, while the outer mAb-1 layer was sparse and molecules had a “side-on” orientation. A non-uniform triple layer was observed at 5000 mg/L, pH 7.4, suggesting mAb-1 adsorbed to the SiO2 surface as oligomers at this concentration and pH. mAb-1 adsorbed as a sparse monolayer to hydrophobized silica, with a layer thickness increasing with bulk concentration - suggesting a near end-on orientation without observable relaxation-unfolding. PMID:23196810

Two duplicated chicken-type lysozyme genes in disc abalone Haliotis discus discus: molecular aspects in relevance to structure, genomic organization, mRNA expression and bacteriolytic function.

PubMed

Umasuthan, Navaneethaiyer; Bathige, S D N K; Kasthuri, Saranya Revathy; Wan, Qiang; Whang, Ilson; Lee, Jehee

2013-08-01

Lysozymes are crucial antibacterial proteins that are associated with catalytic cleavage of peptidoglycan and subsequent bacteriolysis. The present study describes the identification of two lysozyme genes from disc abalone Haliotis discus discus and their characterization at sequence-, genomic-, transcriptional- and functional-levels. Two cDNAs and BAC clones bearing lysozyme genes were isolated from abalone transcriptome and BAC genomic libraries, respectively and sequences were determined. Corresponding deduced amino acid sequences harbored a chicken-type lysozyme (LysC) family profile and exhibited conserved characteristics of LysC family members including active residues (Glu and Asp) and GS(S/T)DYGIFQINS motif suggested that they are LysC counterparts in disc abalone and designated as abLysC1 and abLysC2. While abLysC1 represented the homolog recently reported in Ezo abalone [1], abLysC2 shared significant identity with LysC homologs. Unlike other vertebrate LysCs, coding sequence of abLysCs were distributed within five exons interrupted by four introns. Both abLysCs revealed a broader mRNA distribution with highest levels in mantle (abLysC1) and hepatopancreas (abLysC2) suggesting their likely main role in defense and digestion, respectively. Investigation of temporal transcriptional profiles post-LPS and -pathogen challenges revealed induced-responses of abLysCs in gills and hemocytes. The in vitro muramidase activity of purified recombinant (r) abLysCs proteins was evaluated, and findings indicated that they are active in acidic pH range (3.5-6.5) and over a broad temperature range (20-60 °C) and influenced by ionic strength. When the antibacterial spectra of (r)abLysCs were examined, they displayed differential activities against both Gram positive and Gram negative strains providing evidence for their involvement in bacteriolytic function in abalone physiology. Copyright © 2013 Elsevier Ltd. All rights reserved.

IB-12LUNG MASS AND IMMUNOSUPPRESSANT RESPONSIVE SEIZURES: VGKC AUTOIMMUNITY MASQUERADING AS ETIOLOGY OR ACTING AS REACTIVE MARKER?

PubMed Central

Umemura, Yoshie; Bujarski, Krzysztof; Ronan, Lara

2014-01-01

Voltage gated potassium channel complex antibody (VGKC Ab) has been associated with many neurological illnesses including seizures. VGKC Ab related seizures are less responsive to antiepileptic drugs alone, and often require immunosuppression to achieve seizure freedom. Recently, specific antigenic targets such as LGI1 and CASPR2 within VGKC have been found and have been associated with specific syndromes. It has also been noted that the term "VGKC Ab" itself is problematic because it groups LGI1 and CASPR2 mediated disorders with other illnesses with unknown antigenic specificity. We describe a case of faciobrachial dystonia seizures refractory to multiple antiepileptic agents. Video EEG monitoring revealed mesial frontal epileptic discharges and serum test was positive for VGKC Ab. Patient's seizures were controlled after intravenous corticosteroid then oral mycophenolate maintenance, however, seizures returned with immunotherapy taper and cessation. When the seizures recurred, serum VGKC Ab, CSF CASPR 2 and LGI1 Abs were negative. Interestingly, she was subsequently found to have a lung mass consistent with sarcoidosis. Seizure control was achieved again with restarting of mycophenolate. Most VGKC Ab related illnesses are autoimmune even though it is a part of readily available commercial paraneoplastic testing panels. There has been no previously reported VGKC Ab related seizures associated with sarcoidosis. Although sarcoidosis is also an autoimmune entity, it is of T-cell mediated condition. It has been proposed that VGKC Ab may be a part of normal autoantibodies found in otherwise healthy individuals, which can rise to detectable levels under some circumstances. This patient suffered recurrent seizures responsive to immunotherapy even though VGKC, LGI1, and CASPR2 Abs were negative, suggesting another underlying autoimmune pathogenesis and VGKC Ab may have been a mere reactive marker of such underlying process. This raises the question of clinical significance of these antibodies and whether they are causative or reactive of the pathology.

PirAB protein from Xenorhabdus nematophila HB310 exhibits a binary toxin with insecticidal activity and cytotoxicity in Galleria mellonella.

PubMed

Yang, Qing; Zhang, Jie; Li, Tianhui; Liu, Shen; Song, Ping; Nangong, Ziyan; Wang, Qinying

2017-09-01

PirAB (Photorhabdus insect-related proteins, PirAB) toxin was initially found in the Photorhabdus luminescens TT01 strain and has been shown to be a binary toxin with high insecticidal activity. Based on GenBank data, this gene was also found in the Xenorhabdus nematophila genome sequence. The predicted amino acid sequence of pirA and pirB in the genome of X. nematophila showed 51% and 50% identity with those gene sequences from P. luminescens. The purpose of this experiment is to identify the relevant information for this toxin gene in X. nematophila. The pirA, pirB and pirAB genes of X. nematophila HB310 were cloned and expressed in Escherichia coli BL21 (DE3) using the pET-28a vector. A PirAB-fusion protein (PirAB-F) was constructed by linking the pirA and pirB genes with the flexible linker (Gly) 4 DNA encoding sequence and then efficiently expressed in E. coli. The hemocoel and oral insecticidal activities of the recombinant proteins were analyzed against the larvae of Galleria mellonella. The results show that PirA/B alone, PirA/B mixture, co-expressed PirAB protein, and PirAB-F all had no oral insecticidal activity against the second-instar larvae of G. mellonella. Only PirA/B mixture and co-expressed PirAB protein had hemocoel insecticidal activity against G. mellonella fifth-instar larvae, with an LD 50 of 2.718μg/larva or 1.566μg/larva, respectively. Therefore, we confirmed that PirAB protein of X. nematophila HB310 is a binary insecticidal toxin. The successful expression and purification of PirAB laid a foundation for further studies on the function, insecticidal mechanism and expression regulation of the binary toxin. Copyright © 2017 Elsevier Inc. All rights reserved.

Anti-TIF1-γ antibody and cancer-associated myositis: A clinicohistopathologic study.

PubMed

Hida, Ayumi; Yamashita, Takenari; Hosono, Yuji; Inoue, Manami; Kaida, Kenichi; Kadoya, Masato; Miwa, Yusuke; Yajima, Nobuyuki; Maezawa, Reika; Arai, Satoko; Kurasawa, Kazuhiro; Ito, Kazuhiro; Shimada, Hiroyuki; Iwanami, Tomoko; Sonoo, Masahiro; Hatanaka, Yuki; Murayama, Shigeo; Uchibori, Ayumi; Chiba, Atsuro; Aizawa, Hitoshi; Momoo, Takayuki; Nakae, Yoshiharu; Sakurai, Yasuhisa; Shiio, Yasushi; Hashida, Hideji; Yoshizawa, Toshihiro; Sakiyama, Yoshio; Oda, Aya; Inoue, Kiyoharu; Takeuchi, Sousuke; Iwata, Nobue K; Date, Hidetoshi; Masuda, Naoki; Mikata, Takashi; Motoyoshi, Yasufumi; Uesaka, Yoshikazu; Maeda, Meiko Hashimoto; Nakashima, Ran; Tsuji, Shoji; Kwak, Shin; Mimori, Tsuneyo; Shimizu, Jun

2016-07-19

We aimed to analyze the clinical and histopathologic features of cancer-associated myositis (CAM) in relation to anti-transcriptional intermediary factor 1 γ antibody (anti-TIF1-γ-Ab), a marker of cancer association. We retrospectively studied 349 patients with idiopathic inflammatory myopathies (IIMs), including 284 patients with pretreatment biopsy samples available. For the classification of IIMs, the European Neuromuscular Center criteria were applied. Patients with CAM with (anti-TIF1-γ-Ab[+] CAM) and without anti-TIF1-γ-Ab (anti-TIF1-γ-Ab[-] CAM) were compared with patients with IIM without cancers within and beyond 3 years of myositis diagnosis. Cancer was detected in 75 patients, of whom 36 (48%) were positive for anti-TIF1-γ-Ab. In anti-TIF1-γ-Ab(+) patients with CAM, cancers were detected within 1 year of myositis diagnosis in 35 (97%) and before 1 year of myositis diagnosis in 1. All the anti-TIF1-γ-Ab(+) patients with CAM satisfied the dermatomyositis (DM) criteria, including 2 possible DM sine dermatitis cases, and were characterized histologically by the presence of perifascicular atrophy, vacuolated fibers (VFs), and dense C5b-9 deposits on capillaries (dC5b-9). In contrast, 39 anti-TIF1-γ-Ab(-) patients with CAM were classified into various subgroups, and characterized by a higher frequency of necrotizing autoimmune myopathy (NAM). Notably, all 7 patients with CAM classified into the NAM subgroup were anti-TIF1-γ-Ab(-) and exhibited no dC5b-9 or VFs. CAM includes clinicohistopathologically heterogeneous disease entities. Among CAM entities, anti-TIF1-γ-Ab(+) CAM has characteristically shown a close temporal association with cancer detection and the histopathologic findings of dC5b-9 and VFs, and CAM with NAM is a subset of anti-TIF1-γ-Ab(-) CAM. © 2016 American Academy of Neurology.

Effect of hepatitis B vaccination in hepatitis B surface antibody-negative pregnant mothers on the vertical transmission of hepatitis B virus from father to infant.

PubMed

Cao, Li-Hua; Li, Yun-Ru; Wang, Shou-Yun; Liu, Zhi-Min; Sun, Shao-Chun; Xu, Dong-Bo; Zhang, Ji-Dong

2015-07-01

The aim of the present study was to investigate the effects of vaccination with the hepatitis B vaccine (HBVac) in HB surface antibody (HBsAb)-negative pregnant mothers on the vertical transmission of HB virus (HBV) from father to infant. All the fathers tested positive for the serum HBV DNA and HB surface antigen (HBsAg) markers. The pregnant females were divided into an observation group or a control group depending on whether their serum was HBsAb-negative or positive. A total of 93 healthy individuals without HBV infection were included in a blank group, while 96 females who were serum HBV marker-negative or HB core antibody (HBcAb)-positive/(HBsAb)-negative were included in the observation group. The control group comprised 89 females who all tested positive for serum HBsAb, HB envelope antibodies and HBcAb. In the observation group, the positive rate of HBV DNA in the newborns was 7.29% (7/96), the positive rate of HBsAg was 3.13% (3/96) and the positive rate of HBsAb was 81.3% (78/96). In the control group, the positive rates of HBV DNA, HBsAg and HBsAb in the newborns were 4.49% (4/89), 2.25% (2/89) and 89.9% (80/89), respectively. No statistically significant differences were observed between the two groups. Therefore, the results of the present study indicate that HBVac treatment for HBsAb-negative pregnant females may have a positive role in blocking the vertical transmission of HBV from father to infant, as long as the vaccination is able to induce the production of a sufficient quantity of HBsAb. The HBVac exhibited no difference compared with pre-pregnancy HBsAb in blocking the vertical transmission of HBV from father to infant.

Therapeutic Effects of Monoclonal Antibody against Dengue Virus NS1 in a STAT1 Knockout Mouse Model of Dengue Infection.

PubMed

Wan, Shu-Wen; Chen, Pei-Wei; Chen, Chin-Yu; Lai, Yen-Chung; Chu, Ya-Ting; Hung, Chia-Yi; Lee, Han; Wu, Hsuan Franziska; Chuang, Yung-Chun; Lin, Jessica; Chang, Chih-Peng; Wang, Shuying; Liu, Ching-Chuan; Ho, Tzong-Shiann; Lin, Chiou-Feng; Lee, Chien-Kuo; Wu-Hsieh, Betty A; Anderson, Robert; Yeh, Trai-Ming; Lin, Yee-Shin

2017-10-15

Dengue virus (DENV) is the causative agent of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome and is endemic to tropical and subtropical regions of the world. Our previous studies showed the existence of epitopes in the C-terminal region of DENV nonstructural protein 1 (NS1) which are cross-reactive with host Ags and trigger anti-DENV NS1 Ab-mediated endothelial cell damage and platelet dysfunction. To circumvent these potentially harmful events, we replaced the C-terminal region of DENV NS1 with the corresponding region from Japanese encephalitis virus NS1 to create chimeric DJ NS1 protein. Passive immunization of DENV-infected mice with polyclonal anti-DJ NS1 Abs reduced viral Ag expression at skin inoculation sites and shortened DENV-induced prolonged bleeding time. We also investigated the therapeutic effects of anti-NS1 mAb. One mAb designated 2E8 does not recognize the C-terminal region of DENV NS1 in which host-cross-reactive epitopes reside. Moreover, mAb 2E8 recognizes NS1 of all four DENV serotypes. We also found that mAb 2E8 caused complement-mediated lysis in DENV-infected cells. In mouse model studies, treatment with mAb 2E8 shortened DENV-induced prolonged bleeding time and reduced viral Ag expression in the skin. Importantly, mAb 2E8 provided therapeutic effects against all four serotypes of DENV. We further found that mAb administration to mice as late as 1 d prior to severe bleeding still reduced prolonged bleeding time and hemorrhage. Therefore, administration with a single dose of mAb 2E8 can protect mice against DENV infection and pathological effects, suggesting that NS1-specific mAb may be a therapeutic option against dengue disease. Copyright © 2017 by The American Association of Immunologists, Inc.

[Dynamics of Cry1ab protein content in the rhizosphere soil and straw debris of transgenic Bt corn].

PubMed

Li, Fan; Wang, Min; Sun, Hong-Wei; Yang, Shu-Ke; Lu, Xing-Bo

2013-07-01

By using ELISA test kits, a field investigation was conducted on the degradation dynamics of CrylAb protein in the rhizosphere soil of Bt corn MON810 at its different growth stages and in the MON810 straws returned into field after harvest. Three models (shift-log model, exponential model, and bi-exponential model) were used to fit the degradation dynamics of the Cry1 Ab protein from the straw debris, and the DT50 and DT90, values were estimated. There existed great differences in the CrylAb protein content in the rhizosphere soil of MON810 at its different growth stages, but overall, the CrylAb protein content was decreased remarkably with the growth of MON810. The degradation of Cry1 Ab protein from the straws covered on soil surface and buried in soil showed the same two-stage pattern, i.e., more rapid at early stage and slow-stable in later period. Within the first week after straw return, the degradation rate of the CrylAb protein from the straws covered on soil surface was significantly higher than that from the straws buried in soil. At 10 d, the degradation rate of the CrylAb protein from the straws covered on soil surface and buried in soil was basically the same, being 88.8% and 88.6%, respectively. After 20 days, the degradation of CrylAb protein entered slow-stable stage. Till at 180 d, a small amount of Cry1Ab protein could still be detected in the straw debris. All of the three models used in this study could fit the decay pattern of the CrylAb protein from the straw debris in field. By comparing the correlation coefficient (r) and the consistency between the measured and calculated DT90, bi-exponential model was considered to be the best.

Effectiveness of motorcycle antilock braking systems (ABS) in reducing crashes, the first cross-national study.

PubMed

Rizzi, Matteo; Strandroth, Johan; Kullgren, Anders; Tingvall, Claes; Fildes, Brian

2015-01-01

This study set out to evaluate the effectiveness of motorcycle antilock braking systems (ABS) in reducing real-life crashes. Since the European Parliament has voted on legislation making ABS mandatory on all new motorcycles over 125 cc from 2016, the fitment rate in Europe is likely to increase in the coming years. Though previous research has focused on mostly large displacement motorcycles, this study used police reports from Spain (2006-2009), Italy (2009), and Sweden (2003-2012) in order to analyze a wide range of motorcycles, including scooters, and compare countries with different motorcycling habits. The statistical analysis used odds ratio calculations with an induced exposure approach. Previous research found that head-on crashes were the least ABS-affected crash type and was therefore used as the nonsensitive crash type for ABS in these calculations. The same motorcycle models, with and without ABS, were compared and the calculations were carried out for each country separately. Crashes involving only scooters were further analyzed. The effectiveness of motorcycle ABS in reducing injury crashes ranged from 24% (95% confidence interval [CI], 12-36) in Italy to 29% (95% CI, 20-38) in Spain, and 34% (95% CI, 16-52) in Sweden. The reductions in severe and fatal crashes were even greater, at 34% (95% CI, 24-44) in Spain and 42% (95% CI, 23-61) in Sweden. The overall reductions of crashes involving ABS-equipped scooters (at least 250 cc) were 27% (95% CI, 12-42) in Italy and 22% (95% CI, 2-42) in Spain. ABS on scooters with at least a 250 cc engine reduced severe and fatal crashes by 31% (95% CI, 12-50), based on Spanish data alone. At this stage, there is more than sufficient scientific-based evidence to support the implementation of ABS on all motorcycles, even light ones. Further research should aim at understanding the injury mitigating effects of motorcycle ABS, possibly in combination with combined braking systems.

Complete Genomic Structure of the Cultivated Rice Endophyte Azospirillum sp. B510

PubMed Central

Kaneko, Takakazu; Minamisawa, Kiwamu; Isawa, Tsuyoshi; Nakatsukasa, Hiroki; Mitsui, Hisayuki; Kawaharada, Yasuyuki; Nakamura, Yasukazu; Watanabe, Akiko; Kawashima, Kumiko; Ono, Akiko; Shimizu, Yoshimi; Takahashi, Chika; Minami, Chiharu; Fujishiro, Tsunakazu; Kohara, Mitsuyo; Katoh, Midori; Nakazaki, Naomi; Nakayama, Shinobu; Yamada, Manabu; Tabata, Satoshi; Sato, Shusei

2010-01-01

We determined the nucleotide sequence of the entire genome of a diazotrophic endophyte, Azospirillum sp. B510. Strain B510 is an endophytic bacterium isolated from stems of rice plants (Oryza sativa cv. Nipponbare). The genome of B510 consisted of a single chromosome (3 311 395 bp) and six plasmids, designated as pAB510a (1 455 109 bp), pAB510b (723 779 bp), pAB510c (681 723 bp), pAB510d (628 837 bp), pAB510e (537 299 bp), and pAB510f (261 596 bp). The chromosome bears 2893 potential protein-encoding genes, two sets of rRNA gene clusters (rrns), and 45 tRNA genes representing 37 tRNA species. The genomes of the six plasmids contained a total of 3416 protein-encoding genes, seven sets of rrns, and 34 tRNAs representing 19 tRNA species. Eight genes for plasmid-specific tRNA species are located on either pAB510a or pAB510d. Two out of eight genomic islands are inserted in the plasmids, pAB510b and pAB510e, and one of the islands is inserted into trnfM-CAU in the rrn located on pAB510e. Genes other than the nif gene cluster that are involved in N2 fixation and are homologues of Bradyrhizobium japonicum USDA110 include fixABCX, fixNOQP, fixHIS, fixG, and fixLJK. Three putative plant hormone-related genes encoding tryptophan 2-monooxytenase (iaaM) and indole-3-acetaldehyde hydrolase (iaaH), which are involved in IAA biosynthesis, and ACC deaminase (acdS), which reduces ethylene levels, were identified. Multiple gene-clusters for tripartite ATP-independent periplasmic-transport systems and a diverse set of malic enzymes were identified, suggesting that B510 utilizes C4-dicarboxylate during its symbiotic relationship with the host plant. PMID:20047946

Establishment of a sandwich enzyme-linked immunosorbent assay for specific detection of Bacillus thuringiensis (Bt) Cry1Ab toxin utilizing a monoclonal antibody produced with a novel hapten designed with molecular model.

PubMed

Dong, Sa; Zhang, Xiao; Liu, Yuan; Zhang, Cunzheng; Xie, Yajing; Zhong, Jianfeng; Xu, Chongxin; Liu, Xianjin

2017-03-01

Cry1Ab toxin is commonly expressed in genetically modified crops in order to control chewing pests. At present, the detection method with enzyme-linked immunosorbent assay (ELISA) based on monoclonal antibody cannot specifically detect Cry1Ab toxin for Cry1Ab's amino acid sequence and spatial structure are highly similar to Cry1Ac toxin. In this study, based on molecular design, a novel hapten polypeptide was synthesized and conjugated to keyhole limpet hemocyanin (KLH). Then, through animal immunization with this antigen, a monoclonal antibody named 2C12, showing high affinity to Cry1Ab and having no cross reaction with Cry1Ac, was produced. The equilibrium dissociation constant (K D ) value of Cry1Ab toxin with MAb 2C12 was 1.947 × 10 -8  M. Based on this specific monoclonal antibody, a sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was developed for the specific determination of Cry1Ab toxin and the LOD and LOQ values were determined as 0.47 ± 0.11 and 2.43 ± 0.19 ng mL -1 , respectively. The average recoveries of Cry1Ab from spiked rice leaf and rice flour samples ranged from 75 to 115%, with coefficient of variation (CV) less than 8.6% within the quantitation range (2.5-100 ng mL -1 ), showing good accuracy for the quantitative detection of Cry1Ab toxin in agricultural samples. In conclusion, this study provides a new approach for the production of high specific antibody and the newly developed DAS-ELISA is a useful method for Cry1Ab monitoring in agriculture products. Graphical Abstract Establishment of a DAS-ELISA for the specific detecting of Bacillus thuringiensis (Bt) Cry1Ab toxin.

Selective sequential separation of ABS/HIPS and PVC from automobile and electronic waste shredder residue by hybrid nano-Fe/Ca/CaO assisted ozonisation process.

PubMed

Mallampati, Srinivasa Reddy; Lee, Byoung Ho; Mitoma, Yoshiharu; Simion, Cristian

2017-02-01

The separation of plastics containing brominated flame retardants (BFR) like (acrylonitrile-butadiene-styrene (ABS), high-impact polystyrene (HIPS), and polyvinyl chloride (PVC)) from automobile and electronic waste shredder residue (ASR/ESR) are a major concern for thermal recycling. In laboratory scale tests using a hybrid nano-Fe/Ca/CaO assisted ozonation treatment has been found to selectively hydrophilize the surface of ABS/HIPS and PVC plastics, enhancing ABS wettability and thereby promoting its separation from ASR/ESR by means of froth flotation. The water contact angles, of ABS/HIPS and PVC decreased, about 18.7°, 18.3°, and 17.9° in ASR and about 21.2°, 20.7°, and 20.0° in ESR respectively. SEM-EDS, FT-IR, and XPS analyses demonstrated a marked decrease in [Cl] and a significant increase in the number of hydrophilic groups, such as CO, CO, and (CO)O, on the PVC or ABS surface. Under froth flotation conditions at 50rpm, about 99.1% of combined fraction of ABS/HIPS in ASR samples and 99.6% of ABS/HIPS in ESR samples were separated as settled fraction. After separation, the purity of the recovered combined ABS/HIPS fraction was 96.5% and 97.6% in ASR and ESR samples respectively. Furthermore, at 150rpm a 100% PVC separation in the settled fraction, with 98% and 99% purity in ASR and ESR plastics, respectively. Total recovery of non-ABS/HIPS and PVC plastics reached nearly 100% in the floating fraction. Further, this process improved the quality of recycled ASR/ESR plastics by removing surface contaminants or impurities. Copyright © 2017 Elsevier Ltd. All rights reserved.

Towards the implementation of quality by design to the production of therapeutic monoclonal antibodies with desired glycosylation patterns.

PubMed

del Val, Ioscani Jimenez; Kontoravdi, Cleo; Nagy, Judit M

2010-01-01

Quality by design (QbD) is a scheme for the development, manufacture, and approval of pharmaceutical products. The end goal of QbD is to ensure product quality by building it into the manufacturing process. The main regulatory bodies are encouraging its implementation to the manufacture of all new pharmaceuticals including biological products. Monoclonal antibodies (mAbs) are currently the leading products of the biopharmaceutical industry. It has been widely reported that glycosylation directly influences the therapeutic mechanisms by which mAbs function in vivo. In addition, glycosylation has been identified as one of the main sources of monoclonal antibody heterogeneity, and thus, a critical parameter to follow during mAb manufacture. This article reviews the research on glycosylation of mAbs over the past 2 decades under the QbD scope. The categories presented under this scope are: (a) definition of the desired clinical effects of mAbs, (b) definition of the glycosylation-associated critical quality attributes (glycCQAs) of mAbs, (c) assessment of process parameters that pose a risk for mAb glycCQAs, and (d) methods for accurately quantifying glycCQAs of mAbs. The information available in all four areas leads us to conclude that implementation of QbD to the manufacture of mAbs with specific glycosylation patterns will be a reality in the near future. We also foresee that the implementation of QbD will lead to the development of more robust and efficient manufacturing processes and to a new generation of mAbs with increased clinical efficacy. Copyright © 2010 American Institute of Chemical Engineers (AIChE).

Anti-nucleosome antibodies complexed to nucleosomal antigens show anti-DNA reactivity and bind to rat glomerular basement membrane in vivo.

PubMed Central

Kramers, C; Hylkema, M N; van Bruggen, M C; van de Lagemaat, R; Dijkman, H B; Assmann, K J; Smeenk, R J; Berden, J H

1994-01-01

Histones can mediate the binding of DNA and anti-DNA to the glomerular basement membrane (GBM). In ELISA histone/DNA/anti-DNA complexes are able to bind to heparan sulfate (HS), an intrinsic constituent of the GBM. We questioned whether histone containing immune complexes are able to bind to the GBM, and if so, whether the ligand in the GBM is HS. Monoclonal antibodies (mAbs) complexed to nucleosomal antigens and noncomplexed mAbs were isolated from culture supernatants of four IgG anti-nuclear mAbs. All noncomplexed mAbs showed strong anti-nucleosome reactivity in ELISA. One of them showed in addition anti-DNA reactivity in noncomplexed form. The other three mAbs only showed anti-DNA reactivity when they were complexed to nucleosomal antigens. After renal perfusion a fine granular binding of complexed mAbs to the glomerular capillary wall and activation of complement was observed in immunofluorescence, whereas noncomplexed mAbs did not bind. Immuno-electron microscopy showed binding of complexes to the whole width of the GBM. When HS in the GBM was removed by renal heparinase perfusion the binding of complexed mAb decreased, but did not disappear completely. We conclude that anti-nucleosome mAbs, which do not bind DNA, become DNA reactive once complexed to nucleosomal antigens. These complexed mAbs can bind to the GBM. The binding ligand in the GBM is partly, but not solely, HS. Binding to the GBM of immune complexes containing nucleosomal material might be an important event in the pathogenesis of lupus nephritis. Images PMID:8040312

B-Cell Epitopes in GroEL of Francisella tularensis

PubMed Central

Lu, Zhaohua; Rynkiewicz, Michael J.; Madico, Guillermo; Li, Sheng; Yang, Chiou-Ying; Perkins, Hillary M.; Sompuram, Seshi R.; Kodela, Vani; Liu, Tong; Morris, Timothy; Wang, Daphne; Roche, Marly I.; Seaton, Barbara A.; Sharon, Jacqueline

2014-01-01

The chaperonin protein GroEL, also known as heat shock protein 60 (Hsp60), is a prominent antigen in the human and mouse antibody response to the facultative intracellular bacterium Francisella tularensis (Ft), the causative agent of tularemia. In addition to its presumed cytoplasmic location, FtGroEL has been reported to be a potential component of the bacterial surface and to be released from the bacteria. In the current study, 13 IgG2a and one IgG3 mouse monoclonal antibodies (mAbs) specific for FtGroEL were classified into eleven unique groups based on shared VH-VL germline genes, and seven crossblocking profiles revealing at least three non-overlapping epitope areas in competition ELISA. In a mouse model of respiratory tularemia with the highly pathogenic Ft type A strain SchuS4, the Ab64 and N200 IgG2a mAbs, which block each other’s binding to and are sensitive to the same two point mutations in FtGroEL, reduced bacterial burden indicating that they target protective GroEL B-cell epitopes. The Ab64 and N200 epitopes, as well as those of three other mAbs with different crossblocking profiles, Ab53, N3, and N30, were mapped by hydrogen/deuterium exchange–mass spectrometry (DXMS) and visualized on a homology model of FtGroEL. This model was further supported by its experimentally-validated computational docking to the X-ray crystal structures of Ab64 and Ab53 Fabs. The structural analysis and DXMS profiles of the Ab64 and N200 mAbs suggest that their protective effects may be due to induction or stabilization of a conformational change in FtGroEL. PMID:24968190

Analysis of peripheral B cells and autoantibodies against the anti-nicotinic acetylcholine receptor derived from patients with myasthenia gravis using single-cell manipulation tools

PubMed Central

Terakawa, Maki; Muneoka, Satoshi; Nagahira, Kazuhiro; Nagane, Yuriko; Yamate, Jyoji; Motomura, Masakatsu; Utsugisawa, Kimiaki

2017-01-01

The majority of patients with myasthenia gravis (MG), an organ-specific autoimmune disease, harbor autoantibodies that attack the nicotinic acetylcholine receptor (nAChR-Abs) at the neuromuscular junction of skeletal muscles, resulting in muscle weakness. Single cell manipulation technologies coupled with genetic engineering are very powerful tools to examine T cell and B cell repertoires and the dynamics of adaptive immunity. These tools have been utilized to develop mAbs in parallel with hybridomas, phage display technologies and B-cell immortalization. By applying a single cell technology and novel high-throughput cell-based binding assays, we identified peripheral B cells that produce pathogenic nAChR-Abs in patients with MG. Although anti-nAChR antibodies produced by individual peripheral B cells generally exhibited low binding affinity for the α-subunit of the nAChR and great sequence diversity, a small fraction of these antibodies bound with high affinity to native-structured nAChRs on cell surfaces. B12L, one such Ab isolated here, competed with a rat Ab (mAb35) for binding to the human nAChR and thus considered to recognize the main immunogenic region (MIR). By evaluating the Ab in in vitro cell-based assays and an in vivo rat passive transfer model, B12L was found to act as a pathogenic Ab in rodents and presumably in humans.These findings suggest that B cells in peripheral blood may impact MG pathogenicity. Our methodology can be applied not only to validate pathogenic Abs as molecular target of MG treatment, but also to discover and analyze Ab production systems in other human diseases. PMID:29040265

Stellar Origin of 15N-rich Presolar SiC Grains of Type AB: Supernovae with Explosive Hydrogen Burning

NASA Astrophysics Data System (ADS)

Liu, Nan; Nittler, Larry R.; Pignatari, Marco; O'D. Alexander, Conel M.; Wang, Jianhua

2017-06-01

We report C, N, and Si isotopic data for 59 highly 13C-enriched presolar submicron- to micron-sized SiC grains from the Murchison meteorite, including eight putative nova grains (PNGs) and 29 15N-rich (14N/15N ≤ solar) AB grains, and their Mg-Al, S, and Ca-Ti isotope data when available. These 37 grains are enriched in 13C, 15N, and 26Al with the PNGs showing more extreme enhancements. The 15N-rich AB grains show systematically higher 26Al and 30Si excesses than the 14N-rich AB grains. Thus, we propose to divide the AB grains into groups 1 (14N/15N < solar) and 2 (14N/15N ≥ solar). For the first time, we have obtained both S and Ti isotopic data for five AB1 grains and one PNG and found 32S and/or 50Ti enhancements. Interestingly, one AB1 grain had the largest 32S and 50Ti excesses, strongly suggesting a neutron-capture nucleosynthetic origin of the 32S excess and thus the initial presence of radiogenic 32Si (t 1/2 = 153 years). More importantly, we found that the 15N and 26Al excesses of AB1 grains form a trend that extends to the region in the N-Al isotope plot occupied by C2 grains, strongly indicating a common stellar origin for both AB1 and C2 grains. Comparison of supernova models with the AB1 and C2 grain data indicates that these grains came from supernovae that experienced H ingestion into the He/C zones of their progenitors.

Increased Prevalence of Luminal Narrowing and Stricturing Identified by Enterography in Pediatric Crohn Disease Patients with Elevated Granulocyte-Macrophage Colony Stimulating Factor Auto-antibodies

PubMed Central

Dykes, Dana M.H.; Towbin, Alexander J.; Bonkowski, Erin; Chalk, Claudia; Bezold, Ramona; Lake, Kathleen; Kim, Mi-Ok; Heubi, James E.; Trapnell, Bruce C.; Podberesky, Daniel J.; Denson, Lee A.

2013-01-01

Background Crohn disease (CD) patients with elevated Granulocyte-Macrophage Colony-Stimulating Factor auto-antibodies (GM-CSF Ab) are more likely to develop stricturing behavior requiring surgery. Computed Tomography or Magnetic Resonance Enterography (CTE or MRE) may detect luminal narrowing (LN) prior to stricture development. Objective To determine whether CD patients with elevated GM-CSF Ab (≥ 1.6 mcg/mL) have a higher prevalence of LN and stricturing on CTE or MRE. Methods A single center, cross-sectional study of 153 pediatric CD patients and controls undergoing CTE or MRE. A novel scoring system evaluated for disease activity, presence of LN, stricture, intra-abdominal abscess, or fistulae Ouutcomes were compared with respect to antibody status using Fisher's exact test, logistic regression, and the unpaired t-test. Results GM-CSF Ab were elevated in CD patients (n=114) with a median (IQR) GM-CSF Ab level of 2.3 mcg/mL (0.5, 6.6) compared with healthy and disease controls, p=0.001. Ileal disease location was more common in CD patients with high GM-CSF Ab, p<0.001. Luminal narrowing increased from 39% in CD patients with low GM-CSF Ab to 71% in those with high levels (p=0.004). High GM-CSF Ab remained significantly associated with LN in a multivariate logistic model. Stricturing increased from 4% in CD patients with low GM-CSF Ab to 19% in those with high GM-CSF Ab (p=0.03). Conclusions Pediatric CD patients with high GM-CSF Ab levels have a higher prevalence of LN on CTE or MRE. Further study will be needed to determine whether medical therapy will reduce progression to stricturing behavior in these patients. PMID:23893081

A novel antiplatelet antibody therapy that induces cAMP-dependent endocytosis of the GPVI/Fc receptor γ-chain complex

PubMed Central

Takayama, Hiroshi; Hosaka, Yoshitaka; Nakayama, Kazuyuki; Shirakawa, Kamon; Naitoh, Katsuki; Matsusue, Tomokazu; Shinozaki, Mikihiko; Honda, Motoyasu; Yatagai, Yukiko; Kawahara, Tetsushi; Hirose, Jiro; Yokoyama, Tooru; Kurihara, Michiru; Furusako, Shoji

2008-01-01

Platelet adhesion to vascular subendothelium, mediated in part by interactions between collagen and glycoprotein VI (GPVI) complexed with Fc receptor γ-chain, is crucial for thrombus formation. Antiplatelet therapy benefits patients with various thrombotic and ischemic diseases, but the safety and efficacy of existing treatments are limited. Recent data suggest GPVI as a promising target for a novel antiplatelet therapy, for example, GPVI-specific Abs that deplete GPVI from the surface of platelets. Here, we characterized GPVI-specific auto-Abs (YA-Abs) from the first reported patient with ongoing platelet GPVI deficiency caused by the YA-Abs. To obtain experimentally useful human GPVI–specific mAbs with characteristics similar to YA-Abs, we generated human GPVI–specific mouse mAbs and selected 2 representative mAbs, mF1201 and mF1232, whose binding to GPVI was inhibited by YA-Abs. In vitro, mF1201, but not mF1232, induced human platelet activation and GPVI shedding, and mF1232 inhibited collagen-induced human platelet aggregation. Administration of mF1201 and mF1232 to monkeys caused GPVI immunodepletion with and without both significant thrombocytopenia and GPVI shedding, respectively. When a human/mouse chimeric form of mF1232 (cF1232) was labeled with a fluorescent endocytosis probe and administered to monkeys, fluorescence increased in circulating platelets and surface GPVI was lost. Loss of platelet surface GPVI mediated by cF1232 was successfully reproduced in vitro in the presence of a cAMP-elevating agent. Thus, we have characterized cAMP-dependent endocytosis of GPVI mediated by a human GPVI–specific mAb as what we believe to be a novel antiplatelet therapy. PMID:18382762

Three-dimensional quantitative structure-activity relationship modeling of cocaine binding by a novel human monoclonal antibody.

PubMed

Paula, Stefan; Tabet, Michael R; Farr, Carol D; Norman, Andrew B; Ball, W James

2004-01-01

Human monoclonal antibodies (mAbs) designed for immunotherapy have a high potential for avoiding the complications that may result from human immune system responses to the introduction of nonhuman mAbs into patients. This study presents a characterization of cocaine/antibody interactions that determine the binding properties of the novel human sequence mAb 2E2 using three-dimensional quantitative structure-activity relationship (3D-QSAR) methodology. We have experimentally determined the binding affinities of mAb 2E2 for cocaine and 38 cocaine analogues. The K(d) of mAb 2E2 for cocaine was 4 nM, indicating a high affinity. Also, mAb 2E2 displayed good cocaine specificity, as reflected in its 10-, 1500-, and 25000-fold lower binding affinities for the three physiologically relevant cocaine metabolites benzoylecgonine, ecgonine methyl ester, and ecgonine, respectively. 3D-QSAR models of cocaine binding were developed by comparative molecular similarity index analysis (CoMSIA). A model of high statistical quality was generated showing that cocaine binds to mAb 2E2 in a sterically restricted binding site that leaves the methyl group attached to the ring nitrogen of cocaine solvent-exposed. The methyl ester group of cocaine appears to engage in attractive van der Waals interactions with mAb 2E2, whereas the phenyl group contributes to the binding primarily via hydrophobic interactions. The model further indicated that an increase in partial positive charge near the nitrogen proton and methyl ester carbonyl group enhances binding affinity and that the ester oxygen likely forms an intermolecular hydrogen bond with mAb 2E2. Overall, the cocaine binding properties of mAb 2E2 support its clinical potential for development as a treatment of cocaine overdose and addiction.

Performance of the BioPlex 2200 HIV Ag-Ab assay for identifying acute HIV infection.

PubMed

Eshleman, Susan H; Piwowar-Manning, Estelle; Sivay, Mariya V; Debevec, Barbara; Veater, Stephanie; McKinstry, Laura; Bekker, Linda-Gail; Mannheimer, Sharon; Grant, Robert M; Chesney, Margaret A; Coates, Thomas J; Koblin, Beryl A; Fogel, Jessica M

Assays that detect HIV antigen (Ag) and antibody (Ab) can be used to screen for HIV infection. To compare the performance of the BioPlex 2200 HIV Ag-Ab assay and two other Ag/Ab combination assays for detection of acute HIV infection. Samples were obtained from 24 individuals (18 from the US, 6 from South Africa); these individuals were classified as having acute infection based on the following criteria: positive qualitative RNA assay; two negative rapid tests; negative discriminatory test. The samples were tested with the BioPlex assay, the ARCHITECT HIV Ag/Ab Combo test, the Bio-Rad GS HIV Combo Ag-Ab EIA test, and a viral load assay. Twelve (50.0%) of 24 samples had RNA detected only ( > 40 to 13,476 copies/mL). Ten (43.5%) samples had reactive results with all three Ag/Ab assays, one sample was reactive with the ARCHITECT and Bio-Rad assays, and one sample was reactive with the Bio-Rad and BioPlex assays. The 11 samples that were reactive with the BioPlex assay had viral loads from 83,010 to >750,000 copies/mL; 9/11 samples were classified as Ag positive/Ab negative by the BioPlex assay. Detection of acute HIV infection was similar for the BioPlex assay and two other Ag/Ab assays. All three tests were less sensitive than a qualitative RNA assay and only detected HIV Ag when the viral load was high. The BioPlex assay detected acute infection in about half of the cases, and identified most of those infections as Ag positive/Ab negative. Copyright © 2018 Elsevier B.V. All rights reserved.

Drosophila small heat shock protein CryAB ensures structural integrity of developing muscles, and proper muscle and heart performance.

PubMed

Wójtowicz, Inga; Jabłońska, Jadwiga; Zmojdzian, Monika; Taghli-Lamallem, Ouarda; Renaud, Yoan; Junion, Guillaume; Daczewska, Malgorzata; Huelsmann, Sven; Jagla, Krzysztof; Jagla, Teresa

2015-03-01

Molecular chaperones, such as the small heat shock proteins (sHsps), maintain normal cellular function by controlling protein homeostasis in stress conditions. However, sHsps are not only activated in response to environmental insults, but also exert developmental and tissue-specific functions that are much less known. Here, we show that during normal development the Drosophila sHsp CryAB [L(2)efl] is specifically expressed in larval body wall muscles and accumulates at the level of Z-bands and around myonuclei. CryAB features a conserved actin-binding domain and, when attenuated, leads to clustering of myonuclei and an altered pattern of sarcomeric actin and the Z-band-associated actin crosslinker Cheerio (filamin). Our data suggest that CryAB and Cheerio form a complex essential for muscle integrity: CryAB colocalizes with Cheerio and, as revealed by mass spectrometry and co-immunoprecipitation experiments, binds to Cheerio, and the muscle-specific attenuation of cheerio leads to CryAB-like sarcomeric phenotypes. Furthermore, muscle-targeted expression of CryAB(R120G), which carries a mutation associated with desmin-related myopathy (DRM), results in an altered sarcomeric actin pattern, in affected myofibrillar integrity and in Z-band breaks, leading to reduced muscle performance and to marked cardiac arrhythmia. Taken together, we demonstrate that CryAB ensures myofibrillar integrity in Drosophila muscles during development and propose that it does so by interacting with the actin crosslinker Cheerio. The evidence that a DRM-causing mutation affects CryAB muscle function and leads to DRM-like phenotypes in the fly reveals a conserved stress-independent role of CryAB in maintaining muscle cell cytoarchitecture. © 2015. Published by The Company of Biologists Ltd.

Evaluation of photobiomodulation therapy associated with guided bone regeneration in critical size defects. In vivo study.

PubMed

Freitas, Nicole Rosa de; Guerrini, Luísa Belluco; Esper, Luis Augusto; Sbrana, Michyele Cristhiane; Dalben, Gisele da Silva; Soares, Simone; Almeida, Ana Lúcia Pompéia Fraga de

2018-01-01

The repair of bone defects raises the interest of investigators in several health specialties. Grafting techniques with bone substitutes and laser therapies have been investigated to replace autogenous bone and accelerate the bone healing process. Objective To evaluate the effect of photobiomodulation therapy (PBMT) associated with guided bone regeneration (GBR) in critical size defects. Material and Methods The study was conducted on 80 male rats (Rattus norvegicus albinus, Wistar) submitted to surgical creation of a critical size defect on the calvaria, divided into eight study groups: group C (control - only blood clot); group M (collagen membrane); group PBMT (photobiomodulation therapy); group AB (autogenous bone); group AB+PBMT; group AB+M; group PBMT+M; group AB+PBMT+M. The animals were killed 30 days postoperatively. After tissue processing, bone regeneration was evaluated by histomorphometric analysis and statistical analyses were performed (Tukey test, p<0.05). Results All groups had greater area of newly formed bone compared to group C (9.96±4.49%). The group PBMT+M (achieved the greater quantity of new bone (64.09±7.62%), followed by groups PBMT (47.67±8.66%), M (47.43±15.73%), AB+PBMT (39.15±16.72%) and AB+PBMT+M (35.82±7.68%). After group C, the groups AB (25.10±16.59%) and AB+M (22.72±13.83%) had the smallest quantities of newly formed bone. The area of remaining particles did not have statistically significant difference between groups AB+M (14.93±8.92%) and AB+PBMT+M (14.76±6.58%). Conclusion The PBMT utilization may be effective for bone repair, when associated with bone regeneration techniques.

Modeling neutralization kinetics of HIV by broadly neutralizing monoclonal antibodies in genital secretions coating the cervicovaginal mucosa.

PubMed

McKinley, Scott A; Chen, Alex; Shi, Feng; Wang, Simi; Mucha, Peter J; Forest, M Gregory; Lai, Samuel K

2014-01-01

Eliciting broadly neutralizing antibodies (bnAb) in cervicovaginal mucus (CVM) represents a promising "first line of defense" strategy to reduce vaginal HIV transmission. However, it remains unclear what levels of bnAb must be present in CVM to effectively reduce infection. We approached this complex question by modeling the dynamic tally of bnAb coverage on HIV. This analysis introduces a critical, timescale-dependent competition: to protect, bnAb must accumulate at sufficient stoichiometry to neutralize HIV faster than virions penetrate CVM and reach target cells. We developed a model that incorporates concentrations and diffusivities of HIV and bnAb in semen and CVM, kinetic rates for binding (kon) and unbinding (koff) of select bnAb, and physiologically relevant thicknesses of CVM and semen layers. Comprehensive model simulations lead to robust conclusions about neutralization kinetics in CVM. First, due to the limited time virions in semen need to penetrate CVM, substantially greater bnAb concentrations than in vitro estimates must be present in CVM to neutralize HIV. Second, the model predicts that bnAb with more rapid kon, almost independent of koff, should offer greater neutralization potency in vivo. These findings suggest the fastest arriving virions at target cells present the greatest likelihood of infection. It also implies the marked improvements in in vitro neutralization potency of many recently discovered bnAb may not translate to comparable reduction in the bnAb dose needed to confer protection against initial vaginal infections. Our modeling framework offers a valuable tool to gaining quantitative insights into the dynamics of mucosal immunity against HIV and other infectious diseases.

The presence of anti-thyroglobulin (TgAb) and/or anti-thyroperoxidase antibodies (TPOAb) does not exclude the diagnosis of type 2 amiodarone-induced thyrotoxicosis.

PubMed

Tomisti, L; Urbani, C; Rossi, G; Latrofa, F; Sardella, C; Manetti, L; Lupi, I; Marcocci, C; Bartalena, L; Curzio, O; Martino, E; Bogazzi, F

2016-05-01

It is widely accepted that type 2 amiodarone-induced thyrotoxicosis (AIT) generally occurs in patients with a normal thyroid gland without signs of thyroid autoimmunity. However, it is currently unknown if the presence of anti-thyroglobulin (TgAb) and/or anti-thyroperoxidase antibodies (TPOAb) in AIT patients without other signs of an underlying thyroid disease may impair the response to glucocorticoid therapy. We performed a pilot retrospective cohort study with matched-subject design and an equivalence hypothesis, comparing the response to glucocorticoid therapy between 20 AIT patients with a normal thyroid gland, low radioiodine uptake, undetectable TSH receptor antibodies and positive TgAb and/or TPOAb (Ab+ group), and 40 patients with the same features and absent thyroid antibodies (Ab- group). The mean cure time was 54 ± 68 days in the Ab+ group and 55 ± 49 days in the Ab- group (p = 0.63). The equivalence test revealed an equivalent cure rate after 60, 90 and 180 days (p = 0.67, 0.88 and 0.278, respectively). The occurrence of permanent hypothyroidism was higher in the Ab+ group than in the Ab- group (26.3 vs 5.13 %, p = 0.032). The presence of TgAb and/or TPOAb does not affect the response to glucocorticoid therapy, suggesting that the patients with features of destructive form of AIT should be considered as having a type 2 AIT irrespective of the presence of TGAb or TPOAb. These patients have a higher risk of developing hypothyroidism after the resolution of thyrotoxicosis and should be monitored accordingly.

Deficiency of the ferrous iron transporter FeoAB is linked with metronidazole resistance in Bacteroides fragilis

PubMed Central

Veeranagouda, Yaligara; Husain, Fasahath; Boente, Renata; Moore, Jane; Smith, C. Jeffrey; Rocha, Edson R.; Patrick, Sheila; Wexler, Hannah M.

2014-01-01

Background Metronidazole is the most commonly used antimicrobial for Bacteroides fragilis infections and is recommended for prophylaxis of colorectal surgery. Metronidazole resistance is increasing and the mechanisms of resistance are not clear. Methods A transposon mutant library was generated in B. fragilis 638R (BF638R) to identify the genetic loci associated with resistance to metronidazole. Results Thirty-two independently isolated metronidazole-resistant mutants had a transposon insertion in BF638R_1421 that encodes the ferrous transport fusion protein (feoAB). Deletion of feoAB resulted in a 10-fold increased MIC of metronidazole for the strain. The metronidazole MIC for the feoAB mutant was similar to that for the parent strain when grown on media supplemented with excess iron, suggesting that the increase seen in the MIC of metronidazole was due to reduced cellular iron transport in the feoAB mutant. The furA gene repressed feoAB transcription in an iron-dependent manner and disruption of furA resulted in constitutive transcription of feoAB, regardless of whether or not iron was present. However, disruption of feoAB also diminished the capacity of BF638R to grow in a mouse intraperitoneal abscess model, suggesting that inorganic ferrous iron assimilation is essential for B. fragilis survival in vivo. Conclusions Selection for feoAB mutations as a result of metronidazole treatment will disable the pathogenic potential of B. fragilis and could contribute to the clinical efficacy of metronidazole. While mutations in feoAB are probably not a direct cause of clinical resistance, this study provides a key insight into intracellular metronidazole activity and the link with intracellular iron homeostasis. PMID:25028451

Molecular simulations of the pairwise interaction of monoclonal antibodies.

PubMed

Lapelosa, Mauro; Patapoff, Thomas W; Zarraga, Isidro E

2014-11-20

Molecular simulations are employed to compute the free energy of pairwise monoclonal antibodies (mAbs) association using a conformational sampling algorithm with a scoring function. The work reported here is aimed at investigating the mAb-mAb association driven by weak interactions with a computational method capable of predicting experimental observations of low binding affinity. The simulations are able to explore the free energy landscape. A steric interaction component, electrostatic interactions, and a nonpolar component of the free energy form the energy scoring function. Electrostatic interactions are calculated by solving the Poisson-Boltzmann equation. The nonpolar component is derived from the van der Waals interactions upon close contact of the protein surfaces. Two mAbs with similar IgG1 framework but with small sequence differences, mAb1 and mAb2, are considered for their different viscosity and propensity to form a weak interacting dimer. mAb1 presents favorable free energy of association at pH 6 with 15 mM of ion concentration reproducing experimental trends of high viscosity and dimer formation at high concentration. Free energy landscape and minimum free energy configurations of the dimer, as well as the second virial coefficient (B22) values are calculated. The energy distributions for mAb1 are obtained, and the most probable configurations are seen to be consistent with experimental measurements. In contrast, mAb2 shows an unfavorable average free energy at the same buffer conditions due to poor electrostatic complementarity, and reversible dimer configurations with favorable free energy are found to be unlikely. Finally, the simulations of the mAb association dynamics provide insights on the self-association responsible for bulk solution behavior and aggregation, which are important to the processing and the quality of biopharmaceuticals.

Monoclonal antibody, mAb 4C13, an effective detoxicant antibody against ricin poisoning.

PubMed

Dong, Na; Luo, Longlong; Wu, Junhua; Jia, Peiyuan; Li, Qian; Wang, Yuxia; Gao, Zhongcai; Peng, Hui; Lv, Ming; Huang, Chunqian; Feng, Jiannan; Li, Hua; Shan, Junjie; Han, Gang; Shen, Beifen

2015-07-31

Ricin is a glycoprotein produced in castor seeds and consists of two polypeptide chains named Ricin Toxin A Chain (RTA) and Ricin Toxin B Chain (RTB), linked via a disulfide bridge. Due to its high toxicity, ricin is regarded as a high terrorist risk for the public. However, antibodies can play a pivotal role in neutralizing the toxin. In this research, the anti-toxicant effect of mAb 4C13, a monoclonal antibody (mAb) established using detoxicated ricin as the immunized antigen, was evaluated. Compared with mAb 4F2 and mAb 5G6, the effective mechanism of mAb 4C13 was analyzed by experiments relating to its cytotoxicity, epitope on ricin, binding kinetics with the toxin, its blockage on the protein synthesis inhibition induced by ricin and the intracelluar tracing of its complex with ricin. Our result indicated that mAb 4C13 could recognize and bind to RTA, RTB and exert its high affinity to the holotoxin. Both cytotoxicity and animal toxicity of ricin were well blocked by pre-incubating the toxin with mAb 4C13. By intravenous injection, mAb 4C13 could rescue the mouse intraperitoneally (ip) injected with a lethal dose of ricin (20μg/kg) even at 6h after the intoxication and its efficacy was dependent on its dosage. This research indicated that mAb 4C13 could be an excellent candidate for therapeutic antibodies. Its potent antitoxic efficiency was related to its recognition on the specific epitope with very high affinity and its blockage of protein synthesis inhibition in cytoplasm followed by cellular internalization with ricin. Copyright © 2015 Elsevier Ltd. All rights reserved.

A novel L-asparaginase from Aquabacterium sp. A7-Y with self-cleavage activation.

PubMed

Sun, Zhibin; Li, Ding; Liu, Pingping; Wang, Wenhui; Ji, Kai; Huang, Yan; Cui, Zhongli

2016-01-01

We have identified a novel L-asparaginase, abASNase3, from Aquabacterium sp. A7-Y. abASNase3 is composed of 306 amino acids and exhibits 34 % sequence homology to human asparaginase (hASNase3). Further analysis revealed that abASNase3 belongs to the N-terminal nucleophile (Ntn) family of hydrolases. Previous reports about the Ntn hydrolase family and the results of our study suggest that abASNase3 must form two subunits by self-cleavage between Gly189 and Thr190 to attain catalytic activity. The two subunits remained tightly associated to build a single functional unit. The optimum pH for abASNase3 was found to be 8.0 in Tris-HCl buffer and the enzyme was found to be stable over a broad pH range from pH 6.0 to 12.0. The optimum temperature for abASNase3 was found to be approximately 40 °C, and the enzyme was stable below 65 °C. abASNase3 showed high substrate specificity toward L-asparagine and had no or only slight activity toward D-asparagine, L-glutamine and D-glutamine. abASNase3 was significantly activated by Mg(2+) and was substantially inhibited by Ni(2+), Cu(2+), Mn(2+) and Co(2+). The Michaelis-Menten constant and turnover number of abASNase3 for L-asparagine were estimated to be 3.37 × 10(-2) M and 8.72 × 10(-3) s(-1), respectively. Our results indicate that abASNase3 is a novel L-asparaginase in the Ntn family of hydrolases.

Highly viscous antibody solutions are a consequence of network formation caused by domain-domain electrostatic complementarities: insights from coarse-grained simulations.

PubMed

Buck, Patrick M; Chaudhri, Anuj; Kumar, Sandeep; Singh, Satish K

2015-01-05

Therapeutic monoclonal antibody (mAb) candidates that form highly viscous solutions at concentrations above 100 mg/mL can lead to challenges in bioprocessing, formulation development, and subcutaneous drug delivery. Earlier studies of mAbs with concentration-dependent high viscosity have indicated that mAbs with negatively charged Fv regions have a dipole-like quality that increases the likelihood of reversible self-association. This suggests that weak electrostatic intermolecular interactions can form transient antibody networks that participate in resistance to solution deformation under shear stress. Here this hypothesis is explored by parametrizing a coarse-grained (CG) model of an antibody using the domain charges from four different mAbs that have had their concentration-dependent viscosity behaviors previously determined. Multicopy molecular dynamics simulations were performed for these four CG mAbs at several concentrations to understand the effect of surface charge on mass diffusivity, pairwise interactions, and electrostatic network formation. Diffusion coefficients computed from simulations were in qualitative agreement with experimentally determined viscosities for all four mAbs. Contact analysis revealed an overall greater number of pairwise interactions for the two mAbs in this study with high concentration viscosity issues. Further, using equilibrated solution trajectories, the two mAbs with high concentration viscosity issues quantitatively formed more features of an electrostatic network than the other mAbs. The change in the number of these network features as a function of concentration is related to the number of pairwise interactions formed by electrostatic complementarities between antibody domains. Thus, transient antibody network formation caused by domain-domain electrostatic complementarities is the most probable origin of high concentration viscosity for mAbs in this study.

Neuropsychiatric disease relevance of circulating anti-NMDA receptor autoantibodies depends on blood-brain barrier integrity.

PubMed

Hammer, C; Stepniak, B; Schneider, A; Papiol, S; Tantra, M; Begemann, M; Sirén, A-L; Pardo, L A; Sperling, S; Mohd Jofrry, S; Gurvich, A; Jensen, N; Ostmeier, K; Lühder, F; Probst, C; Martens, H; Gillis, M; Saher, G; Assogna, F; Spalletta, G; Stöcker, W; Schulz, T F; Nave, K-A; Ehrenreich, H

2014-10-01

In 2007, a multifaceted syndrome, associated with anti-NMDA receptor autoantibodies (NMDAR-AB) of immunoglobulin-G isotype, has been described, which variably consists of psychosis, epilepsy, cognitive decline and extrapyramidal symptoms. Prevalence and significance of NMDAR-AB in complex neuropsychiatric disease versus health, however, have remained unclear. We tested sera of 2817 subjects (1325 healthy, 1081 schizophrenic, 263 Parkinson and 148 affective-disorder subjects) for presence of NMDAR-AB, conducted a genome-wide genetic association study, comparing AB carriers versus non-carriers, and assessed their influenza AB status. For mechanistic insight and documentation of AB functionality, in vivo experiments involving mice with deficient blood-brain barrier (ApoE(-/-)) and in vitro endocytosis assays in primary cortical neurons were performed. In 10.5% of subjects, NMDAR-AB (NR1 subunit) of any immunoglobulin isotype were detected, with no difference in seroprevalence, titer or in vitro functionality between patients and healthy controls. Administration of extracted human serum to mice influenced basal and MK-801-induced activity in the open field only in ApoE(-/-) mice injected with NMDAR-AB-positive serum but not in respective controls. Seropositive schizophrenic patients with a history of neurotrauma or birth complications, indicating an at least temporarily compromised blood-brain barrier, had more neurological abnormalities than seronegative patients with comparable history. A common genetic variant (rs524991, P=6.15E-08) as well as past influenza A (P=0.024) or B (P=0.006) infection were identified as predisposing factors for NMDAR-AB seropositivity. The >10% overall seroprevalence of NMDAR-AB of both healthy individuals and patients is unexpectedly high. Clinical significance, however, apparently depends on association with past or present perturbations of blood-brain barrier function.

Toxicokinetic and toxicodynamic analyses of Androctonus australis hector venom in rats: Optimization of antivenom therapy

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

Hammoudi-Triki, D.; Laboratoire de Biologie Cellulaire et Moleculaire, Faculte des Sciences Biologiques, Universite des Sciences et de la Technologie 'Houari Boumedienne' Bab Ezzouar, Alger, Algerie; Laboratoire de Recherche et de Developpement sur les Venins, Institut Pasteur d'Algerie, Algerie

2007-02-01

This paper reports the simultaneous determination of toxicokinetic and toxicodynamic properties of Androctonus australis hector venom, in the absence and presence of antivenom (F(ab'){sub 2} and Fab), in envenomed rats. After subcutaneous injection of the venom, toxins showed a complete absorption phase from the site of injection associated with a distribution into a large extravascular compartment. The injection of Fab and F(ab'){sub 2} induced the neutralization of venom antigens in the blood compartment, as well as the redistribution of venom components from the extravascular compartment to the blood compartment. Interestingly, F(ab'){sub 2} and Fab showed distinct efficiencies depending on theirmore » route of injection. F(ab'){sub 2} induced a faster venom neutralization and redistribution than Fab when injected intravenously. Fab was more effective than F(ab'){sub 2} by the intramuscular route. The hemodynamic effects of Aah venom were further investigated. Changes in mean arterial pressure and heart rate were observed in parallel with an upper airway obstruction. Fab was more effective than F(ab'){sub 2} for preventing early symptoms of envenomation, whatever their route of administration. Intraperitoneal injection of F(ab'){sub 2} and Fab was similar for the prevention of the delayed symptoms, even after a late administration. Fab was more effective than F(ab'){sub 2} in the inhibition of airway resistance, independent of the route and time of administration. These results show that the treatment for scorpion stings might be improved by the intravascular injection of a mixture of Fab and F(ab'){sub 2}. If antivenom cannot be administered intravenously, Fab might be an alternative as they are more effective than F(ab'){sub 2} when injected intramuscularly.« less

Analysis of peripheral B cells and autoantibodies against the anti-nicotinic acetylcholine receptor derived from patients with myasthenia gravis using single-cell manipulation tools.

PubMed

Makino, Tomohiro; Nakamura, Ryuichi; Terakawa, Maki; Muneoka, Satoshi; Nagahira, Kazuhiro; Nagane, Yuriko; Yamate, Jyoji; Motomura, Masakatsu; Utsugisawa, Kimiaki

2017-01-01

The majority of patients with myasthenia gravis (MG), an organ-specific autoimmune disease, harbor autoantibodies that attack the nicotinic acetylcholine receptor (nAChR-Abs) at the neuromuscular junction of skeletal muscles, resulting in muscle weakness. Single cell manipulation technologies coupled with genetic engineering are very powerful tools to examine T cell and B cell repertoires and the dynamics of adaptive immunity. These tools have been utilized to develop mAbs in parallel with hybridomas, phage display technologies and B-cell immortalization. By applying a single cell technology and novel high-throughput cell-based binding assays, we identified peripheral B cells that produce pathogenic nAChR-Abs in patients with MG. Although anti-nAChR antibodies produced by individual peripheral B cells generally exhibited low binding affinity for the α-subunit of the nAChR and great sequence diversity, a small fraction of these antibodies bound with high affinity to native-structured nAChRs on cell surfaces. B12L, one such Ab isolated here, competed with a rat Ab (mAb35) for binding to the human nAChR and thus considered to recognize the main immunogenic region (MIR). By evaluating the Ab in in vitro cell-based assays and an in vivo rat passive transfer model, B12L was found to act as a pathogenic Ab in rodents and presumably in humans.These findings suggest that B cells in peripheral blood may impact MG pathogenicity. Our methodology can be applied not only to validate pathogenic Abs as molecular target of MG treatment, but also to discover and analyze Ab production systems in other human diseases.

Marker of cemento-periodontal ligament junction associated with periodontal regeneration.

PubMed

Hara, Ryohko; Wato, Masahiro; Tanaka, Akio

2005-06-01

The purpose of this study was to identify factors promoting formation of the cemento-periodontal ligament junction. Regeneration of the cemento-periodontal ligament junction is an important factor in recovery of the connective tissue attachment to the cementum and it is important to identify all specific substances that promote its formation. To clarify the substances involved in cemento-periodontal ligament junction formation, we produced a monoclonal antibody (mAb) to human cemento-periodontal ligament junction (designated as the anti-TAP mAb) and examined its immunostaining properties and reactive antigen. Hybridomas producing monoclonal antibody against human cemento-periodontal ligament junction antigens were established by fusing P3U1 mouse myeloma cells with spleen cells from BALB/c mice immunized with homogenized human cemento-periodontal ligament junction. The mAb, the anti-TAP mAb for cemento-periodontal ligament junction, was then isolated. The immunoglobulin class and light chain of the mAb were examined using an isotyping kit. Before immunostaining, antigen determination using an enzymatic method or heating was conducted. Human teeth, hard tissue-forming lesions, and animal tissues were immunostained by the anti-TAP mAb. The anti-TAP mAb was positive in human cemento-periodontal ligament junction and predentin but negative in all other human and animal tissues examined. In the cemento-osseous lesions, the anti-TAP mAb was positive in the peripheral area of the cementum and cementum-like hard tissues and not in the bone and bone-like tissues. The anti-TAP mAb showed IgM (kappa) and recognized phosphoprotein. The anti-TAP mAb is potentially useful for developing new agents promoting cementogenesis and periodontal regeneration.

Macrolide resistance in Legionella pneumophila: the role of LpeAB efflux pump.

PubMed

Massip, Clémence; Descours, Ghislaine; Ginevra, Christophe; Doublet, Patricia; Jarraud, Sophie; Gilbert, Christophe

2017-05-01

A previous study on 12 in vitro -selected azithromycin-resistant Legionella pneumophila lineages showed that ribosomal mutations were major macrolide resistance determinants. In addition to these mechanisms that have been well described in many species, mutations upstream of lpeAB operon, homologous to acrAB in Escherichia coli , were identified in two lineages. In this study, we investigated the role of LpeAB and of these mutations in macrolide resistance of L. pneumophila . The role of LpeAB was studied by testing the antibiotic susceptibility of WT, deleted and complemented L. pneumophila Paris strains. Translational fusion experiments using GFP as a reporter were conducted to investigate the consequences of the mutations observed in the upstream sequence of lpeAB operon. We demonstrated the involvement of LpeAB in an efflux pump responsible for a macrolide-specific reduced susceptibility of L. pneumophila Paris strain. Mutations in the upstream sequence of lpeAB operon were associated with an increased protein expression. Increased expression was also observed under sub-inhibitory macrolide concentrations in strains with both mutated and WT promoting regions. LpeAB are components of an efflux pump, which is a macrolide resistance determinant in L. pneumophila Paris strain. Mutations observed in the upstream sequence of lpeAB operon in resistant lineages led to an overexpression of this efflux pump. Sub-inhibitory concentrations of macrolides themselves participated in upregulating this efflux and could constitute a first step in the acquisition of a high macrolide resistance level. © The Author 2017. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Is there a place for nutrition-sensitive agriculture?

PubMed

Wambugu, Florence; Obukosia, Silas; Gaffney, Jim; Kamanga, Daniel; Che, Ping; Albertsen, Marc C; Zhao, Zuo-Yu; Ragland, Lonnetta; Yeye, Mary; Kimani, Esther; Aba, Daniel; Gidado, Rose; Solomon, B O; Njuguna, Michael

2015-11-01

The focus of the review paper is to discuss how biotechnological innovations are opening new frontiers to mitigate nutrition in key agricultural crops with potential for large-scale health impact to people in Africa. The general objective of the Africa Biofortified Sorghum (ABS) project is to develop and deploy sorghum with enhanced pro-vitamin A to farmers and end-users in Africa to alleviate vitamin A-related micronutrient deficiency diseases. To achieve this objective the project technology development team has developed several promising high pro-vitamin A sorghum events. ABS 203 events are so far the most advanced and well-characterised lead events with about 12 μg β-carotene/g tissue which would supply about 40-50 % of the daily recommended vitamin A at harvest. Through gene expression optimisation other events with higher amounts of pro-vitamin A, including ABS 214, ABS 235, ABS 239 with 25, 30-40, 40-50 μg β-carotene/g tissue, respectively, have been developed. ABS 239 would provide twice recommended pro-vitamin A at harvest, 50-90 % after 3 months storage and 13-45 % after 6 months storage for children. Preliminary results of introgression of ABS pro-vitamin A traits into local sorghum varieties in target countries Nigeria and Kenya show stable introgression of ABS vitamin A into local farmer-preferred sorghums varieties. ABS gene Intellectual Property Rights and Freedom to Operate have been donated for use royalty free for Africa. Prior to the focus on the current target countries, the project was implemented by fourteen institutions in Africa and the USA. For the next 5 years, the project will complete ABS product development, complete regulatory science data package and apply for product deregulation in target African countries.

Selection of Single Domain Antibodies from Immune Libraries Displayed on the Surface of E. coli Cells with Two β-Domains of Opposite Topologies

PubMed Central

Martínez-Arteaga, Rocio; Ruano-Gallego, David; Fraile, Sofía; Margolles, Yago; Teira, Xema; Gutierrez, Carlos; Bodelón, Gustavo; Fernández, Luis Ángel

2013-01-01

Screening of antibody (Ab) libraries by direct display on the surface of E. coli cells is hampered by the presence of the outer membrane (OM). In this work we demonstrate that the native β-domains of EhaA autotransporter and intimin, two proteins from enterohemorrhagic E. coli O157:H7 (EHEC) with opposite topologies in the OM, are effective systems for the display of immune libraries of single domain Abs (sdAbs) from camelids (nanobodies or VHH) on the surface of E. coli K-12 cells and for the selection of high affinity sdAbs using magnetic cell sorting (MACS). We analyzed the capacity of EhaA and intimin β-domains to display individual sdAbs and sdAb libraries obtained after immunization with the extracellular domain of the translocated intimin receptor from EHEC (TirMEHEC). We demonstrated that both systems displayed functional sdAbs on the surface of E. coli cells with little proteolysis and cellular toxicity, although E. coli cells displaying sdAbs with the β-domain of intimin showed higher antigen-binding capacity. Both E. coli display libraries were screened for TirMEHEC binding clones by MACS. High affinity binders were selected by both display systems, although more efficiently with the intimin β-domain. The specificity of the selected clones against TirMEHEC was demonstrated by flow cytometry of E. coli cells, along with ELISA and surface plasmon resonance with purified sdAbs. Finally, we employed the E. coli cell display systems to provide an estimation of the affinity of the selected sdAb by flow cytometry analysis under equilibrium conditions. PMID:24086454

Neutron Reflection Study of Surface Adsorption of Fc, Fab, and the Whole mAb.

PubMed

Li, Zongyi; Li, Ruiheng; Smith, Charles; Pan, Fang; Campana, Mario; Webster, John R P; van der Walle, Christopher F; Uddin, Shahid; Bishop, Steve M; Narwal, Rojaramani; Warwicker, Jim; Lu, Jian Ren

2017-07-12

Characterizing the influence of fragment crystallization (Fc) and antigen-binding fragment (Fab) on monoclonal antibody (mAb) adsorption at the air/water interface is an important step to understanding liquid mAb drug product stability during manufacture, shipping, and storage. Here, neutron reflection is used to study the air/water adsorption of a mAb and its Fc and Fab fragments. By varying the isotopic contrast, the adsorbed amount, thickness, orientation, and immersion of the adsorbed layers could be determined unambiguously. While Fc adsorption reached saturation within the hour, its surface adsorbed amount showed little variation with bulk concentration. In contrast, Fab adsorption was slower and the adsorbed amount was concentration dependent. The much higher Fc adsorption, as compared to Fab, was linked to its lower surface charge. Time and concentration dependence of mAb adsorption was dominated by Fab behavior, although both Fab and Fc behaviors contributed to the amount of mAb adsorbed. Changing the pH from 5.5 to 8.8 did not much perturb the adsorbed amount of Fc, Fab, or mAb. However, a small decrease in adsorption was observed for the Fc over pH 8-8.8 and vice versa for the Fab and mAb, consistent with a dominant Fab behavior. As bulk concentration increased from 5 to 50 ppm, the thicknesses of the Fc layers were almost constant at 40 Å, while Fab and mAb layers increased from 45 to 50 Å. These results imply that the adsorbed mAb, Fc, and Fab all retained their globular structures and were oriented with their short axial lengths perpendicular to the interface.

Dopamine and temporal attention: An attentional blink study in Parkinson's disease patients on and off medication.

PubMed

Slagter, H A; van Wouwe, N C; Kanoff, K; Grasman, R P P P; Claassen, D O; van den Wildenberg, W P M; Wylie, S A

2016-10-01

The current study aimed to shed more light on the role of dopamine in temporal attention. To this end, we pharmacologically manipulated dopamine levels in a large sample of Parkinson's disease patients (n=63) while they performed an attentional blink (AB) task in which they had to identify two targets (T1 and T2) presented in close temporal proximity among distractors. We specifically examined 1) differences in the magnitude of the AB between unmedicated Parkinson patients, who have depleted levels of striatal dopamine, and healthy controls, and 2) effects of two dopaminergic medications (l-DOPA and dopamine agonists) on the AB in the Parkinson patients at the group level and as a function of individual baseline performance. In line with the notion that relatively low levels of striatal dopamine may impair target detection in general, Parkinson patients OFF medications displayed overall poor target perception compared to healthy controls. Moreover, as predicted, effects of dopaminergic medication on AB performance critically depended on individual baseline AB size, although this effect was only observed for l-DOPA. l-DOPA generally decreased the size of the AB in patients with a large baseline AB (i.e., OFF medications), while l-DOPA generally increased the AB in patients with a small baseline AB. These findings may support a role for dopamine in the AB and temporal attention, more generally and corroborate the notion that there is an optimum dopamine level for cognitive function. They also emphasize the need for more studies that examine the separate effects of DA agonists and l-DOPA on cognitive functioning. Copyright © 2016 Elsevier Ltd. All rights reserved.

Autonomic control of post-air-breathing tachycardia in Clarias gariepinus (Teleostei: Clariidae).

PubMed

Teixeira, Mariana Teodoro; Armelin, Vinicius Araújo; Abe, Augusto Shinya; Rantin, Francisco Tadeu; Florindo, Luiz Henrique

2015-08-01

The African catfish (Clarias gariepinus) is a teleost with bimodal respiration that utilizes a paired suprabranchial chamber located in the gill cavity as an air-breathing organ. Like all air-breathing fishes studied to date, the African catfish exhibits pronounced changes in heart rate (f H) that are associated with air-breathing events. We acquired f H, gill-breathing frequency (f G) and air-breathing frequency (f AB) in situations that require or do not require air breathing (during normoxia and hypoxia), and we assessed the autonomic control of post-air-breathing tachycardia using an infusion of the β-adrenergic antagonist propranolol and the muscarinic cholinergic antagonist atropine. During normoxia, C. gariepinus presented low f AB (1.85 ± 0.73 AB h(-1)) and a constant f G (43.16 ± 1.74 breaths min(-1)). During non-critical hypoxia (PO2 = 60 mmHg), f AB in the African catfish increased to 5.42 ± 1.19 AB h(-1) and f G decreased to 39.12 ± 1.58 breaths min(-1). During critical hypoxia (PO2 = 20 mmHg), f AB increased to 7.4 ± 1.39 AB h(-1) and f G decreased to 34.97 ± 1.78 breaths min(-1). These results were expected for a facultative air breather. Each air breath (AB) was followed by a brief but significant tachycardia, which in the critical hypoxia trials, reached a maximum of 143 % of the pre-AB f H values of untreated animals. Pharmacological blockade allowed the calculation of cardiac autonomic tones, which showed that post-AB tachycardia is predominantly regulated by the parasympathetic subdivision of the autonomic nervous system.

Efficient clearance of Aβ protofibrils in AβPP-transgenic mice treated with a brain-penetrating bifunctional antibody.

PubMed

Syvänen, Stina; Hultqvist, Greta; Gustavsson, Tobias; Gumucio, Astrid; Laudon, Hanna; Söderberg, Linda; Ingelsson, Martin; Lannfelt, Lars; Sehlin, Dag

2018-05-24

Amyloid-β (Aβ) immunotherapy is one of the most promising disease-modifying strategies for Alzheimer's disease (AD). Despite recent progress targeting aggregated forms of Aβ, low antibody brain penetrance remains a challenge. In the present study, we used transferrin receptor (TfR)-mediated transcytosis to facilitate brain uptake of our previously developed Aβ protofibril-selective mAb158, with the aim of increasing the efficacy of immunotherapy directed toward soluble Aβ protofibrils. Aβ protein precursor (AβPP)-transgenic mice (tg-ArcSwe) were given a single dose of mAb158, modified for TfR-mediated transcytosis (RmAb158-scFv8D3), in comparison with an equimolar dose or a tenfold higher dose of unmodified recombinant mAb158 (RmAb158). Soluble Aβ protofibrils and total Aβ in the brain were measured by enzyme-linked immunosorbent assay (ELISA). Brain distribution of radiolabeled antibodies was visualized by positron emission tomography (PET) and ex vivo autoradiography. ELISA analysis of Tris-buffered saline brain extracts demonstrated a 40% reduction of soluble Aβ protofibrils in both RmAb158-scFv8D3- and high-dose RmAb158-treated mice, whereas there was no Aβ protofibril reduction in mice treated with a low dose of RmAb158. Further, ex vivo autoradiography and PET imaging revealed different brain distribution patterns of RmAb158-scFv8D3 and RmAb158, suggesting that these antibodies may affect Aβ levels by different mechanisms. With a combination of biochemical and imaging analyses, this study demonstrates that antibodies engineered to be transported across the blood-brain barrier can be used to increase the efficacy of Aβ immunotherapy. This strategy may allow for decreased antibody doses and thereby reduced side effects and treatment costs.

Monoclonal antibodies and toxins--a perspective on function and isotype.

PubMed

Chow, Siu-Kei; Casadevall, Arturo

2012-06-01

Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins--Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)--and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions.

Monoclonal Antibodies and Toxins—A Perspective on Function and Isotype

PubMed Central

Chow, Siu-Kei; Casadevall, Arturo

2012-01-01

Antibody therapy remains the only effective treatment for toxin-mediated diseases. The development of hybridoma technology has allowed the isolation of monoclonal antibodies (mAbs) with high specificity and defined properties, and numerous mAbs have been purified and characterized for their protective efficacy against different toxins. This review summarizes the mAb studies for 6 toxins—Shiga toxin, pertussis toxin, anthrax toxin, ricin toxin, botulinum toxin, and Staphylococcal enterotoxin B (SEB)—and analyzes the prevalence of mAb functions and their isotypes. Here we show that most toxin-binding mAbs resulted from immunization are non-protective and that mAbs with potential therapeutic use are preferably characterized. Various common practices and caveats of protection studies are discussed, with the goal of providing insights for the design of future research on antibody-toxin interactions. PMID:22822456

FTIR analysis on aging characteristics of ABS/PC blend under UV-irradiation in air

NASA Astrophysics Data System (ADS)

Li, Jiarong; Chen, Fu; Yang, Long; Jiang, Long; Dan, Yi

2017-09-01

Fourier Transform Infrared Spectroscopy (FTIR) is adopted to study the aging characteristics of poly(acrylonitrile-butadiene-styrene)/polycarbonate (ABS/PC) blend under UV-irradiation in air by analyzing the variation of the three main absorbance at about 967 cm- 1, 1720 cm- 1 and 3420 cm- 1 associated with carbon-hydrogen bonds belonging to 1,4 butadiene, carbonyl and hydroxyl groups, respectively. Results indicate that, under UV-irradiation in air, the photo-oxidation of the blend is not a simple combination of the photo-oxidation of corresponding ABS and PC themselves and takes place predominantly at the ABS component. Due to the interaction between the two components and the Fries rearrangement taken place in the PC component during the UV-irradiation in air, the ABS/PC blends behave higher photo-stability than ABS has.

Specificity of Mimotope-Induced Anti-High Molecular Weight-Melanoma Associated Antigen (HMW-MAA) Antibodies Does Not Ensure Biological Activity

PubMed Central

Hofstetter, Gerlinde; Balazs, Nina; Smole, Ursula; Ferrone, Soldano; Scheiner, Otto; Breiteneder, Heimo; Pehamberger, Hubert; Wagner, Stefan

2011-01-01

Vaccines based on peptide mimics (mimotopes) of conformational tumor antigen epitopes have been investigated for a variety of human tumors including breast cancer, tumors expressing the carcinoembryonic antigen, B cell lymphoma, neuroblastoma, and melanoma. In our previous work, we designed a vaccine based on a mimotope of the high molecular weight-melanoma associated antigen (HMW-MAA) that elicited HMW-MAA-specific antibodies (Abs) with anti-tumor activity in vitro and in vivo. In this study, we aimed to identify mimotopes of additional distinct HMW-MAA epitopes, since they could be used to construct a polymimotope melanoma vaccine. For this purpose, random peptide phage libraries were screened with the anti-HMW-MAA monoclonal antibodies (mAbs) VT80.12 and VF1-TP43 yielding one peptide ligand for each mAb. Both peptides inhibited the binding of the corresponding mAb to the HMW-MAA. Furthermore, when coupled to the carrier protein keyhole limpet hemocyanin (KLH), both HMW-MAA mimotopes elicited peptide-specific Abs in rabbits or BALB/c mice, but only the mimotope isolated with the mAb VT80.12 elicited HMW-MAA-specific Abs and only in mice. However, the latter Abs had no detectable effect on HMW-MAA expressing human melanoma cells in vitro. These results describe limitations related to the phage display technique and emphasize the need to characterize the functional properties of the mAb utilized to isolate mimotopes of the corresponding epitopes. PMID:21573118

Specificity of mimotope-induced anti-high molecular weight-melanoma associated antigen (HMW-MAA) antibodies does not ensure biological activity.

PubMed

Latzka, Julia; Gaier, Sonja; Hofstetter, Gerlinde; Balazs, Nina; Smole, Ursula; Ferrone, Soldano; Scheiner, Otto; Breiteneder, Heimo; Pehamberger, Hubert; Wagner, Stefan

2011-05-06

Vaccines based on peptide mimics (mimotopes) of conformational tumor antigen epitopes have been investigated for a variety of human tumors including breast cancer, tumors expressing the carcinoembryonic antigen, B cell lymphoma, neuroblastoma, and melanoma. In our previous work, we designed a vaccine based on a mimotope of the high molecular weight-melanoma associated antigen (HMW-MAA) that elicited HMW-MAA-specific antibodies (Abs) with anti-tumor activity in vitro and in vivo. In this study, we aimed to identify mimotopes of additional distinct HMW-MAA epitopes, since they could be used to construct a polymimotope melanoma vaccine. For this purpose, random peptide phage libraries were screened with the anti-HMW-MAA monoclonal antibodies (mAbs) VT80.12 and VF1-TP43 yielding one peptide ligand for each mAb. Both peptides inhibited the binding of the corresponding mAb to the HMW-MAA. Furthermore, when coupled to the carrier protein keyhole limpet hemocyanin (KLH), both HMW-MAA mimotopes elicited peptide-specific Abs in rabbits or BALB/c mice, but only the mimotope isolated with the mAb VT80.12 elicited HMW-MAA-specific Abs and only in mice. However, the latter Abs had no detectable effect on HMW-MAA expressing human melanoma cells in vitro. These results describe limitations related to the phage display technique and emphasize the need to characterize the functional properties of the mAb utilized to isolate mimotopes of the corresponding epitopes.

Separation of polycarbonate and acrylonitrile–butadiene–styrene waste plastics by froth flotation combined with ammonia pretreatment

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

Wang, Chong-qing; Wang, Hui, E-mail: huiwang1968@163.com; Liu, Qun

Highlights: • Ammonia treatment changes selectively floatability of PC. • The effects of ammonia on PC were investigated through contact angle and XPS. • Reactions between ammonia and PC surface make PC more hydrophilic. • PC and ABS mixtures with different particle sizes were separated effectively. - Abstract: The objective of this research is flotation separation of polycarbonate (PC) and acrylonitrile–butadiene–styrene (ABS) waste plastics combined with ammonia pretreatment. The PC and ABS plastics show similar hydrophobicity, and ammonia treatment changes selectively floatability of PC plastic while ABS is insensitive to ammonia treatment. The contact angle measurement indicates the dropping ofmore » flotation recovery of PC is ascribed to a decline of contact angle. X-ray photoelectron spectroscopy demonstrates reactions occur on PC surface, which makes PC surface more hydrophilic. Separation of PC and ABS waste plastics was conducted based on the flotation behavior of single plastic. At different temperatures, PC and ABS mixtures were separated efficiently through froth flotation with ammonia pretreatment for different time (13 min at 23 °C, 18 min at 18 °C and 30 min at 23 °C). For both PC and ABS, the purity and recovery is more than 95.31% and 95.35%, respectively; the purity of PC and ABS is up to 99.72% and 99.23%, respectively. PC and ABS mixtures with different particle sizes were separated effectively, implying that ammonia treatment possesses superior applicability.« less

Evaluation of three fully automated immunoassay systems for detection of IgA anti-beta 2-glycoprotein I antibodies.

PubMed

Pérez, D; Martínez-Flores, J A; Serrano, M; Lora, D; Paz-Artal, E; Morales, J M; Serrano, A

2016-10-01

In recent years, we have been witnessing increased clinical interest in the determination of IgA anti-beta 2-glycoprotein I (aB2GPI) antibodies as well as increased demand for this test. Some ELISA-based diagnostic systems for IgA aB2GPI antibodies detection are suboptimal to detect it. The aim of our study was to determine whether the diagnostic yield of modern detection systems based on automatic platforms to measure IgA aB2GPI is equivalent to that of the well-optimized ELISA-based assays. In total, 130 patients were analyzed for IgA aB2GPI by three fully automated immunoassays using an ELISA-based assay as reference. The three systems were also analyzed for IgG aB2GPI with 58 patients. System 1 was able to detect IgA aB2GPI with good sensitivity and kappa index (99% and 0.72, respectively). The other two systems had also poor sensitivity (20% and 15%) and kappa index (0.10 and 0.07), respectively. On the other hand, kappa index for IgG aB2GPI was >0.89 in the three systems. Some analytical methods to detect IgA aB2GPI are suboptimal as well as some ELISA-based diagnostic systems. It is important that the scientific community work to standardize analytical methods to determine IgA aB2GPI antibodies. © 2016 John Wiley & Sons Ltd.

Common Occurrence of Explosive Hydrogen Burning in Type II Supernovae

NASA Astrophysics Data System (ADS)

Liu, Nan; Stephan, Thomas; Boehnke, Patrick; Nittler, Larry R.; Meyer, Bradley S.; O’D. Alexander, Conel M.; Davis, Andrew M.; Trappitsch, Reto; Pellin, Michael J.

2018-03-01

We report Mo isotopic data for 16 15N-rich presolar SiC grains of type AB (14N/15N < solar, AB1) and their correlated Sr and Ba isotope ratios when available. Of the 16 AB1 grains, 8 show s-process Mo isotopic compositions, together with s-process Ba and/or Sr isotopic compositions. We found that a higher percentage of AB1 grains show anomalous isotopic compositions than that of AB2 grains (14N/15N > solar), thus providing further support to the division of the two AB subgroups recently proposed by Liu et al., who showed that AB1 grains most likely originated from Type II supernovae (SNe) with explosive H burning. Comparison of the Sr, Mo, and Ba isotopic compositions of the AB1 grains with SN model predictions indicates that the s-process isotopic compositions of AB1 grains resulted from neutron-capture processes occurring during the progenitor massive stars’ pre-SN evolution rather than from an explosive neutron-capture process. In addition, the observations of (1) explosive H burning occurring in the C-rich regions of the progenitor SNe of X grains as suggested by the isotopic compositions of X grains, and (2) explosive H burning occurring both at the bottom of the He/C zone and at the top of the He/N zone as suggested by model simulations, imply that explosive H burning is a common phenomenon in outer SN zones.

Common Occurrence of Explosive Hydrogen Burning in Type II Supernovae

DOE PAGES

Liu, Nan; Stephan, Thomas; Boehnke, Patrick; ...

2018-03-16

In this paper, we report Mo isotopic data for 16 15N-rich presolar SiC grains of type AB ( 14N/ 15N < solar, AB1) and their correlated Sr and Ba isotope ratios when available. Of the 16 AB1 grains, 8 show s-process Mo isotopic compositions, together with s-process Ba and/or Sr isotopic compositions. We found that a higher percentage of AB1 grains show anomalous isotopic compositions than that of AB2 grains ( 14N/ 15N > solar), thus providing further support to the division of the two AB subgroups recently proposed by Liu et al., who showed that AB1 grains most likelymore » originated from Type II supernovae (SNe) with explosive H burning. Comparison of the Sr, Mo, and Ba isotopic compositions of the AB1 grains with SN model predictions indicates that the s-process isotopic compositions of AB1 grains resulted from neutron-capture processes occurring during the progenitor massive stars' pre-SN evolution rather than from an explosive neutron-capture process. Finally, in addition, the observations of (1) explosive H burning occurring in the C-rich regions of the progenitor SNe of X grains as suggested by the isotopic compositions of X grains, and (2) explosive H burning occurring both at the bottom of the He/C zone and at the top of the He/N zone as suggested by model simulations, imply that explosive H burning is a common phenomenon in outer SN zones.« less

Production and characterization of monoclonal antibodies (mAbs) against human serum albumin (HSA) for the development of an immunoaffinity system with oriented anti-HSA mAbs as immobilized ligand.

PubMed

Rajak, Poonam; Vijayalakshmi, M A; Jayaprakash, N S

2013-05-05

Proteins present in human serum are of immense importance in the field of biomarker discovery. But, the presence of high-abundant proteins like albumin makes the analysis more challenging because of masking effect on low-abundant proteins. Therefore, removal of albumin using highly specific monoclonal antibodies (mAbs) can potentiate the discovery of low-abundant proteins. In the present study, mAbs against human serum albumin (HSA) were developed and integrated in to an immunoaffinity based system for specific removal of albumin from the serum. Hybridomas were obtained by fusion of Sp2/0 mouse myeloma cells with spleen cells from the mouse immunized with HSA. Five clones (AHSA1-5) producing mAbs specific to HSA were established and characterized by enzyme linked immunosorbent assay (ELISA) and immunoblotting for specificity, sensitivity and affinity in terms of antigen binding. The mAbs were able to bind to both native albumin as well as its glycated isoform. Reactivity of mAbs with different mammalian sera was tested. The affinity constant of the mAbs ranged from 10(8) to 10(9)M(-1). An approach based on oriented immobilization was followed to immobilize purified anti-HSA mAbs on hydrazine activated agarose gel and the dynamic binding capacity of the column was determined. Copyright © 2013 Elsevier B.V. All rights reserved.

A targeted complement-dependent strategy to improve the outcome of mAb therapy, and characterization in a murine model of metastatic cancer

PubMed Central

Elvington, Michelle; Huang, Yuxiang; Morgan, B. Paul; Qiao, Fei; van Rooijen, Nico; Atkinson, Carl

2012-01-01

Complement inhibitors expressed on tumor cells provide an evasion mechanism against mAb therapy and may modulate the development of an acquired antitumor immune response. Here we investigate a strategy to amplify mAb-targeted complement activation on a tumor cell, independent of a requirement to target and block complement inhibitor expression or function, which is difficult to achieve in vivo. We constructed a murine fusion protein, CR2Fc, and demonstrated that the protein targets to C3 activation products deposited on a tumor cell by a specific mAb, and amplifies mAb-dependent complement activation and tumor cell lysis in vitro. In syngeneic models of metastatic lymphoma (EL4) and melanoma (B16), CR2Fc significantly enhanced the outcome of mAb therapy. Subsequent studies using the EL4 model with various genetically modified mice and macrophage-depleted mice revealed that CR2Fc enhanced the therapeutic effect of mAb therapy via both macrophage-dependent FcγR-mediated antibody-dependent cellular cytotoxicity, and by direct complement-mediated lysis. Complement activation products can also modulate adaptive immunity, but we found no evidence that either mAb or CR2Fc treatment had any effect on an antitumor humoral or cellular immune response. CR2Fc represents a potential adjuvant treatment to increase the effectiveness of mAb therapy of cancer. PMID:22442351

Single domain antibody-quantum dot conjugates for ricin detection by both fluoroimmunoassay and surface plasmon resonance.

PubMed

Anderson, George P; Glaven, Richard H; Algar, W Russ; Susumu, Kimihiro; Stewart, Michael H; Medintz, Igor L; Goldman, Ellen R

2013-07-05

The combination of stable biorecognition elements and robust quantum dots (QDs) has the potential to yield highly effective reporters for bioanalyses. Llama-derived single domain antibodies (sdAb) provide small thermostable recognition elements that can be easily manipulated using standard DNA methods. The sdAb was self-assembled on dihydrolipoic acid (DHLA) ligand-capped CdSe-ZnS core-shell QDs made in our laboratory through the polyhistidine tail of the protein, which coordinated to zinc ions on the QD surface. The sdAb-QD bioconjugates were then applied in both fluorometric and surface plasmon resonance (SPR) immunoassays for the detection of ricin, a potential biothreat agent. The sdAb-QD conjugates functioned in fluoroimmunoassays for the detection of ricin, providing equivalent limits of detection when compared to the same anti-ricin sdAb labeled with a conventional fluorophore. In addition, the DHLA-QD-sdAb conjugates were very effective reporter elements in SPR sandwich assays, providing more sensitive detection with a signal enhancement of ~10-fold over sdAb reporters and 2-4 fold over full sized antibody reporters. Commercially prepared streptavidin-modified polymer-coated QDs also amplified the SPR signal for the detection of ricin when applied to locations where biotinylated anti-ricin sdAb was bound to target; however, we observed a 4-fold greater amplification when using the DHLA-QD-sdAb conjugates in this format. Copyright © 2013. Published by Elsevier B.V.

Prospective iterative trial of proteasome inhibitor-based desensitization.

PubMed

Woodle, E S; Shields, A R; Ejaz, N S; Sadaka, B; Girnita, A; Walsh, R C; Alloway, R R; Brailey, P; Cardi, M A; Abu Jawdeh, B G; Roy-Chaudhury, P; Govil, A; Mogilishetty, G

2015-01-01

A prospective iterative trial of proteasome inhibitor (PI)-based therapy for reducing HLA antibody (Ab) levels was conducted in five phases differing in bortezomib dosing density and plasmapheresis timing. Phases included 1 or 2 bortezomib cycles (1.3 mg/m(2) × 6-8 doses), one rituximab dose and plasmapheresis. HLA Abs were measured by solid phase and flow cytometry (FCM) assays. Immunodominant Ab (iAb) was defined as highest HLA Ab level. Forty-four patients received 52 desensitization courses (7 patients enrolled in multiple phases): Phase 1 (n = 20), Phase 2 (n = 12), Phase 3 (n = 10), Phase 4 (n = 5), Phase 5 (n = 5). iAb reductions were observed in 38 of 44 (86%) patients and persisted up to 10 months. In Phase 1, a 51.5% iAb reduction was observed at 28 days with bortezomib alone. iAb reductions increased with higher bortezomib dosing densities and included class I, II, and public antigens (HLA DRβ3, HLA DRβ4 and HLA DRβ5). FCM median channel shifts decreased in 11/11 (100%) patients by a mean of 103 ± 54 mean channel shifts (log scale). Nineteen out of 44 patients (43.2%) were transplanted with low acute rejection rates (18.8%) and de novo DSA formation (12.5%). In conclusion, PI-based desensitization consistently and durably reduces HLA Ab levels providing an alternative to intravenous immune globulin-based desensitization. © Copyright 2014 The American Society of Transplantation and the American Society of Transplant Surgeons.

Common Occurrence of Explosive Hydrogen Burning in Type II Supernovae

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

Liu, Nan; Stephan, Thomas; Boehnke, Patrick

In this paper, we report Mo isotopic data for 16 15N-rich presolar SiC grains of type AB ( 14N/ 15N < solar, AB1) and their correlated Sr and Ba isotope ratios when available. Of the 16 AB1 grains, 8 show s-process Mo isotopic compositions, together with s-process Ba and/or Sr isotopic compositions. We found that a higher percentage of AB1 grains show anomalous isotopic compositions than that of AB2 grains ( 14N/ 15N > solar), thus providing further support to the division of the two AB subgroups recently proposed by Liu et al., who showed that AB1 grains most likelymore » originated from Type II supernovae (SNe) with explosive H burning. Comparison of the Sr, Mo, and Ba isotopic compositions of the AB1 grains with SN model predictions indicates that the s-process isotopic compositions of AB1 grains resulted from neutron-capture processes occurring during the progenitor massive stars' pre-SN evolution rather than from an explosive neutron-capture process. Finally, in addition, the observations of (1) explosive H burning occurring in the C-rich regions of the progenitor SNe of X grains as suggested by the isotopic compositions of X grains, and (2) explosive H burning occurring both at the bottom of the He/C zone and at the top of the He/N zone as suggested by model simulations, imply that explosive H burning is a common phenomenon in outer SN zones.« less

Treatment of psoriasis with interleukin-12/23 monoclonal antibody: a systematic review.

PubMed

Wu, Yan; Chen, Jing; Li, Yuan-Hong; Ma, Guo-Zhang; Chen, John Z S; Gao, Xing-Hua; Chen, Hong-Duo

2012-01-01

To systematically review the efficacy and safety of interleukin-12/23 monoclonal antibody (IL-12/23 mAb) on psoriasis. Relevant randomized controlled trials (RCTs) were identified by systematic literature searches in MEDLINE, OVID, EMBASE, Cochrane Library, and the metaRegister of Controlled Trials. The efficacy outcomes and adverse effects of included RCTs were critically assessed. A total of 3365 participants in 5 multicenter RCTs were included. The RRs of most efficacy outcomes showed significant differences between i) IL-12/23 mAb and placebo at week 12/16; ii) IL-12/23 mAb and etanercept at week 12; iii) IL-12/23 mAb in high dose and IL-12/23 mAb in low dose at week 24/28. Increasing treatment times did not obviously provide additional benefit to efficacy improvement. The adverse events of IL-12/23 mAb were similar to those of controls. Antibodies to IL-12/23 mAb were mostly undetected or shown at low titer. Treatment with IL-12/23 mAb did not influence related biochemical markers. IL-12/23 mAb was effective in the treatment of psoriasis on skin lesions, health-related quality of life and psoriatic arthritis in the short-term. The increase in treatment time points was not associated with additional efficacy and dose-dependence was observed with the ongoing treatment up to week 24/28. The adverse effects were minimal and tolerable.

A genetic variant of NLRP1 gene is associated with asbestos body burden in patients with malignant pleural mesothelioma.

PubMed

Crovella, S; Moura, R R; Cappellani, S; Celsi, F; Trevisan, E; Schneider, M; Brollo, A; Nicastro, E M; Vita, F; Finotto, L; Zabucchi, G; Borelli, V

2018-01-01

The presence of asbestos bodies (ABs) in lung parenchyma is considered a histopathologic hallmark of past exposure to asbestos fibers, of which there was a population of longer fibers. The mechanisms underlying AB formation are complex, involving inflammatory responses and iron (Fe) metabolism. Thus, the responsiveness to AB formation is variable, with some individuals appearing to be poor AB formers. The aim of this study was to disclose the possible role of genetic variants of genes encoding inflammasome and iron metabolism proteins in the ability to form ABs in a population of 81 individuals from North East Italy, who died after having developed malignant pleural mesothelioma (MPM). This study included 86 genetic variants distributed in 10 genes involved in Fe metabolism and 7 genetic variants in two genes encoding for inflammasome molecules. Genotypes/haplotypes were compared according to the number of lung ABs. Data showed that the NLRP1 rs12150220 missense variant (H155L) was significantly correlated with numbers of ABs in MPM patients. Specifically, a low number of ABs was detected in individuals carrying the NLRP1 rs12150220 A/T genotype. Our findings suggest that the NLRP1 inflammasome might contribute in the development of lung ABs. It is postulated that the NLRP1 missense variant may be considered as one of the possible host genetic factors contributing to individual variability in coating efficiency, which needs to be taken when assessing occupational exposure to asbestos.

An Anti-C1s Monoclonal, TNT003, Inhibits Complement Activation Induced by Antibodies Against HLA.

PubMed

Thomas, K A; Valenzuela, N M; Gjertson, D; Mulder, A; Fishbein, M C; Parry, G C; Panicker, S; Reed, E F

2015-08-01

Antibody-mediated rejection (AMR) of solid organ transplants (SOT) is characterized by damage triggered by donor-specific antibodies (DSA) binding donor Class I and II HLA (HLA-I and HLA-II) expressed on endothelial cells. While F(ab')2 portions of DSA cause cellular activation and proliferation, Fc regions activate the classical complement cascade, resulting in complement deposition and leukocyte recruitment, both hallmark features of AMR. We characterized the ability of an anti-C1s monoclonal antibody, TNT003, to inhibit HLA antibody (HLA-Ab)-induced complement activation. Complement deposition induced by HLA-Ab was evaluated using novel cell- and bead-based assays. Human aortic endothelial cells (HAEC) were cultured with HLA-Ab and human complement; production of activated complement proteins was measured by flow cytometry. Additionally, C3d deposition was measured on single antigen beads (SAB) mixed with HLA-Ab and human complement. TNT003 inhibited HLA-Ab mediated complement deposition on HAEC in a concentration-dependent manner; C3a, C4a and C5a anaphylatoxin production was also diminished by TNT003. Finally, TNT003 blocked C3d deposition induced by Class I (HLAI-Ab)- and Class II (HLAII-Ab)-specific antibodies on SAB. These data suggest TNT003 may be useful for modulating the effects of DSA, as TNT003 inhibits complement deposition and split product formation generated by HLA-I/II-Ab in vitro. © 2015 The Authors. American Journal of Transplantation Published by Wiley Periodicals, Inc.

Properties of blocking and non-blocking monoclonal antibodies specific for human macrophage galactose-type C-type lectin (MGL/ClecSF10A/CD301).

PubMed

Sano, Yoshihiko; Usami, Katsuaki; Izawa, Ryota; Denda-Nagai, Kaori; Higashi, Nobuaki; Kimura, Toshifumi; Suzuki, Noriko; Irimura, Tatsuro

2007-01-01

Monoclonal antibodies (mAbs) specific for the human macrophage galactose-type calcium-type lectin (MGL) were established. The recombinant extracellular domain of MGL was used to immunize a mouse, and 10 hybridoma clones were obtained. Binding of recombinant MGL to asialo-bovine submaxillary mucin was shown to be blocked by mAbs MLD-1, 4 and 6. Immunoprecipitation of MGL from lysates of COS-1 cells transfected with MGL cDNA (form 6A) was achieved with mAbs MLD-1, 4, 7, 8 and 16. Chimeric recombinant proteins between human MGL and mouse MGL1 were used to determine the location of the epitopes for these mAbs. mAbs MLD-8, 13, 15 and 16 interacted with the amino terminal side of the conserved WVDGTD sequence immediately upstream of QPD, whereas mAbs MLD-7, 12 and 17 interacted with the other side. mAbs MLD-1, 4, and 6 apparently required both sides of this boundary. mAbs MLD-15 and 16 were shown to recognize the protein products of alternatively spliced mRNA 6A/8A and 6C/8A, having deletions at the boundary of exons 7 and 8, in addition to full length and other spliced forms of MGL (6A, 6B and 6C), whereas the other mAbs bound only full length and forms 6A, 6B and 6C.

Ab interno trabeculectomy: ultrastructural evidence and early tissue response in a human eye.

PubMed

Ferrari, Ettore; Ortolani, Fulvia; Petrelli, Lucia; Contin, Magali; Pognuz, Derri Roman; Marchini, Maurizio; Bandello, Francesco

2007-10-01

To report the results of ultrastructural analysis of the postoperative effects of ab interno trabeculectomy in a human eye. Department of Ophthalmology, Palmanova Hospital, Palmanova, Udine, Italy. A 60-year-old woman with cataract and glaucoma had enucleation for a choroidal melanoma 10 days after ab interno trabeculectomy combined with phacoemulsification. A second ab interno trabeculectomy was performed after enucleation to evaluate the outcomes of the previous trabeculectomy. Light and transmission electron microscopy analyses were performed on samples excised from areas (1) not subjected to a procedure (control samples), (2) that had ab interno trabeculectomy before enucleation, and (3) that had ab interno trabeculectomy immediately after enucleation. Control samples showed normal trabecular features. Semithin sections of all ab interno trabeculectomy samples showed full-thickness removal of trabeculum segments, with Schlemm's canal lumen opening into the anterior chamber and apparent preservation of the adjacent structures. On ultrathin sections of samples that had ab interno trabeculectomy before enucleation, the endothelium lining the outer wall of Schlemm's canal and other angle components showed intact ultrastructural features. In trabecular beams that were not removed, the extracellular matrix appeared to have maintained its fine texture and was free of activated fibroblasts or leucocyte infiltrates. Observations confirm that ab interno trabeculectomy causes direct communication between Schlemm's canal lumen and the anterior chamber in vivo and immediately after enucleation during the early postoperative period. The absence of an evident inflammatory reaction in the examined case should be considered with caution because of possible tumor-induced immune suppression.

Diode laser surgery. Ab interno and ab externo versus conventional surgery in rabbits.

PubMed

Karp, C L; Higginbotham, E J; Edward, D P; Musch, D C

1993-10-01

Fibroblastic proliferation of subconjunctival tissues remains a primary mechanism of failure in filtration surgery. Minimizing the surgical manipulation of episcleral tissues may reduce scarring. Laser sclerostomy surgery involves minimal tissue dissection, and is gaining attention as a method of potentially improving filter duration in high-risk cases. Twenty-five New Zealand rabbits underwent filtration surgery in one eye, and the fellow eye remained as the unoperated control. Ten rabbits underwent ab externo diode laser sclerostomy surgery, ten underwent ab interno diode sclerostomy surgery, and five had posterior sclerostomy procedures. Filtration failure was defined as a less-than-4-mmHg intraocular pressure (IOP) difference between the operative and control eyes. The mean time to failure for the ab externo, ab interno, and conventional posterior sclerostomy techniques measured 17.4 +/- 11.5, 13.1 +/- 6.7, and 6.0 +/- 3.1 days, respectively. In a comparison of the laser-treated groups with the conventional procedure, the time to failure was significantly longer (P = 0.02) for the ab externo filter. The mean ab interno sclerostomy duration was longer than the posterior lip procedure, but this difference was not statistically significant (P = 0.15). The overall level of IOP reduction was similar in the three groups. These data suggest that diode laser sclerostomy is a feasible technique in rabbits, and the ab externo approach resulted in longer filter duration than the conventional posterior lip procedure in this model.

[VDRL and FTA-ABS reactivity in cerebrospinal fluid: our experience].

PubMed

García-Rodríguez, J A; Martín-Sánchez, A M; Canut, A; García-García, L; Cacho, J

1990-01-01

The reactivity of 194 samples of CSF against VDRL and FTA-ABS was studied in patients attending the Clinical Hospital in Salamanca over a five years period. This laboratory was asked to rule out an etiology of syphilis. Twelve samples of CSF proved to be reactive (6.2%) against VDRL and/or FTA-ABS. Seven of these corresponded to six adults diagnosed as suffering from neurosyphilis and one to an infant with early congenital syphilis without neurological alterations; these had in common the presence of active syphilis and a reactive FTA-ABS in serum. In the CSF of the six cases of neurosyphilis, VDRL was reactive in two patients (33.3%) and FTA-ABS in five (83.3%). One minimally reactive VDRL and four FTA-ABS were detected in the remaining five patients, with no known previous history of syphilis, that were suffering from different neurological alterations and that had a nonreactive FTA-ABS in serum. The results obtained in this study point to inappropriate use in CSF of VDRL and FTA-ABS to exclude neurosyphilis in our hospital since only 3.6% of the CSF studied corresponded to patients diagnosed as suffering from neurosyphilis and also to the need for improving the criteria for patient selection.

Overexpression of the ABC transporter AvtAB increases avermectin production in Streptomyces avermitilis.

PubMed

Qiu, Jingfan; Zhuo, Ying; Zhu, Dongqing; Zhou, Xiufen; Zhang, Lixin; Bai, Linquan; Deng, Zixin

2011-10-01

Avermectins are 16-membered macrocyclic polyketides with potent antiparasitic activities, produced by Streptomyces avermitilis. Upstream of the avermectin biosynthetic gene cluster, there is the avtAB operon encoding the ABC transporter AvtAB, which is highly homologous to the mammalian multidrug efflux pump P-glycoprotein (Pgp). Inactivation of avtAB had no effect, but increasing the concentration of avtAB mRNA 30-500-fold, using a multi-copy plasmid in S. avermitilis, enhanced avermectin production about two-fold both in the wild-type and in a high-yield producer strain on agar plates. In liquid industrial fermentation medium, the overall productivity of avermectin B1a in the engineered high-yield producer was improved for about 50%, from 3.3 to 4.8 g/l. In liquid YMG medium, moreover, the ratio of intracellular to extracellular accumulation of avermectin B1a was dropped from 6:1 to 4.5:1 in response to multiple copies of avtAB. Additionally, the overexpression of avtAB did not cause any increased expression of the avermectin biosynthetic genes through RT-PCR analysis. We propose that the AvtAB transporter exports avermectin, and thus reduces the feedback inhibition on avermectin production inside the cell. This strategy may be useful for enhancing the production of other antibiotics.

Stacking stability of MoS2 bilayer: An ab initio study

NASA Astrophysics Data System (ADS)

Tao, Peng; Guo, Huai-Hong; Yang, Teng; Zhang, Zhi-Dong

2014-10-01

The study of the stacking stability of bilayer MoS2 is essential since a bilayer has exhibited advantages over single layer MoS2 in many aspects for nanoelectronic applications. We explored the relative stability, optimal sliding path between different stacking orders of bilayer MoS2, and (especially) the effect of inter-layer stress, by combining first-principles density functional total energy calculations and the climbing-image nudge-elastic-band (CI-NEB) method. Among five typical stacking orders, which can be categorized into two kinds (I: AA, AB and II: AA', AB', A'B), we found that stacking orders with Mo and S superposing from both layers, such as AA' and AB, is more stable than the others. With smaller computational efforts than potential energy profile searching, we can study the effect of inter-layer stress on the stacking stability. Under isobaric condition, the sliding barrier increases by a few eV/(ucGPa) from AA' to AB', compared to 0.1 eV/(ucGPa) from AB to [AB]. Moreover, we found that interlayer compressive stress can help enhance the transport properties of AA'. This study can help understand why inter-layer stress by dielectric gating materials can be an effective means to improving MoS2 on nanoelectronic applications.

Evidence for three-dimensional XY critical properties in underdoped YBa2Cu3O7-δ

NASA Astrophysics Data System (ADS)

Schneider, T.

2007-05-01

We perform a detailed analysis of the reversible magnetization data of Salem-Sugui and Babíc of underdoped and optimally doped YBa2Cu3O7-δ single crystals. Near the zero field transition temperature we observe extended consistency with the properties of the three-dimensional XY universality class, even though the attained critical regime is limited by an inhomogeneity induced finite size effect. Nevertheless, as Tc falls from 93.5to41.5K , the critical amplitude of the in-plane correlation length ξab0 , the anisotropy γ=ξab0/ξc0 and the critical amplitude of the in-plane penetration depth λab0 increase substantially, while the critical amplitude of the c -axis correlation length ξc0 does not change much. As a consequence, the correlation volume Vcorr- increases and the critical amplitude of the specific heat singularity A- decreases dramatically, while the rise of λab0 reflects the behavior of the zero temperature counterpart. Conversely, although ξab0 and λab0 increase with reduced Tc , the ratio λab0/ξab0- , corresponding to the Ginzburg-Landau parameter κab , decreases substantially and YBa2Cu3O7-δ crosses over from an extreme to a weak type-II superconductor.

A novel blocking monoclonal antibody recognizing a distinct epitope of human CD40 molecule.

PubMed

Zhuang, Y; Huang, J; Zhou, Z; Ge, Y; Fan, Y; Qi, C; Zhen, L; Monchatre, E; Edelman, L; Zhang, X

2005-01-01

CD40, a member of the tumor necrosis factor receptor superfamily, is an important costimulatory molecule during the immune response. Here, we report a blocking mouse antihuman CD40 monoclonal antibody, mAb 3G3, of which the specificity was verified by flow cytometry and Western blot. It was shown by competition test that 3G3 bound to a different site (epitope) of CD40 from the reported CD40 mAbs, including clone mAb89, 3B2, and 5C11. It was also found that mAb 3G3 could inhibit homotypic aggregation of Daudi cells induced by the agonistic anti-CD40 mAb 5C11. Furthermore, mAb 3G3 effectively inhibited the proliferation of peripheral blood mononuclear cells in mixed lymphocyte reaction assay. Finally, a sensitive and specific soluble CD40 (sCD40) ELISA kit was established by matching mAb 3G3 with 5C11, and it was found that the levels of sCD40 in sera from patients with immune disorders such as hyperthyroidism, chronic nephritis, and rheumatoid arthritis were obviously higher than those from normal individuals. Thus, this blocking anti-CD40 mAb provides a novel tool for the study of CD40.

Antibody therapeutics for treating prostate cancer: where are we now and what comes next?

PubMed

Vlachostergios, Panagiotis J; Galletti, Giuseppe; Palmer, Jessica; Lam, Linda; Karir, Beerinder S; Tagawa, Scott T

2017-02-01

Progress in the understanding of molecular events of carcinogenesis and cancer evolution as well as the identification of tumor antigens has led to the development of different targeted therapeutic approaches, including the use of monoclonal antibodies (mAbs). Prostate cancer (PC) is highly amenable to mAb targeting given the existence of prostate-specific targets and the natural history and localization of metastatic disease. Areas covered: Several aspects of the PC phenotype, including growth factors, angiogenesis mediators, bone microenvironment signals, and immune evasion pathways, have become areas of ongoing investigation in terms of mAb targeting. These are reviewed. The greatest success so far has been the development of mAbs against prostate-specific tumor antigen (PSMA), which opened an opportunity to improve diagnostic accuracy and simultaneously target metastatic disease. Expert opinion: As mAb use in PC continues to evolve, more accurate imaging of the extent of disease and more effective mAb therapies (naked or conjugated with drugs, toxins or radioactive molecules) are emerging. In addition, the combination of mAbs with other treatment modalities is expected to further improve responses and overall survival. Identification of validated biomarkers is necessary for better recognition of patient subgroups who will derive the greatest benefit from mAb therapy.

J-type Carbon Stars: A Dominant Source of 14 N-rich Presolar SiC Grains of Type AB

DOE PAGES

Liu, Nan; Stephan, Thomas; Boehnke, Patrick; ...

2017-07-21

Here, we report Mo isotopic data of 27 new presolar SiC grains, including 12 14N-rich AB ( 14N/ 15N > 440, AB2) and 15 mainstream (MS) grains, and their correlated Sr and Ba isotope ratios when available. Direct comparison of the data for the MS grains, which came from low-mass asymptotic giant branch (AGB) stars with large s-process isotope enhancements, with the AB2 grain data demonstrates that AB2 grains show near-solar isotopic compositions and lack s-process enhancements. The near-normal Sr, Mo, and Ba isotopic compositions of AB2 grains clearly exclude born-again AGB stars, where the intermediate neutron-capture process (i-process) takesmore » place, as their stellar source. On the other hand, low-mass CO novae and early R- and J-type carbon stars show 13C and 14N excesses but no s-process enhancements and are thus potential stellar sources of AB2 grains. And because both early R-type carbon stars and CO novae are rare objects, the abundant J-type carbon stars (10%–15% of all carbon stars) are thus likely to be a dominant source of AB2 grains.« less

Immunization of A4galt-deficient mice with glycosphingolipids from renal cell cancers resulted in the generation of anti-sulfoglycolipid monoclonal antibodies.

PubMed

Ando, Reiko; Tokuda, Noriyo; Yamamoto, Tokunori; Ikeda, Kazutaka; Hashimoto, Noboru; Taguchi, Ryo; Fan, Xiaoen; Furukawa, Keiko; Niimura, Yukio; Suzuki, Akemi; Goto, Momokazu; Furukawa, Koichi

2016-04-01

In this study, we immunized Gb3/CD77 synthase gene (A4galt) knockout (KO) mice with glycosphingolipids (GSLs) extracted from 3 renal cell cancer (RCC) cell lines to raise monoclonal antibodies (mAbs) reactive with globo-series GSLs specifically expressed in RCCs. Although a number of mAbs reactive with globo-series GSLs were generated, they reacted with both RCC cell lines and normal kidney cells. When we analyzed recognized antigens by mAbs that were specifically reactive with RCC, but not with normal kidney cells at least on the cell surface, many of them turned out to be reactive with sulfoglycolipids. Eight out of 11 RCC-specific mAbs were reactive with SM2 alone, and the other 3 mAbs were more broadly reactive with sulfated glycolipids, i.e. SM3 and SM4 as well as SM2. In the immunohistochemistry, these anti-sulfoglycolipids mAbs showed RCC-specific reaction, with no or minimal reaction with adjacent normal tissues. Thus, immunization of A4galt KO mice with RCC-derived GSLs resulted in the generation of anti sulfated GSL mAbs, and these mAbs may be applicable for the therapeutics for RCC patients.

Enhanced removal of bisphenol-AF by activated carbon-alginate beads with cetyltrimethyl ammonium bromide.

PubMed

Tang, Zheng; Peng, Sha; Hu, Shuya; Hong, Song

2017-06-01

Adsorption removal of bisphenol-AF (BPAF) from aqueous solutions by synthesized activated carbon-alginate beads (AC-AB) with cetyltrimethyl ammonium bromide (CTAB) has been studied using two ways. The traditional method (two-step) first synthesized CTAB-modified AC-AB (AC-AB-CTAB), then used it to remove BPAF by adsorption. And one-step method dispersed AC-AB and CTAB in wastewater, followed by the removal of BPAF accompanied with the synthesis of AC-AB-CTAB. The one-step method showed a better performance than the two-step method, achieving a maximum removal of BPAF with 284.6mg/g. Kinetic studies and adsorption isotherms indicated that adsorption process of BPAF on AC-AB by the one-step method could be expressed by a pseudo-second-order model and a Dubinin-Ashtakhov (D-A) isotherm, respectively. The effects of pH, ionic strength, and inorganic ions on BPAF adsorption were also investigated. Furthermore, hydrophobic interactions, hydrogen bonds, and π-π electron donor-acceptor (EDA) interactions were discussed to explain the enhanced adsorption behavior of BPAF on AC-AB with CTAB. The findings verified the effectiveness of AC-AB for the removal of BPAF from wastewater and its high stability within five regeneration cycles. Copyright © 2017 Elsevier Inc. All rights reserved.

Impact-induced fracture mechanisms of immiscible PC/ABS (50/50) blends

NASA Astrophysics Data System (ADS)

Machmud, M. N.; Omiya, M.; Inoue, H.; Kishimoto, K.

2018-03-01

This paper presents a study on fracture mechanisms of polycarbonate (PC)/acrylonitrile-butadiene-styrene (ABS) (50/50) blends with different ABS types under a drop weight impact test (DWIT) using a circular sheet specimen. Formation of secondary crack indicated by a stress-whitening layer on the mid-plane of scattered specimens and secondary surface of fracture perpendicular to primary fracture surface were captured under scanning electron microscope (SEM). Although the both blends finally failed in brittle modes, SEM observation showed that their secondary fracture mechanisms were completely different. Observation through the thickness of the etched PC/ABS specimen samples using SEM also clearly showed that PC and ABS were immiscible. The immiscibility between PC and ABS was indicated by presence of their layer structures through the thickness of the blends. It was revealed that layer of ABS structure was influenced by size of rubber particle and this latter parameter then affected microstructure and fracture mechanisms of the blends. Impact-induced fracture mechanisms of the blends due to such microstructures are discussed in this paper. It was also pointed out that the secondary cracking was likely caused by interface delamination between PC and ABS layers in the core due to transverse shear stress generated during the impact test.

J-type Carbon Stars: A Dominant Source of {sup 14}N-rich Presolar SiC Grains of Type AB

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

Liu, Nan; Nittler, Larry R.; Alexander, Conel M. O’D.

We report Mo isotopic data of 27 new presolar SiC grains, including 12 {sup 14}N-rich AB ({sup 14}N/{sup 15}N > 440, AB2) and 15 mainstream (MS) grains, and their correlated Sr and Ba isotope ratios when available. Direct comparison of the data for the MS grains, which came from low-mass asymptotic giant branch (AGB) stars with large s -process isotope enhancements, with the AB2 grain data demonstrates that AB2 grains show near-solar isotopic compositions and lack s -process enhancements. The near-normal Sr, Mo, and Ba isotopic compositions of AB2 grains clearly exclude born-again AGB stars, where the intermediate neutron-capture processmore » ( i -process) takes place, as their stellar source. On the other hand, low-mass CO novae and early R- and J-type carbon stars show {sup 13}C and {sup 14}N excesses but no s -process enhancements and are thus potential stellar sources of AB2 grains. Because both early R-type carbon stars and CO novae are rare objects, the abundant J-type carbon stars (10%–15% of all carbon stars) are thus likely to be a dominant source of AB2 grains.« less

J-type Carbon Stars: A Dominant Source of 14 N-rich Presolar SiC Grains of Type AB

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

Liu, Nan; Stephan, Thomas; Boehnke, Patrick

Here, we report Mo isotopic data of 27 new presolar SiC grains, including 12 14N-rich AB ( 14N/ 15N > 440, AB2) and 15 mainstream (MS) grains, and their correlated Sr and Ba isotope ratios when available. Direct comparison of the data for the MS grains, which came from low-mass asymptotic giant branch (AGB) stars with large s-process isotope enhancements, with the AB2 grain data demonstrates that AB2 grains show near-solar isotopic compositions and lack s-process enhancements. The near-normal Sr, Mo, and Ba isotopic compositions of AB2 grains clearly exclude born-again AGB stars, where the intermediate neutron-capture process (i-process) takesmore » place, as their stellar source. On the other hand, low-mass CO novae and early R- and J-type carbon stars show 13C and 14N excesses but no s-process enhancements and are thus potential stellar sources of AB2 grains. And because both early R-type carbon stars and CO novae are rare objects, the abundant J-type carbon stars (10%–15% of all carbon stars) are thus likely to be a dominant source of AB2 grains.« less

J-type Carbon Stars: A Dominant Source of 14 N-rich Presolar SiC Grains of Type AB

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

Liu, Nan; Stephan, Thomas; Boehnke, Patrick

We report Mo isotopic data of 27 new presolar SiC grains, including 12 N-14-rich AB (N-14/N-15 > 440, AB2) and 15 mainstream (MS) grains, and their correlated Sr and Ba isotope ratios when available. Direct comparison of the data for the MS grains, which came from low-mass asymptotic giant branch (AGB) stars with large s-process isotope enhancements, with the AB2 grain data demonstrates that AB2 grains show near-solar isotopic compositions and lack s-process enhancements. The near-normal Sr, Mo, and Ba isotopic compositions of AB2 grains clearly exclude born-again AGB stars, where the intermediate neutron-capture process (i-process) takes place, as theirmore » stellar source. On the other hand, low-mass CO novae and early R-and J-type carbon stars show C-13 and N-14 excesses but no s-process enhancements and are thus potential stellar sources of AB2 grains. Because both early R-type carbon stars and CO novae are rare objects, the abundant J-type carbon stars (10%-15% of all carbon stars) are thus likely to be a dominant source of AB2 grains.« less

Naturally selected hepatitis C virus polymorphisms confer broad neutralizing antibody resistance.

PubMed

Bailey, Justin R; Wasilewski, Lisa N; Snider, Anna E; El-Diwany, Ramy; Osburn, William O; Keck, Zhenyong; Foung, Steven K H; Ray, Stuart C

2015-01-01

For hepatitis C virus (HCV) and other highly variable viruses, broadly neutralizing mAbs are an important guide for vaccine development. The development of resistance to anti-HCV mAbs is poorly understood, in part due to a lack of neutralization testing against diverse, representative panels of HCV variants. Here, we developed a neutralization panel expressing diverse, naturally occurring HCV envelopes (E1E2s) and used this panel to characterize neutralizing breadth and resistance mechanisms of 18 previously described broadly neutralizing anti-HCV human mAbs. The observed mAb resistance could not be attributed to polymorphisms in E1E2 at known mAb-binding residues. Additionally, hierarchical clustering analysis of neutralization resistance patterns revealed relationships between mAbs that were not predicted by prior epitope mapping, identifying 3 distinct neutralization clusters. Using this clustering analysis and envelope sequence data, we identified polymorphisms in E2 that confer resistance to multiple broadly neutralizing mAbs. These polymorphisms, which are not at mAb contact residues, also conferred resistance to neutralization by plasma from HCV-infected subjects. Together, our method of neutralization clustering with sequence analysis reveals that polymorphisms at noncontact residues may be a major immune evasion mechanism for HCV, facilitating viral persistence and presenting a challenge for HCV vaccine development.

Thyroid Stimulating Hormone Receptor Antibodies in Thyroid Eye Disease-Methodology and Clinical Applications.

PubMed

Diana, Tanja; Kahaly, George J

2018-05-02

Thyroid stimulating hormone receptor antibodies (TSHR-Ab) cause autoimmune hyperthyroidism and are prevalent in patients with related thyroid eye disease (TED). To provide a historical perspective on TSHR-Ab and to present evidence-based recommendations for clinical contemporary use. The authors review the recent literature pertaining to TSHR-Ab in patients with TED and describe the various immunoassays currently used for detecting TSHR-Ab and their clinical applications. We provide a historical summary and description of the various methods used to detect TSHR-Ab, foremost, the functional TSHR-Ab. Increasing experimental and clinical data demonstrate the clinical usefulness of cell-based bioassays for measurements of functional TSHR-Ab in the diagnosis and management of patients with autoimmune TED and in the characterization of patients with autoimmune-induced hyperthyroidism and hypothyroidism. Thyroid stimulating hormone receptor antibodies, especially the functional stimulating antibodies, are sensitive, specific, and reproducible biomarkers for patients with autoimmune TED and correlate well with clinical disease activity and clinical severity. Unlike competitive-binding assays, bioassays have the advantage of indicating not only the presence of antibodies but also their functional activity and potency. Measurement of TSHR-Ab (especially stimulating antibodies) is a clinically useful tool for the management of patients with TED.

Vaccine Induction of Heterologous Tier 2 HIV-1 Neutralizing Antibodies in Animal Models.

PubMed

Saunders, Kevin O; Verkoczy, Laurent K; Jiang, Chuancang; Zhang, Jinsong; Parks, Robert; Chen, Haiyan; Housman, Max; Bouton-Verville, Hilary; Shen, Xiaoying; Trama, Ashley M; Scearce, Richard; Sutherland, Laura; Santra, Sampa; Newman, Amanda; Eaton, Amanda; Xu, Kai; Georgiev, Ivelin S; Joyce, M Gordon; Tomaras, Georgia D; Bonsignori, Mattia; Reed, Steven G; Salazar, Andres; Mascola, John R; Moody, M Anthony; Cain, Derek W; Centlivre, Mireille; Zurawski, Sandra; Zurawski, Gerard; Erickson, Harold P; Kwong, Peter D; Alam, S Munir; Levy, Yves; Montefiori, David C; Haynes, Barton F

2017-12-26

The events required for the induction of broad neutralizing antibodies (bnAbs) following HIV-1 envelope (Env) vaccination are unknown, and their induction in animal models as proof of concept would be critical. Here, we describe the induction of plasma antibodies capable of neutralizing heterologous primary (tier 2) HIV-1 strains in one macaque and two rabbits. Env immunogens were designed to induce CD4 binding site (CD4bs) bnAbs, but surprisingly, the macaque developed V1V2-glycan bnAbs. Env immunization of CD4bs bnAb heavy chain rearrangement (V H DJ H ) knockin mice similarly induced V1V2-glycan neutralizing antibodies (nAbs), wherein the human CD4bs V H  chains were replaced with mouse rearrangements bearing diversity region (D)-D fusions, creating antibodies with long, tyrosine-rich HCDR3s. Our results show that Env vaccination can elicit broad neutralization of tier 2 HIV-1, demonstrate that V1V2-glycan bnAbs are more readily induced than CD4bs bnAbs, and define V H replacement and diversity region fusion as potential mechanisms for generating V1V2-glycan bnAb site antibodies. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Antigen analyses of serotypes of streptococcus mutans using a monoclonal antibody elaborated against serotype g polysaccharide antigen.

PubMed

Okahashi, N; Nishida, Y; Futakami, K; Hamada, S

1985-04-01

A hybridoma (F4B) which produced a monoclonal antibody (mAb) specific for serotype g carbohydrate antigen (RRg) of Streptococcus mutans 6715 was obtained. The F4B mAb cross-reacted with purified carbohydrate antigens of serotype d (RRd) and serotype h (TCAh). In immunodiffusion tests, F4B mAb produced a stable precipitin band with RRg, while the band developed between the mAb and RRd/TCAh in the cold disappeared when incubated at room temperature. The immunoprecipitin reaction between F4B mAb and RRg was strongly inhibited upon addition of lactose.

Comparative study of thiophilic functionalised matrices for polyclonal F(ab')2 purification.

PubMed

Kumpalume, Peter; Slater, Nigel K H

2004-01-02

Thiophilic adsorbents have been developed using divinyl sulfone or epoxy activated Streamline quartz base matrix. Their capacity and selectivity for binding polyclonal F(ab')2 fragments generated by whole serum proteolysis was tested. Except for epoxy activated guanidine, all the adsorbents displayed high selectivity for F(ab')2 with dynamic binding capacities ranging from 3 to 10 mg/ml of adsorbent. Thiol immobilised ligands adsorbed more F(ab')2 and the recovery was equal to or more than that from amino immobilised ligands. All adsorbents showed good selectivity for IgG and the dynamic binding capacities were better than for F(ab')2.

Calorimetric investigation of the excess entropy of mixing in analbite-sanidine solid solutions: lack of evidence for Na,K short- range order and implications for two-feldspar thermometry.

USGS Publications Warehouse

Haselton, H.T.; Hovis, G.L.; Hemingway, B.S.; Robie, R.A.

1983-01-01

Heat capacities (5-380 K) have been measured by adiabatic calorimetry for five highly disordered alkali feldspars (Ab99Or1, Ab85Or15, Ab55Or45, Ab25Or75 and Ab1Or99). The thermodynamic and mineralogical implications of the results are discussed. The new data are also combined with recent data for plagioclases in order to derive a revised expression for the two-feldspar thermometer. T calculated from the revised expression tend to be higher than previous calculations.-J.A.Z.

Nickel-metal hydride (Ni-MH) technology evaluation for aircraft battery applications

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

Loeber, G.; Vukson, S.P.; Erbacher, J.K.

1996-12-31

Available cylindrical and prismatic commercial Ni-MH batteries using AB{sub 5} and AB{sub 2} cathodes were evaluated for possible application to military aircraft batteries. Commercial AB{sub 5} technology is further advanced than AB{sub 2} technology and would require less alloy, electrolyte and single cell/battery development for near term (3--5 years) applications. Tested AB{sub 2} technology appears inadequate to meet the near term military requirements and would require a major development in the alloy to overcome the irreversible capacity loss at temperatures above 49 C. In addition, significant advances in alloy, electrolyte and single cell/battery development would also be needed.

[Analysis of drug resistance of Acinetobacter baumannii in wound of children with traffic injury and its relationship with antibiotic use].

PubMed

Liu, S; Wang, C; Fu, Y X

2017-07-20

Objective: To know the drug resistance of Acinetobacter baumannii (AB) in wound of children with traffic injury and its relationship with antibiotic use. Methods: Wound exudate of 226 children with traffic injury admitted to our unit from January 2010 to December 2015 were collected. API bacteria identification panels and fully automatic microbiological identification system were used to identify pathogens. Kirby-Bauer paper disk diffusion method was used to detect the drug resistance of pathogens to 18 antibiotics including amoxycillin/clavulanic acid, piperacillin/tazobactam, and imipenem. The detection situation of pathogen of children's wounds and drug resistance of detected AB to 18 antibiotics in each year were collected. Forty-six AB positive children (2 children excluded) were divided into imipenem-resistant group (IR, n =19) and non imipenem-resistant group (NIR, n =25) according to whether AB was 100% resistant to imipenem. Drug resistance of AB in wounds of children to 18 antibiotics in two groups was compared. The antibiotic use of AB positive children was collected, and the antibiotic use intensity of children in two groups was compared. Data were processed with Fisher's exact test, independent sample t test, and corrected t test. Results: (1) The detection rates of pathogen in wounds of children in 2010-2015 were 95.6% (43/45), 89.8% (53/59), 81.3% (148/182), 81.1% (107/132), 81.6% (120/147), and 77.5% (62/80), respectively, showing a trend of decreasing year by year. A total of 665 strains and 75 pathogens were detected, and the top 5 pathogens with detection rate from high to low were AB, Pseudomonas aeruginosa, Enterobacter cloacae, Staphylococcus epidermidis, and Escherichia coli, respectively. (2) Drug resistance rates of AB to amoxycillin/clavulanic acid, cefazolin, aztreonam, and piperacillin were all 100%, while AB was 100% sensitive to polymyxin, and the total drug resistance rates of AB to the other 13 antibiotics were all above 50%. The drug resistance rate of AB in wounds of children to piperacillin was higher than that to piperacillin/tazobactam in 2010-2015. (3) Except for imipenem, amoxycillin/clavulanic acid, cefazolin, aztreonam, piperacillin, and polymyxin, the drug resistance rates of AB in wounds of children in group IR to the other 12 antibiotics were higher than those in group NIR (with P values below 0.01). Besides, AB strains in wounds of children in group IR were completely resistant to at least 3 kinds of antibiotics including carbapenems, aminoglycosides, and quinolones, so that they were multidrug-resistant AB. (4) A total of 32 antibiotics were used in 46 AB positive children, and the 10-top-used antibiotics with use intensity from high to low were cefoperazone/sulbactam, piperacillin/tazobactam, cefazolin, imipenem, ceftizoxime, amoxycillin/clavulanate, ceftazidime, cefepime, amoxycillin/sulbactam, and cefmetazole, respectively. (5) Twenty-one antibiotics were not included in the comparison because of their small amount of usage. For the other 11 antibiotics, only the use intensity of metronidazole of children in two groups was statistically different ( t =-3.104, P <0.05). There was no statistically significant difference in total antibiotic use of children in two groups ( t =0.368, P >0.05). Conclusions: AB is one of the main pathogens in wounds of children with traffic injury, with high drug resistant rate. The high intensity of antibiotic use may lead to its drug resistance. In this study, the top-used antibiotics were in accord with AB resistant drugs, indicating a lack of normative use of antibiotics.

40 CFR 89.126 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2010 CFR

2010-07-01

... certificate void ab initio. (d) When the Administrator denies, suspends, revokes, or voids ab initio a... such fraud or other misconduct that makes the certification invalid ab initio. [59 FR 31335, June 17...

40 CFR 89.126 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2014 CFR

2014-07-01

... certificate void ab initio. (d) When the Administrator denies, suspends, revokes, or voids ab initio a... such fraud or other misconduct that makes the certification invalid ab initio. [59 FR 31335, June 17...

40 CFR 89.126 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2012 CFR

2012-07-01

... certificate void ab initio. (d) When the Administrator denies, suspends, revokes, or voids ab initio a... such fraud or other misconduct that makes the certification invalid ab initio. [59 FR 31335, June 17...

40 CFR 89.126 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2013 CFR

2013-07-01

... certificate void ab initio. (d) When the Administrator denies, suspends, revokes, or voids ab initio a... such fraud or other misconduct that makes the certification invalid ab initio. [59 FR 31335, June 17...

40 CFR 89.126 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2011 CFR

2011-07-01

... certificate void ab initio. (d) When the Administrator denies, suspends, revokes, or voids ab initio a... such fraud or other misconduct that makes the certification invalid ab initio. [59 FR 31335, June 17...

Optimal immunization cocktails can promote induction of broadly neutralizing Abs against highly mutable pathogens.

PubMed

Shaffer, J Scott; Moore, Penny L; Kardar, Mehran; Chakraborty, Arup K

2016-10-24

Strategies to elicit Abs that can neutralize diverse strains of a highly mutable pathogen are likely to result in a potent vaccine. Broadly neutralizing Abs (bnAbs) against HIV have been isolated from patients, proving that the human immune system can evolve them. Using computer simulations and theory, we study immunization with diverse mixtures of variant antigens (Ags). Our results show that particular choices for the number of variant Ags and the mutational distances separating them maximize the probability of inducing bnAbs. The variant Ags represent potentially conflicting selection forces that can frustrate the Darwinian evolutionary process of affinity maturation. An intermediate level of frustration maximizes the chance of evolving bnAbs. A simple model makes vivid the origin of this principle of optimal frustration. Our results, combined with past studies, suggest that an appropriately chosen permutation of immunization with an optimally designed mixture (using the principles that we describe) and sequential immunization with variant Ags that are separated by relatively large mutational distances may best promote the evolution of bnAbs.

Unexpected effects of gene deletion on mercury interactions with the methylation-deficient mutant hgcAB

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

Lin, Hui; Hurt, Jr., Richard Ashley; Johs, Alexander

2014-01-01

The hgcA and hgcB gene pair is essential for mercury (Hg) methylation by certain anaerobic bacteria,1 but little is known about how deletion of hgcAB affects cell surface interactions and intracellular uptake of Hg. Here, we compare hgcAB mutants with the wild-type (WT) strains of both Geobacter sulfurreducens PCA and Desulfovibrio desulfuricans ND132 and observe differences in Hg redox transformations, adsorption, and uptake in laboratory incubation studies. In both strains, deletion of hgcAB increased the reduction of Hg(II) but decreased the oxidation of Hg(0) under anaerobic conditions. The measured cellular thiol content in hgcAB mutants was lower than the WT,more » accounting for decreased adsorption and uptake of Hg. Despite the lack of methylation activity, Hg uptake by the hgcAB continued, albeit at a slower rate than the WT. These findings demonstrate that deletion of the hgcAB gene not only eliminates Hg methylation but also alters cell physiology, resulting in changes to Hg redox reactions, sorption, and uptake by cells.« less

Detection, Size, Measurement, and Structural Analysis Limits for the 2MASS, UKIDSS-LAS, and VISTA VIKING Surveys

NASA Astrophysics Data System (ADS)

Andrews, Stephen K.; Kelvin, Lee S.; Driver, Simon P.; Robotham, Aaron S. G.

2014-01-01

The 2MASS, UKIDSS-LAS, and VISTA VIKING surveys have all now observed the GAMA 9hr region in the Ks band. Here we compare the detection rates, photometry, basic size measurements, and single-component GALFIT structural measurements for a sample of 37 591 galaxies. We explore the sensitivity limits where the data agree for a variety of issues including: detection, star-galaxy separation, photometric measurements, size and ellipticity measurements, and Sérsic measurements. We find that 2MASS fails to detect at least 20% of the galaxy population within all magnitude bins, however for those that are detected we find photometry is robust (± 0.2 mag) to 14.7 AB mag and star-galaxy separation to 14.8 AB mag. For UKIDSS-LAS we find incompleteness starts to enter at a flux limit of 18.9 AB mag, star-galaxy separation is robust to 16.3 AB mag, and structural measurements are robust to 17.7 AB mag. VISTA VIKING data are complete to approximately 20.0 AB mag and structural measurements appear robust to 18.8 AB mag.

Optimal immunization cocktails can promote induction of broadly neutralizing Abs against highly mutable pathogens

PubMed Central

Shaffer, J. Scott; Moore, Penny L.; Kardar, Mehran; Chakraborty, Arup K.

2016-01-01

Strategies to elicit Abs that can neutralize diverse strains of a highly mutable pathogen are likely to result in a potent vaccine. Broadly neutralizing Abs (bnAbs) against HIV have been isolated from patients, proving that the human immune system can evolve them. Using computer simulations and theory, we study immunization with diverse mixtures of variant antigens (Ags). Our results show that particular choices for the number of variant Ags and the mutational distances separating them maximize the probability of inducing bnAbs. The variant Ags represent potentially conflicting selection forces that can frustrate the Darwinian evolutionary process of affinity maturation. An intermediate level of frustration maximizes the chance of evolving bnAbs. A simple model makes vivid the origin of this principle of optimal frustration. Our results, combined with past studies, suggest that an appropriately chosen permutation of immunization with an optimally designed mixture (using the principles that we describe) and sequential immunization with variant Ags that are separated by relatively large mutational distances may best promote the evolution of bnAbs. PMID:27791170

A caged Ab reveals an immediate/instructive effect of BDNF during hippocampal synaptic potentiation

PubMed Central

Kossel, Albrecht H.; Cambridge, Sidney B.; Wagner, Uta; Bonhoeffer, Tobias

2001-01-01

Neurotrophins have been shown to be involved in functional strengthening of central nervous system synapses. Although their general importance in this process is undisputed, it remains unresolved whether neurotrophins are truly mediators of synaptic strengthening or merely important cofactors. To address this question, we have devised a method to inactivate endogenous brain-derived neurotrophic factor (BDNF) with high time resolution by “caging” a function-blocking mAb against BDNF with a photosensitive protecting compound. Different assays were used to show that this inactivation of the Ab is reversible by UV light. Synaptic potentiation after τ-burst stimulation in the CA1 region of acute hippocampal slices was significantly less when applying the unmodified Ab compared with the caged Ab. Importantly, photoactivation of the caged Ab during the time of induction of synaptic enhancement led to a marked decrease in potentiation. Our experiments therefore strengthen the view that endogenous BDNF has fast effects during induction of synaptic plasticity. The results additionally show that caged Abs can provide a tool for precise spatiotemporal control over endogenous protein levels. PMID:11724927

Immunoglobulin Gene Insertions and Deletions in the Affinity Maturation of HIV-1 Broadly Reactive Neutralizing Antibodies

PubMed Central

Kepler, Thomas B.; Liao, Hua-Xin; Alam, S. Munir; Bhaskarabhatla, Rekha; Zhang, Ruijun; Stewart, Shelley; Anasti, Kara; Kelsoe, Garnett; Parks, Robert; Lloyd, Krissey E.; Stolarchuk, Christina; Pritchett, Jamie; Solomon, Erika; Friberg, Emma; Morris, Lynn; Karim, Salim S. Abdool; Cohen, Myron S.; Walter, Emmanuel; Moody, M. Anthony; Wu, Xueling; Altae-Tran, Han R.; Georgiev, Ivelin S.; Kwong, Peter D.; Boyd, Scott D.; Fire, Andrew Z.; Mascola, John R.; Haynes, Barton F.

2014-01-01

Summary Induction of HIV-1 broad neutralizing antibodies (bnAbs) is a goal of HIV-1 vaccine development but has remained challenging partially due to unusual traits of bnAbs, including high somatic hypermutation (SHM) frequencies and in-frame insertions and deletions (indels). Here we examined the propensity and functional requirement for indels within HIV-1 bnAbs. High-throughput sequencing of the immunoglobulin (Ig) VHDJH genes in HIV-1 infected and uninfected individuals revealed that the indel frequency was elevated among HIV-1-infected subjects, with no unique properties attributable to bnAb-producing individuals. This increased indel occurrence depended only on the frequency of SHM point-mutations. Indel-encoded regions were generally proximal to antigen binding sites. Additionally, reconstruction of a HIV-1 CD4-binding site bnAb clonal lineage revealed that a large compound VHDJH indel was required for bnAb activity. Thus, vaccine development should focus on designing regimens targeted at sustained activation of bnAb lineages to achieve the required SHM and indel events. PMID:25211073

Beyond Simple AB Diblock Copolymers: Application of Bifunctional and Trifunctional RAFT Agents to PISA in Water.

PubMed

Mellot, Gaëlle; Beaunier, Patricia; Guigner, Jean-Michel; Bouteiller, Laurent; Rieger, Jutta; Stoffelbach, François

2018-06-20

The influence of the macromolecular reversible addition-fragmentation chain transfer (macro-RAFT) agent architecture on the morphology of the self-assemblies obtained by aqueous RAFT dispersion polymerization in polymerization-induced self-assembly (PISA) is studied by comparing amphiphilic AB diblock, (AB) 2 triblock, and triarm star-shaped (AB) 3 copolymers, constituted of N,N-dimethylacrylamide (DMAc = A) and diacetone acrylamide (DAAm = B). Symmetrical triarm (AB) 3 copolymers could be synthesized for the first time in a PISA process. Spheres and higher order morphologies, such as worms or vesicles, could be obtained for all types of architectures and the parameters that determine their formation have been studied. In particular, we found that the total DP n of the PDMAc and the PDAAm segments, i.e., the same overall molar mass, at the same M n (PDMAc)/M n (PDAAm) ratio, rather than the individual length of the arms determined the morphologies for the linear (AB) 2 and star shaped (AB) 3 copolymers obtained by using the bi- and trifunctional macro-RAFT agents. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Unique synergism in flame retardancy in ABS based composites through blending PVDF and halloysite nanotubes

NASA Astrophysics Data System (ADS)

Remanan, Sanjay; Sharma, Maya; Jayashree, Priyadarshini; Parameswaranpillai, Jyotishkumar; Fabian, Thomas; Shih, Julie; Shankarappa, Prasad; Nuggehalli, Bharath; Bose, Suryasarathi

2017-06-01

This study demonstrates flame retardant materials designed using bi-phasic polymer blends of acrylonitrile butadiene styrene (ABS) and polyvinylidene fluoride (PVDF) containing halloysite nanotubes (HNTs) and Cloisite 30B nanoclay. The prepared blends with and without nanoparticles were extensively characterized. The nanoparticles were added in different weight concentrations to improve the flame retardancy. It was observed that prepared ABS/PVDF blends showed better flame retardancy than ABS based composites. The flame resistance was further improved by the addition of nanoparticles in the blends. The microscale combustion calorimetry (MCC) test showed better flame resistance in ABS/PVDF blends filled with 5 wt% HNTs than other composites. The total heat release of ABS/PVDF blend filled with 5 wt% HNTs decreased by 31% and also the heat of combustion decreased by 26% as compared to neat ABS. When compared with nanoparticles, the addition of PVDF reduced the peak heat release rate (PHRR) and increased the char residue more effectively. A synergistic improvement was observed from both PVDF and HNTs on the flame resistance properties.

HIV Envelope Glycoform Heterogeneity and Localized Diversity Govern the Initiation and Maturation of a V2 Apex Broadly Neutralizing Antibody Lineage.

PubMed

Landais, Elise; Murrell, Ben; Briney, Bryan; Murrell, Sasha; Rantalainen, Kimmo; Berndsen, Zachary T; Ramos, Alejandra; Wickramasinghe, Lalinda; Smith, Melissa Laird; Eren, Kemal; de Val, Natalia; Wu, Mengyu; Cappelletti, Audrey; Umotoy, Jeffrey; Lie, Yolanda; Wrin, Terri; Algate, Paul; Chan-Hui, Po-Ying; Karita, Etienne; Ward, Andrew B; Wilson, Ian A; Burton, Dennis R; Smith, Davey; Pond, Sergei L Kosakovsky; Poignard, Pascal

2017-11-21

Understanding how broadly neutralizing antibodies (bnAbs) to HIV envelope (Env) develop during natural infection can help guide the rational design of an HIV vaccine. Here, we described a bnAb lineage targeting the Env V2 apex and the Ab-Env co-evolution that led to development of neutralization breadth. The lineage Abs bore an anionic heavy chain complementarity-determining region 3 (CDRH3) of 25 amino acids, among the shortest known for this class of Abs, and achieved breadth with only 10% nucleotide somatic hypermutation and no insertions or deletions. The data suggested a role for Env glycoform heterogeneity in the activation of the lineage germline B cell. Finally, we showed that localized diversity at key V2 epitope residues drove bnAb maturation toward breadth, mirroring the Env evolution pattern described for another donor who developed V2-apex targeting bnAbs. Overall, these findings suggest potential strategies for vaccine approaches based on germline-targeting and serial immunogen design. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Solvent (acetone-butanol: ab) production

USDA-ARS?s Scientific Manuscript database

This article describes production of butanol [acetone-butanol-ethanol, (also called AB or ABE or solvent)] by fermentation using both traditional and current technologies. AB production from agricultural commodities, such as corn and molasses, was an important historical fermentation. Unfortunately,...

Determining Binary Star Orbits Using Kepler's Equation

NASA Astrophysics Data System (ADS)

Boule, Cory; Andrews, Kaitlyn; Penfield, Andrew; Puckette, Ian; Goodale, Keith; Harfenist, Steven

2017-04-01

Students calculated ephemerides and generated orbits of four well-known binary systems. Using an iterative technique in Microsoft® Excel® to solve Kepler's equation, separation and position angle values were generated as well as plots of the apparent orbits. Current position angle and separation values were measured in the field and compared well to the calculated values for the stars: STF1196AB,C, STF296AB, STF296AB and STF60AB.

Surf Zone Currents. Volume I. State of Knowledge.

DTIC Science & Technology

1982-09-01

elevation above an arbitrary datum a angle between wave crest and bottom contour a angle between wave crest and the shoreline . ab angle between breaking...b- Note that neglecting wave setup, refraction and for small ab , equation (74) reduces to that employed by Longuet-Higgins (eq. 48). These researchers...28. As ab o (Note that ab = o means theory reduces to original order (zero order) solution given by Longuet-Higgins, 1970, the triangular solution is

Will studies in individuals with systemic lupus erythematosus be the key to future HIV vaccine design?

PubMed

Bonsignori, Mattia

2014-11-01

The induction of HIV-1 broadly neutralizing antibodies (bnAbs) remains the primary goal of a preventive HIV-1 vaccine but no HIV-1 vaccine candidate has succeeded in inducing bnAbs. All the bnAbs isolated from chronically HIV-1 infected subjects display one or more traits associated with control by host tolerance and immunoregulatory mechanisms, including reactivity against self antigens. Recent studies on a HIV-1 patient with concurrent systemic lupus erythematosus have informed on how similar bnAbs are to typical autoantibodies controlled by immune tolerance mechanisms. Future studies aimed at elucidating the intersection between autoantibodies generated in the context of systemic lupus erythematosus and the development of HIV-1 bnAbs will further our knowledge of specific roadblocks that hamper the production of bnAbs and, ultimately, inform us on how to implement vaccine strategies to circumvent them.

Mapping of cat albumin using monoclonal antibodies: identification of determinants common to cat and dog.

PubMed Central

Boutin, Y; Hébert, J; Vrancken, E R; Mourad, W

1989-01-01

Cat and dog albumins from commercial extracts were used to produce monoclonal antibodies (MoAb). Anti-cat albumin MoAb recognized both cat and dog albumin equally, as did anti-dog albumin MoAb; this confirms cross-reactivity between cat and dog. The MoAb were separated into two groups according to their epitopic specificity; they recognized two overlapping epitopes of cat albumin. Furthermore, by competitive inhibition of radio-allergosorbent test (RAST), it was shown that one MoAb group inhibited significantly the binding of human IgE antibodies (from a pool of 13 patients allergic to both cats and dogs) to insolubilized cat or dog extracts. These observations suggest that murine anti-cat or anti-dog MoAb and human IgE antibodies recognize identical or closely related determinants on cat and dog albumin. Images Fig. 1 Fig. 2 PMID:2478325

Ab-initio calculation of electronic structure and optical properties of AB-stacked bilayer α-graphyne

NASA Astrophysics Data System (ADS)

Behzad, Somayeh

2016-09-01

Monolayer α-graphyne is a new two-dimensional carbon allotrope with many special features. In this work the electronic properties of AA- and AB-stacked bilayers of this material and then the optical properties are studied, using first principle plane wave method. The electronic spectrum has two Dirac cones for AA stacked bilayer α-graphyne. For AB-stacked bilayer, the interlayer interaction changes the linear bands into parabolic bands. The optical spectra of the most stable AB-stacked bilayer closely resemble to that of the monolayer, except for small shifts of peak positions and increasing of their intensity. For AB-stacked bilayer, a pronounced peak has been found at low energies under the perpendicular polarization. This peak can be clearly ascribed to the transitions at the Dirac point as a result of the small degeneracy lift in the band structure.

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

Wheeler, T.M.; Johnson, E.H.; Coughlin, D.

While asbestos bodies (ABs) in sputum and/or bronchial washings are highly specific markers for significant asbestos exposure, comparison of the sensitivity between sputum cytology and bronchial washing cytology for the detection of ABs had not been documented. Review of the files of the Cytopathology Laboratory, Methodist Hospital, Houston, Texas, for the period 1973 to 1984 identified 11 patients with slides available for review who (1) had been examined by both sputum cytology and bronchial washing cytology and (2) had at least one specimen positive for ABs. Of the 11 evaluable cases, all had ABs in the bronchial washings but onymore » 6 had ABs in the sputum. In addition, iron stain (e.g., the Prussian blue stain) was found to be more sensitive than the Papanicolaou stain for the detection of ABs in these cases. These findings indicate that iron-stained bronchial washing specimens should be preferred for the cytologic detection of asbestos exposure.« less

FREE RADICALS IN THERMALLY STERILIZED ACIDUM BORICUM AND OPTIMIZATION OF THIS PROCESS.

PubMed

Ramos, Paweł; Pilawa, Barbara

2015-01-01

Free radicals formation in the acidum boricum (AB) during thermal sterilization process was examined by an X-band (9.3 GHz) electron paramagnetic resonance (EPR) spectroscopy. Acidum boricum was sterilized according to the pharmacopea norms at temperatures 160°C (120 min), 170°C (60 min), and 180°C (30 min). Free radicals (~10(17) spin/g) were thermally formed in these drug. The free radicals system revealed complex character, and the asymmetrical EPR spectra were measured. Mainly oxygen free radicals exist in the tested heated AB. Slower spin-lattice relaxation processes exist in AB sterilized at 160, 170 and 180°C. AB may be sterilized at temperatures 160, 170 and 180°C. For AB thermal sterilization at temperature 170°C is recommended. Free radicals concentrations changes during storage of the examined AB, and probably interactions with oxygen molecules may be responsible for this effect.

Development and application of ab initio QM/MM methods for mechanistic simulation of reactions in solution and in enzymes

PubMed Central

Hu, Hao; Yang, Weitao

2013-01-01

Determining the free energies and mechanisms of chemical reactions in solution and enzymes is a major challenge. For such complex reaction processes, combined quantum mechanics/molecular mechanics (QM/MM) method is the most effective simulation method to provide an accurate and efficient theoretical description of the molecular system. The computational costs of ab initio QM methods, however, have limited the application of ab initio QM/MM methods. Recent advances in ab initio QM/MM methods allowed the accurate simulation of the free energies for reactions in solution and in enzymes and thus paved the way for broader application of the ab initio QM/MM methods. We review here the theoretical developments and applications of the ab initio QM/MM methods, focusing on the determination of reaction path and the free energies of the reaction processes in solution and enzymes. PMID:24146439

Predicting the clinical efficacy and potential adverse effects of a humanized anticocaine monoclonal antibody

PubMed Central

Norman, Andrew B; Ball, William J

2012-01-01

The effects of a humanized monoclonal antibody (mAb) having high affinity and specificity for cocaine in animal models are reviewed. The mAb reduced the concentration of cocaine in the brain of mice after intravenous injection of cocaine. In addition, the mAb increased the concentration of cocaine required to reinstate cocaine self-administration. These effects may predict clinical efficacy of a passive immunotherapy for reducing the probability of cocaine-induced relapse. However, in the presence of the mAb, once cocaine self-administration was reinstated, the consumption rate of cocaine was increased. This effect is hypothesized to result from a pharmacokinetic/pharmacodynamic interaction. A humanized mAb should minimize adverse events related to the immunogenicity of the mAb protein, and the specificity for cocaine should avoid adverse events related to interactions with physiologically relevant endogenous proteins. PMID:22401638

Antibody Production in Plants and Green Algae.

PubMed

Yusibov, Vidadi; Kushnir, Natasha; Streatfield, Stephen J

2016-04-29

Monoclonal antibodies (mAbs) have a wide range of modern applications, including research, diagnostic, therapeutic, and industrial uses. Market demand for mAbs is high and continues to grow. Although mammalian systems, which currently dominate the biomanufacturing industry, produce effective and safe recombinant mAbs, they have a limited manufacturing capacity and high costs. Bacteria, yeast, and insect cell systems are highly scalable and cost effective but vary in their ability to produce appropriate posttranslationally modified mAbs. Plants and green algae are emerging as promising production platforms because of their time and cost efficiencies, scalability, lack of mammalian pathogens, and eukaryotic posttranslational protein modification machinery. So far, plant- and algae-derived mAbs have been produced predominantly as candidate therapeutics for infectious diseases and cancer. These candidates have been extensively evaluated in animal models, and some have shown efficacy in clinical trials. Here, we review ongoing efforts to advance the production of mAbs in plants and algae.

Combining ZnO/microwave treatment for changing wettability of WEEE styrene plastics (ABS and HIPS) and their selective separation by froth flotation

NASA Astrophysics Data System (ADS)

Thanh Truc, Nguyen Thi; Lee, Byeong-Kyu

2017-10-01

This study reports a simple froth flotation method to separate plastic wastes of acrylonitrile-butadiene-styrene (ABS) and high impact polystyrene (HIPS) after initial hydrophilization by coating the plastics with ZnO and microwave treatment. ABS and HIPS are typical styrene-based WEEE plastics having similar density and hydrophobicity, which hinders their separation for recycling. After coating with ZnO, 2-min microwave treatment rearranged the ABS surface and thus changed its molecular mobility and increased its hydrophilicity. The combined ZnO coating/microwave treatment facilitated the selective separation of ABS and HIPS with 100% and 95.2% recovery and 95.4% and 100% purity in froth flotation, respectively. The combination of ZnO coating-microwave treatment and froth flotation can be utilized as a selective ABS/HIPS separation technique for improved recycling of WEEE plastics.

(abstract) Studies on AB(sub 5) Metal Hydride Alloys with Sn Additives

NASA Technical Reports Server (NTRS)

Ratnakumar, B. V.; Surampudi, S.; Stefano, S. Di; Halpert, G.; Witham, C.; Fultz, B.

1994-01-01

The use of metal hydrides as negative electrodes in alkaline rechargeable cells is becoming increasingly popular, due to several advantages offered by the metal hydrides over conventional anode materials (such as Zn, Cd) in terms of specific energy environmental cycle life and compatibility. Besides, the similarities in the cell voltage pressure characteristics, and charge control methods of the Ni-MH cells to the commonly used Ni-Cd point to a projected take over of 25% of the Ni-Cd market for consumer electronics by the Ni-MH cells in the next couple of years. Two classes of metal hydrides alloys based on rare earth metals (AB(sub 5)) and titanium (AB(sub 2)) are being currently developed at various laboratories. AB(sub 2) alloys exhibit higher specific energy than the AB(sub 5) alloys but the state of the art commercial Ni-MH cells are predominately manufactured using AB(sub 5) alloys.

American foulbrood of the honey bee: occurrence and distribution of different genotypes of Paenibacillus larvae in the administrative district of Arnsberg (North Rhine-Westphalia).

PubMed

Peters, M; Kilwinski, J; Beringhoff, A; Reckling, D; Genersch, E

2006-03-01

Between March 2003 and October 2004, Paenibacillus larvae, the aetiological agent of American foulbrood disease of the honey bee, was isolated from broodcombs and honey samples of 54 apiaries in the administrative district of Arnsberg (North Rhine-Westphalia, Germany). Genotyping of 176 P. larvae isolates with repetitive element polymerase chain reaction fingerprinting (rep-PCR) using BOX A1R and MBO REP1 primers revealed five different genotypes (AB, Ab, ab, ass, Acapital BE, Cyrillic). In samples of three apiaries, more than one genotype was detected. A combination of two genotypes was isolated from honey samples of the same hive two times (ab/ass and Ab/ab). The five genotypes were not randomly distributed in the district, but revealed a certain geographical clustering. Possible factors with impact on the genotype diversity and the distribution pattern are discussed.

Halo-substituted azobenzenes adsorbed at Ag(111) and Au(111) interfaces: Structures and optical properties

NASA Astrophysics Data System (ADS)

Hughes, Zak E.; Baev, Alexander; Prasad, Paras N.; Walsh, Tiffany R.

2017-05-01

The adsorption of azobenzene (AB), ortho fluoro-azobenzene (FAB) and ortho chlor-azobenzol (ClAB), in both the cis and trans isomers, at the Au(111) and Ag(111) surfaces is investigated using plane-wave density functional calculations with the revPBE-vdW-DF functional. The resulting adsorption energies and internal structures of AB adsorbed to both metal surfaces are in broad agreement with available experimental data. In the gas phase, FAB and ClAB feature a significant reduction in the energy difference between the two isomeric states, compared with AB. This relative reduction in the energy difference is still significant for the adsorbed form of FAB but is only weakly apparent for ClAB. The absorption spectra of the molecules have also been calculated, with the halogen substituents generating significant changes in the gas phase, but only a modest difference for the adsorbed molecules.

Development of the Systems Thinking Scale for Adolescent Behavior Change.

PubMed

Moore, Shirley M; Komton, Vilailert; Adegbite-Adeniyi, Clara; Dolansky, Mary A; Hardin, Heather K; Borawski, Elaine A

2018-03-01

This report describes the development and psychometric testing of the Systems Thinking Scale for Adolescent Behavior Change (STS-AB). Following item development, initial assessments of understandability and stability of the STS-AB were conducted in a sample of nine adolescents enrolled in a weight management program. Exploratory factor analysis of the 16-item STS-AB and internal consistency assessments were then done with 359 adolescents enrolled in a weight management program. Test-retest reliability of the STS-AB was .71, p = .03; internal consistency reliability was .87. Factor analysis of the 16-item STS-AB indicated a one-factor solution with good factor loadings, ranging from .40 to .67. Evidence of construct validity was supported by significant correlations with established measures of variables associated with health behavior change. We provide beginning evidence of the reliability and validity of the STS-AB to measure systems thinking for health behavior change in young adolescents.

Development of the Systems Thinking Scale for Adolescent Behavior Change

PubMed Central

Moore, Shirley M.; Komton, Vilailert; Adegbite-Adeniyi, Clara; Dolansky, Mary A.; Hardin, Heather K.; Borawski, Elaine A.

2017-01-01

This report describes the development and psychometric testing of the Systems Thinking Scale for Adolescent Behavior Change (STS-AB). Following item development, initial assessments of understandability and stability of the STS-AB were conducted in a sample of nine adolescents enrolled in a weight management program. Exploratory factor analysis of the 16-item STS-AB and internal consistency assessments were then done with 359 adolescents enrolled in a weight management program. Test–retest reliability of the STS-AB was .71, p = .03; internal consistency reliability was .87. Factor analysis of the 16-item STS-AB indicated a one-factor solution with good factor loadings, ranging from .40 to .67. Evidence of construct validity was supported by significant correlations with established measures of variables associated with health behavior change. We provide beginning evidence of the reliability and validity of the STS-AB to measure systems thinking for health behavior change in young adolescents. PMID:28303755

A KINE-CHEMICAL INVESTIGATION OF THE AB DOR MOVING GROUP 'STREAM'

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

Barenfeld, Scott A.; Bubar, Eric J.; Mamajek, Eric E.

2013-03-20

The AB Dor Moving Group consists of a 'nucleus' of {approx}10 stars at d {approx_equal} 20 pc, along with dozens of purported 'stream' members distributed across the sky. We perform a chemical and kinematic analysis of a subsample of AB Dor stream stars to test whether they constitute a physical stellar group. We use the NEMO Galactic kinematic code to investigate the orbits of the stream members, and perform a chemical abundance analysis using high resolution spectra taken with the Magellan Clay 6.5 m telescope. Using a {chi}{sup 2} test with the measured abundances for 10 different elements, we findmore » that only half of the purported AB Dor stream members could possibly constitute a statistically chemically homogeneous sample. Some stream members with three-dimensional velocities were hundreds of parsecs from the AB Dor nucleus {approx}10{sup 8} yr ago, and hence were unlikely to share a common origin. We conclude that the published lists of AB Dor moving group stream members are unlikely to represent the dispersed remnant of a single star formation episode. A subsample of the stream stars appears to be both statistically chemically homogeneous and in the vicinity of the AB Dor nucleus at birth. Their mean metallicity is [Fe/H] = 0.02 {+-} 0.02 dex, which we consider representative for the AB Dor group. Finally, we report a strong lower limit on the age of the AB Dor nucleus of >110 Myr based on the pre-main sequence contraction times for K-type members which have reached the main sequence.« less

Higher cortisol levels at diurnal trough predict greater attentional bias towards threat in healthy young adults.

PubMed

Hakamata, Yuko; Izawa, Shuhei; Sato, Eisuke; Komi, Shotaro; Murayama, Norio; Moriguchi, Yoshiya; Hanakawa, Takashi; Inoue, Yusuke; Tagaya, Hirokuni

2013-11-01

Attentional bias (AB), selective information processing towards threat, can exacerbate anxiety and depression. Despite growing interest, physiological determinants of AB are yet to be understood. We examined whether stress hormone cortisol and its diurnal variation pattern contribute to AB. Eighty-seven healthy young adults underwent assessments for AB, anxious personality traits, depressive symptoms, and attentional function. Salivary cortisol was collected at three time points daily (at awakening, 30 min after awakening, and bedtime) for 2 consecutive days. We performed: (1) multiple regression analysis to examine the relationships between AB and the other measures and (2) analysis of variance (ANOVA) between groups with different cortisol variation patterns for the other measures. Multiple regression analysis revealed that higher cortisol levels at bedtime (p<0.001), an anxious personality trait (p=0.011), and years of education (p=0.036) were included in the optimal model to predict AB (adjusted R(2)=0.234, p<0.001). ANOVA further demonstrated significant mean differences in AB and depressive symptoms; individuals with blunted cortisol variation exhibited significantly greater AB and depression than those with moderate variation (p=0.037 and p=0.009, respectively). Neuropsychological assessment focused on attention and cortisol measurement at three time points daily. We showed that higher cortisol levels at bedtime and blunted cortisol variation are associated with greater AB. Individuals who have higher cortisol levels at diurnal trough might be at risk of clinical anxiety or depression but could also derive more benefits from the attentional-bias-modification program. © 2013 Elsevier B.V. All rights reserved.

A Kine-chemical Investigation of the AB Dor Moving Group "Stream"

NASA Astrophysics Data System (ADS)

Barenfeld, Scott A.; Bubar, Eric J.; Mamajek, Eric E.; Young, Patrick A.

2013-03-01

The AB Dor Moving Group consists of a "nucleus" of ~10 stars at d ~= 20 pc, along with dozens of purported "stream" members distributed across the sky. We perform a chemical and kinematic analysis of a subsample of AB Dor stream stars to test whether they constitute a physical stellar group. We use the NEMO Galactic kinematic code to investigate the orbits of the stream members, and perform a chemical abundance analysis using high resolution spectra taken with the Magellan Clay 6.5 m telescope. Using a χ2 test with the measured abundances for 10 different elements, we find that only half of the purported AB Dor stream members could possibly constitute a statistically chemically homogeneous sample. Some stream members with three-dimensional velocities were hundreds of parsecs from the AB Dor nucleus ~108 yr ago, and hence were unlikely to share a common origin. We conclude that the published lists of AB Dor moving group stream members are unlikely to represent the dispersed remnant of a single star formation episode. A subsample of the stream stars appears to be both statistically chemically homogeneous and in the vicinity of the AB Dor nucleus at birth. Their mean metallicity is [Fe/H] = 0.02 ± 0.02 dex, which we consider representative for the AB Dor group. Finally, we report a strong lower limit on the age of the AB Dor nucleus of >110 Myr based on the pre-main sequence contraction times for K-type members which have reached the main sequence.

Seroprevalence of neutralizing antibodies against adenovirus type 14 and 55 in healthy adults in Southern China

PubMed Central

Zheng, Xuehua; Rong, Xia; Feng, Ying; Sun, Xikui; Li, Liang; Wang, Qian; Wang, Min; Liu, Wenkuan; Li, Chufang; Yang, Yiyu; Zhou, Rong; Lu, Jiahai; Feng, Liqiang; Chen, Ling

2017-01-01

Re-emerging human adenovirus types 14 (Ad14) and 55 (Ad55) have caused severe respiratory diseases and even deaths during recent outbreaks. However, the seroprevalence of neutralizing antibodies (nAbs) in healthy adults, which may reflect previous circulation and help to predict potential outbreaks, remains unclear. In this study, we established micro-neutralizing (MN) assays on the basis of recombinant Ad14 and Ad55 reporter viruses, and we investigated serum nAbs in healthy blood donors from Southern China. We found that the overall seropositive rates were 24.8% and 22.4% for Ad14 and Ad55 nAbs, respectively. The seropositive rates were low in individuals younger than 20, and they gradually increased with age. Ad55-seropositive individuals tended to have high nAb titers (>1000), while low (72–200) and moderate (201–1000) nAb levels were frequently observed in Ad14-seropositive ones. Surprisingly, the seropositive rates and nAb levels were associated with the blood type but not the gender of the blood donors, with type AB individuals displaying higher seropositive rates and nAb levels. Interestingly, a significant positive correlation was observed between Ad14 and Ad55 seroprevalence, and higher titers of nAbs were detected in double-positive individuals compared to single-positive ones. These results clarified the human humoral immune responses against Ad14 and Ad55 and revealed a low level of herd immunity in some subpopulations, which emphasized the importance of monitoring these two highly virulent adenoviruses and reinforced the development of prophylactic vaccines. PMID:28588291

Phage display antibodies against ectromelia virus that neutralize variola virus: Selection and implementation for p35 neutralizing epitope mapping.

PubMed

Khlusevich, Yana; Matveev, Andrey; Baykov, Ivan; Bulychev, Leonid; Bormotov, Nikolai; Ilyichev, Ivan; Shevelev, Georgiy; Morozova, Vera; Pyshnyi, Dmitrii; Tikunova, Nina

2018-04-01

In this study, five phage display antibodies (pdAbs) against ectromelia virus (ECTV) were selected from vaccinia virus (VACV)-immune phage-display library of human single chain variable fragments (scFv). ELISA demonstrated that selected pdAbs could recognize ECTV, VACV, and cowpox virus (CPXV). Atomic force microscopy visualized binding of the pdAbs to VACV. Three of the selected pdAbs neutralized variola virus (VARV) in the plaque reduction neutralization test. Western blot analysis of ECTV, VARV, VACV, and CPXV proteins indicated that neutralizing pdAbs bound orthopoxvirus 35 kDa proteins, which are encoded by the open reading frames orthologous to the ORF H3L in VACV. The fully human antibody fh1A was constructed on the base of the VH and VL domains of pdAb, which demonstrated a dose-dependent inhibition of plaque formation after infection with VARV, VACV, and CPXV. To determine the p35 region responsible for binding to neutralizing pdAbs, a panel of truncated p35 proteins was designed and expressed in Escherichia coli cells, and a minimal p35 fragment recognized by selected neutralizing pdAbs was identified. In addition, peptide phage-display combinatorial libraries were applied to localize the epitope. The obtained data indicated that the epitope responsible for recognition by the neutralizing pdAbs is discontinuous and amino acid residues located within two p35 regions, 15-19 aa and 232-237 aa, are involved in binding with neutralizing anti-p35 antibodies. Copyright © 2018. Published by Elsevier B.V.

Radioiodine Labeled Anti-MIF McAb: A Potential Agent for Inflammation Imaging

PubMed Central

Zhang, Chao; Hou, Gui-hua; Han, Jian-kui; Song, Jing; Liang, Ting

2007-01-01

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that may play a role in the pathogenesis of inflammation. Radiolabeled anti-MIF McAb can be used to detect in vivo inflammatory changes. The objective of this study was to investigate in vivo biology of radioiodinated anti-MIF McAb using the inflammation model mice. Anti-MIF McAb was radioiodinated with NaI125 by Iodogen method. Animal models were induced in the mice by intramuscular injection of S. aureus, E. coli, and turpentine oil. The biodistribution studies with radioiodinated anti-MIF McAb were performed on inflammation mice. The relationship between inflammatory lesions and anti-MIF McAb binding was investigated using the percent of injected dose per gram tissue (% ID/g) of tissue samples and whole-body autoradiography. The radioactivity of I125-anti-MIF McAb in the inflammatory tissue increased gradually for three inflammation models. The highest uptake was found in S. aureus group and the lowest was in E. coli group. The uptake in turpentine oil group was average. Whole-body autoradiography showed that all inflammation foci could be visualized clearly from 24 hours after injection, but 48 hours images were much clearer in accordance with the high T/NT ratio. These results demonstrate the ability of radioiodinated anti-MIF McAb to measure in vivo inflammatory events represented by high expression of MIF and suggests that radiolabeled anti-MIF McAb warrants further investigation as a potential inflammation-seeking agent for imaging to detect inflammatory disorders. PMID:18317509

Attentional bias and executive control in treatment-seeking substance-dependent adolescents: A cross-sectional and follow-up study.

PubMed

van Hemel-Ruiter, Madelon E; Wiers, Reinout W; Brook, Frank G; de Jong, Peter J

2016-02-01

Research in adults shows that substance dependent individuals demonstrate attentional bias (AB) for substance-related stimuli. This study investigated the role of AB in adolescents diagnosed with alcohol, cannabis, amphetamine or GHB dependency on entering therapy and six months later, and the role of executive control (EC) as a moderator of the relationship between problem severity and AB. Seventy-eight young substance-dependent (SD) patients (mean age=19.5), and 64 healthy controls (HC; mean age=19.0) were tested. Thirty-eight SD patients took part at 6-month follow-up (FU). AB was indexed by a visual probe task, EC by the attention network task, problem severity by the short alcohol (or drug) use disorder identification test and the severity of dependence questionnaire. SD patients demonstrated an AB for substance stimuli presented for 500 ms and 1250 ms, with the latter related to severity of dependence. There was a nonsignificant tendency indicating that EC was higher in HC than SD participants, but EC did not moderate the relationship between AB and dependency. Substance use, dependency, EC and AB remained unchanged in the 6 month FU period. Young SD patients showed a stronger relatively early as well as maintained AB toward substance cues. A stronger maintained attention was related to higher severity of dependence. Further, there were some indications that EC might play a role in adolescent substance use. The finding that at FU AB and problem severity were not decreased, and EC was not increased underlines the persistent character of addiction. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Single N277A substitution in C2 of simian immunodeficiency virus envelope influences vaccine-elicited CD4i neutralizing and anti-V2 antibody responses.

PubMed

Tang, Xian; Guo, Jia; Cheng, Lin; Sun, Caijun; Liu, Li; Zuo, Teng; Wang, Hui; Chen, Ling; Zhang, Linqi; Chen, Zhiwei

2017-05-02

An effective HIV vaccine remains elusive, and immunogens capable of eliciting protective host humoral immunity have not yet been identified. Although HIV/SIV infections result in the abundant production of CD4-induced (CD4i) antibodies (Abs), these Abs are not protective due to steric restrictions following gp120 binding to CD4 on target cells. Here we report that both DNA- and vaccinia-based vaccines encoding SIV mac239 gp160 readily elicited high levels of CD4i Abs in experimental animals. We identified a highly conserved N-linked glycosylation site N277 in the C2 region which strongly affected the immunogenicity of the CD4i Ab domain. Moreover, a single N277A substitution significantly enhanced the immunogenicity of the V2 domain yielding higher titers and frequency of anti-V2 Ab responses as determined by ELISA and yeast antigen display mapping, respectively. Importantly, immune sera elicited by the N277A-mutated gp160 exhibited elevated antibody-dependent cellular cytotoxicity (ADCC) activity. ADCC activity correlated positively with the anti-V2 Ab titer yet, inversely with CD4i Ab titer. Thus, we identified a determinant of the CD4i domain that might affect vaccine-elicited anti-V2 Ab and ADCC responses to SIV mac239 . Our findings may have implications for design of immunogens to direct B cell recognition in the development of an Ab-based HIV vaccine. Copyright © 2017 Elsevier Ltd. All rights reserved.

Characterization of Pharmaceutical IgG and Biosimilars Using Miniaturized Platforms and LC-MS/MS

PubMed Central

Wooding, Kerry M.; Peng, Wenjing; Mechref, Yehia

2016-01-01

Therapeutic monoclonal antibodies (mAbs) have made a tremendous impact in treating patients with various diseases. MAbs are designed to target specifically a cell and illicit a response from the immune system to destroy the cell. As originator mAb drug patents are coming to an end, generic pharmaceutical companies are poised to replicate and produce so-called biosimilar drugs. MAbs are significantly more complicated than small drugs to analyze and produce. The mAb proteoform and glycoform must be as similar to the original drug as possible to be a viable replacement. The mAb proteoform is well characterized but can be altered through various undesirable reactions such as deamidation. The mAb glycoform is harder to replicate as the glycan formation is a complicated template-less one; it is proving difficult for the originator companies to produce a homogenous population of mAbs from batch to batch. Severe side-effects have occurred in patients taking mAbs with immunogenic glycans, highlighting the importance of quality control mechanisms. The complex nature of mAbs requires sensitive and robust tools amenable to the high-throughput analysis required by a manufacturing setting. Miniaturized analytical platforms for complex biosimilar analysis are still in their infancy but have shown great promise for sample preparation. Capillary electrophoresis-laser induced fluorescence remains a powerful and fast technique for routine glycan analysis. Mass spectrometry is the method of choice for the analysis of mAb proteoforms and is emerging as a powerful tool for glycoform analysis. PMID:27033511

Global analysis of double-strand break processing reveals in vivo properties of the helicase-nuclease complex AddAB

PubMed Central

Badrinarayanan, Anjana; Cisse, Ibrahim I.

2017-01-01

In bacteria, double-strand break (DSB) repair via homologous recombination is thought to be initiated through the bi-directional degradation and resection of DNA ends by a helicase-nuclease complex such as AddAB. The activity of AddAB has been well-studied in vitro, with translocation speeds between 400–2000 bp/s on linear DNA suggesting that a large section of DNA around a break site is processed for repair. However, the translocation rate and activity of AddAB in vivo is not known, and how AddAB is regulated to prevent excessive DNA degradation around a break site is unclear. To examine the functions and mechanistic regulation of AddAB inside bacterial cells, we developed a next-generation sequencing-based approach to assay DNA processing after a site-specific DSB was introduced on the chromosome of Caulobacter crescentus. Using this assay we determined the in vivo rates of DSB processing by AddAB and found that putative chi sites attenuate processing in a RecA-dependent manner. This RecA-mediated regulation of AddAB prevents the excessive loss of DNA around a break site, limiting the effects of DSB processing on transcription. In sum, our results, taken together with prior studies, support a mechanism for regulating AddAB that couples two key events of DSB repair–the attenuation of DNA-end processing and the initiation of homology search by RecA–thereby helping to ensure that genomic integrity is maintained during DSB repair. PMID:28489851

Comparison of Passively Transferred Antibodies in Bighorn and Domestic Lambs Reveals One Factor in Differential Susceptibility of These Species to Mannheimia haemolytica-Induced Pneumonia ▿

PubMed Central

Herndon, Caroline N.; Shanthalingam, Sudarvili; Knowles, Donald P.; Call, Douglas R.; Srikumaran, Subramaniam

2011-01-01

Mannheimia haemolytica consistently causes fatal bronchopneumonia in bighorn sheep (BHS; Ovis canadensis) under natural and experimental conditions. Leukotoxin is the primary virulence factor of this organism. BHS are more susceptible to developing fatal pneumonia than the related species Ovis aries (domestic sheep [DS]). In BHS herds affected by pneumonia, lamb recruitment is severely impaired for years subsequent to an outbreak. We hypothesized that a lack of maternally derived antibodies (Abs) against M. haemolytica provides an immunologic basis for enhanced susceptibility of BH lambs to population-limiting pneumonia. Therefore, the objective of this study was to determine the titers of Abs directed against M. haemolytica in the sera of BH and domestic lambs at birth through 12 weeks of age. Results revealed that BH lambs had approximately 18-fold lower titers of Ab against surface antigens of M. haemolytica and approximately 20-fold lower titers of leukotoxin-neutralizing Abs than domestic lambs. The titers of leukotoxin-neutralizing Abs in the serum and colostrum samples of BH ewes were approximately 157- and 50-fold lower than those for domestic ewes, respectively. Comparatively, the higher titers of parainfluenza 3 virus-neutralizing Abs in the BH lambs ruled out the possibility that these BHS had an impaired ability to passively transfer Abs to their lambs. These results suggest that lower levels of leukotoxin-neutralizing Abs in the sera of BH ewes, and resultant low Ab titers in their lambs, may be a critical factor in the poor lamb recruitment in herds affected by pneumonia. PMID:21613459

Combination sclerostin antibody and zoledronic acid treatment outperforms either treatment alone in a mouse model of osteogenesis imperfecta.

PubMed

Little, David G; Peacock, Lauren; Mikulec, Kathy; Kneissel, Michaela; Kramer, Ina; Cheng, Tegan L; Schindeler, Aaron; Munns, Craig

2017-08-01

In this study, we examined the therapeutic potential of anti-Sclerostin Antibody (Scl-Ab) and bisphosphonate treatments for the bone fragility disorder Osteogenesis Imperfecta (OI). Mice with the Amish OI mutation (Col1a2 G610C mice) and control wild type littermates (WT) were treated from week 5 to week 9 of life with (1) saline (control), (2) zoledronic acid given 0.025mg/kg s.c. weekly (ZA), (3) Scl-Ab given 50mg/kg IV weekly (Scl-Ab), or (4) a combination of both (Scl-Ab/ZA). Functional outcomes were prioritized and included bone mineral density (BMD), bone microarchitecture, long bone bending strength, and vertebral compression strength. By dual-energy absorptiometry, Scl-Ab treatment alone had no effect on tibial BMD, while ZA and Scl-Ab/ZA significantly enhanced BMD by week 4 (+16% and +27% respectively, P<0.05). Scl-Ab/ZA treatment also led to increases in cortical thickness and tissue mineral density, and restored the tibial 4-point bending strength to that of control WT mice. In the spine, all treatments increased compression strength over controls, but only the combined group reached the strength of WT controls. Scl-Ab showed greater anabolic effects in the trabecular bone than in cortical bone. In summary, the Scl-Ab/ZA intervention was superior to either treatment alone in this OI mouse model, however further studies are required to establish its efficacy in other preclinical and clinical scenarios. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

Next-Generation Autoantibody Testing by Combination of Screening and Confirmation-the CytoBead® Technology.

PubMed

Sowa, Mandy; Hiemann, Rico; Schierack, Peter; Reinhold, Dirk; Conrad, Karsten; Roggenbuck, Dirk

2017-08-01

Occurrence of autoantibodies (autoAbs) is a hallmark of autoimmune diseases, and the analysis thereof is an essential part in the diagnosis of organ-specific autoimmune and systemic autoimmune rheumatic diseases (SARD), especially connective tissue diseases (CTDs). Due to the appearance of autoAb profiles in SARD patients and the complexity of the corresponding serological diagnosis, different diagnostic strategies have been suggested for appropriate autoAb testing. Thus, evolving assay techniques and the continuous discovery of novel autoantigens have greatly influenced the development of these strategies. Antinuclear antibody (ANA) analysis by indirect immunofluorescence (IIF) on tissue and later cellular substrates was one of the first tests introduced into clinical routine and is still an indispensable tool for CTD serology. Thus, screening for ANA by IIF is recommended to be followed by confirmatory testing of positive findings employing different assay techniques. Given the continuous growth in the demand for autoAb testing, IIF has been challenged as the standard method for ANA and other autoAb analyses due to lacking automation, standardization, modern data management, and human bias in IIF pattern interpretation. To address these limitations of autoAb testing, the CytoBead® technique has been introduced recently which enables automated interpretation of cell-based IIF and quantitative autoAb multiplexing by addressable microbead immunoassays in one reaction environment. Thus, autoAb screening and confirmatory testing can be combined for the first time. The present review discusses the history of autoAb assay techniques in this context and gives an overview and outlook of the recent progress in emerging technologies.

Manufacturing of High-Concentration Monoclonal Antibody Formulations via Spray Drying-the Road to Manufacturing Scale.

PubMed

Gikanga, Benson; Turok, Robert; Hui, Ada; Bowen, Mayumi; Stauch, Oliver B; Maa, Yuh-Fun

2015-01-01

Spray-dried monoclonal antibody (mAb) powders may offer applications more versatile than the freeze-dried cake, including preparing high-concentration formulations for subcutaneous administration. Published studies on this topic, however, are generally scarce. This study evaluates a pilot-scale spray dryer against a laboratory-scale dryer to spray-dry multiple mAbs in consideration of scale-up, impact on mAb stability, and feasibility of a high-concentration preparation. Under similar conditions, both dryers produced powders of similar properties-for example, water content, particle size and morphology, and mAb stability profile-despite a 4-fold faster output by the pilot-scale unit. All formulations containing arginine salt or a combination of arginine salt and trehalose were able to be spray-dried with high powder collection efficiency (>95%), but yield was adversely affected in formulations with high trehalose content due to powder sticking to the drying chamber. Spray-drying production output was dictated by the size of the dryer operated at an optimal liquid feed rate. Spray-dried powders could be reconstituted to high-viscosity liquids, >300 cP, substantially beyond what an ultrafiltration process can achieve. The molar ratio of trehalose to mAb needed to be reduced to 50:1 in consideration of isotonicity of the formulation with mAb concentration at 250 mg/mL. Even with this low level of sugar protection, long-term stability of spray-dried formulations remained superior to their liquid counterparts based on size variant and potency data. This study offers a commercially viable spray-drying process for biological bulk storage and an option for high-concentration mAb manufacturing. This study evaluates a pilot-scale spray dryer against a laboratory-scale dryer to spray-dry multiple monoclonal antibodies (mAbs) from the perspective of scale-up, impact on mAb stability, and feasibility of a high-concentration preparation. The data demonstrated that there is no process limitation in solution viscosity when high-concentration mAb formulations are prepared from spray-dried powder reconstitution compared with concentration via the conventional ultrafiltration process. This study offers a commercially viable spray-drying process for biological bulk storage and a high-concentration mAb manufacturing option for subcutaneous administration. The outcomes of this study will benefit scientists and engineers who develop high-concentration mAb products by providing a viable manufacturing alternative. © PDA, Inc. 2015.

Intracellular Crosslinking of Filoviral Nucleoproteins with Xintrabodies Restricts Viral Packaging

PubMed Central

Darling, Tamarand Lee; Sherwood, Laura Jo; Hayhurst, Andrew

2017-01-01

Viruses assemble large macromolecular repeat structures that become part of the infectious particles or virions. Ribonucleocapsids (RNCs) of negative strand RNA viruses are a prime example where repetition of nucleoprotein (NP) along the genome creates a core polymeric helical scaffold that accommodates other nucleocapsid proteins including viral polymerase. The RNCs are transported through the cytosol for packaging into virions through association with viral matrix proteins at cell membranes. We hypothesized that RNC would be ideal targets for crosslinkers engineered to promote aberrant protein–protein interactions, thereby blocking their orderly transport and packaging. Previously, we had generated single-domain antibodies (sdAbs) against Filoviruses that have all targeted highly conserved C-terminal regions of NP known to be repetitively exposed along the length of the RNCs of Marburgvirus (MARV) and Ebolavirus (EBOV). Our crosslinker design consisted of dimeric sdAb expressed intracellularly, which we call Xintrabodies (X- for crosslinking). Electron microscopy of purified NP polymers incubated with purified sdAb constructs showed NP aggregation occurred in a genus-specific manner with dimeric and not monomeric sdAb. A virus-like particle (VLP) assay was used for initial evaluation where we found that dimeric sdAb inhibited NP incorporation into VP40-based VLPs whereas monomeric sdAb did not. Inhibition of NP packaging was genus specific. Confocal microscopy revealed dimeric sdAb was diffuse when expressed alone but focused on pools of NP when the two were coexpressed, while monomeric sdAb showed ambivalent partition. Infection of stable Vero cell lines expressing dimeric sdAb specific for either MARV or EBOV NP resulted in smaller plaques and reduced progeny of cognate virus relative to wild-type Vero cells. Though the impact was marginal at later time-points, the collective data suggest that viral replication can be reduced by crosslinking intracellular NP using relatively small amounts of dimeric sdAb to restrict NP packaging. The stoichiometry and ease of application of the approach would likely benefit from transitioning away from intracellular expression of crosslinking sdAb to exogenous delivery of antibody. By retuning sdAb specificity, the approach of crosslinking highly conserved regions of assembly critical proteins may well be applicable to inhibiting replication processes of a broad spectrum of viruses. PMID:29021793

Comparison of the osteogenesis and fusion rates between activin A/BMP-2 chimera (AB204) and rhBMP-2 in a beagle's posterolateral lumbar spine model.

PubMed

Zheng, Guang Bin; Yoon, Byung-Hak; Lee, Jae Hyup

2017-10-01

Activin A/BMP-2 chimera (AB204) could promote bone healing more effectively than recombinant bone morphogenetic protein 2 (rhBMP-2) with much lower dose in a rodent model, but there is no report about the effectiveness of AB204 in a large animal model. The purpose of this study was to compare the osteogenesis and fusion rate between AB204 and rhBMP-2 using biphasic calcium phosphate (BCP) as a carrier in a beagle's posterolateral lumbar fusion model. This is a randomized control animal study. Seventeen male beagle dogs were included. Bilateral posterolateral fusion was performed at the L1-L2 and L4-L5 levels. Biphasic calcium phosphate (2 cc), rhBMP-2 (50 µg)+BCP (2 cc), or AB204 (50 µg)+BCP (2 cc) were implanted into the intertransverse space randomly. X-ray was performed at 4 and 8 weeks. After 8 weeks, the animals were sacrificed, and new bone formation and fusion rate were evaluated by manual palpation, computed tomography (CT), and undecalcified histology. The AB204 group showed significantly higher fusion rate (90%) than the rhBMP-2 group (15%) or the Osteon group (6.3%) by manual palpation. On x-ray and CT assessment, fusion rate and the volume of newly formed bone were also significantly higher in AB204 group than other groups. In contrast, more osteolysis was found in rhBMP-2 group (40%) than in AB204 group (10%) on CT study. In histologic results, new bone formation was sufficient between transverse processes in AB204 group, and obvious trabeculation and bone remodeling were observed. But in rhBMP-2 group, new bone formation was less than AB204 group and osteolysis was observed between the intertransverse spaces. A low dose of AB204 with BCP as a carrier significantly promotes the fusion rate in a large animal model when compared with the rhBMP-2. These findings demonstrate that AB204 could be an alternative to rhBMP-2 to improve fusion rate. Copyright © 2017 Elsevier Inc. All rights reserved.

Clinical aggressiveness and long-term outcome in patients with papillary thyroid cancer and circulating anti-thyroglobulin autoantibodies.

PubMed

Durante, Cosimo; Tognini, Sara; Montesano, Teresa; Orlandi, Fabio; Torlontano, Massimo; Puxeddu, Efisio; Attard, Marco; Costante, Giuseppe; Tumino, Salvatore; Meringolo, Domenico; Bruno, Rocco; Trulli, Fabiana; Toteda, Maria; Redler, Adriano; Ronga, Giuseppe; Filetti, Sebastiano; Monzani, Fabio

2014-07-01

The association between papillary thyroid cancer (PTC) and Hashimoto's thyroiditis is widely recognized, but less is known about the possible link between circulating anti-thyroglobulin antibody (TgAb) titers and PTC aggressiveness. To shed light on this issue, we retrospectively examined a large series of PTC patients with and without positive TgAb. Data on 220 TgAb-positive PTC patients (study cohort) were retrospectively collected in 10 hospital-based referral centers. All the patients had undergone near-total thyroidectomy with or without radioiodine remnant ablation. Tumor characteristics and long-term outcomes (follow-up range: 2.5-24.8 years) were compared with those recently reported in 1020 TgAb-negative PTC patients with similar demographic characteristics. We also assessed the impact on clinical outcome of early titer disappearance in the TgAb-positive group. At baseline, the study cohort (mean age 45.9 years, range 12.5-84.1 years; 85% female) had a significantly higher prevalence of high-risk patients (6.9% vs. 3.2%, p<0.05) and extrathyroidal tumor extension (28.2% vs. 24%; p<0.0001) than TgAb-negative controls. Study cohort patients were also more likely than controls to have persistent disease at the 1-year visit (13.6% vs. 7.0%, p=0.001) or recurrence during subsequent follow-up (5.8% vs. 1.4%, p=0.0001). At the final follow-up visit, the percentage of patients with either persistent or recurrent disease in the two cohorts was significantly different (6.4% of TgAb-positive patients vs. 1.7% in the TgAb-negative group, p<0.0001). At the 1-year visit, titer normalization was observed in 85 of the 220 TgAb-positive individuals. These patients had a significantly lower rate of persistent disease than those who were still TgAb positive (8.2% vs. 17.3%. p=0.05), and no relapses were observed among patients with no evidence of disease during subsequent follow-up. PTC patients with positive serum TgAb titer during the first year after primary treatment were more likely to have persistent/recurrent disease than those who were consistently TgAb-negative. Negative titers at 1 year may be associated with more favorable outcomes.

Non Target Effect of Cry1 Ab and Cry Ab x Cry3 Bb1 Bt Transgenic Maize on Orius Insidiosus (Hemiptera: Anthocoridae) Abundance

USDA-ARS?s Scientific Manuscript database

Non-target effects of Cry1Ab x CP4 EPSPS and Cry1Ab + Cry3Bb1 x CP4 EPSPS Bt transgenic new maize hybrids on insidious flower bugs [Orius insidiosus (Say)] was studied in Nebraska (Mead, C lay Center, and Concord) during 2007 and 2008. The Bt effect was compared to CP4 EPSPS maize (isoline), convent...

Mac-1 (CD11b/CD18) is essential for Fc receptor-mediated neutrophil cytotoxicity and immunologic synapse formation.

PubMed

van Spriel, A B; Leusen, J H; van Egmond, M; Dijkman, H B; Assmann, K J; Mayadas, T N; van de Winkel, J G

2001-04-15

Receptors for human immunoglobulin (Ig)G and IgA initiate potent cytolysis of antibody (Ab)-coated targets by polymorphonuclear leukocytes (PMNs). Mac-1 (complement receptor type 3, CD11b/CD18) has previously been implicated in receptor cooperation with Fc receptors (FcRs). The role of Mac-1 in FcR-mediated lysis of tumor cells was characterized by studying normal human PMNs, Mac-1-deficient mouse PMNs, and mouse PMNs transgenic for human FcR. All PMNs efficiently phagocytosed Ab-coated particles. However, antibody-dependent cellular cytotoxicity (ADCC) was abrogated in Mac-1(-/-) PMNs and in human PMNs blocked with anti-Mac-1 monoclonal Ab (mAb). Mac-1(-/-) PMNs were unable to spread on Ab-opsonized target cells and other Ab-coated surfaces. Confocal laser scanning and electron microscopy revealed a striking difference in immunologic synapse formation between Mac-1(-/-) and wild-type PMNs. Also, respiratory burst activity could be measured outside membrane-enclosed compartments by using Mac-1(-/-) PMNs bound to Ab-coated tumor cells, in contrast to wild-type PMNs. In summary, these data document an absolute requirement of Mac-1 for FcR-mediated PMN cytotoxicity toward tumor targets. Mac-1(-/-) PMNs exhibit defective spreading on Ab-coated targets, impaired formation of immunologic synapses, and absent tumor cytolysis.

Hexon and fiber of adenovirus type 14 and 55 are major targets of neutralizing antibody but only fiber-specific antibody contributes to cross-neutralizing activity.

PubMed

Feng, Ying; Sun, Xikui; Ye, Xianmiao; Feng, Yupeng; Wang, Jinlin; Zheng, Xuehua; Liu, Xinglong; Yi, Changhua; Hao, Mingli; Wang, Qian; Li, Feng; Xu, Wei; Li, Liang; Li, Chufang; Zhou, Rong; Chen, Ling; Feng, Liqiang

2018-05-01

Re-emerging human adenoviruses type 14 (HAdV14) and 55 (HAdV55) represent two highly virulent adenoviruses. The neutralizing antibody (nAb) responses elicited by infection or immunization remain largely unknown. Herein, we generated hexon-chimeric HAdV14 viruses harboring each single or entire hexon hyper-variable-regions (HVR) from HAdV55, and determined the neutralizing epitopes of human and mouse nAbs. In human sera, hexon-targeting nAbs are type-specific and mainly recognize HVR2, 5, and 7. Fiber-targeting nAbs are only detectable in sera cross-neutralizing HAdV14 and HAdV55 and contribute substantially to cross-neutralization. Penton-binding antibodies, however, show no significant neutralizing activities. In mice immunized with HAdV14 or HAdV55, a single immunization mainly elicited hexon-specific nAbs, which recognized HAdV14 HVR1, 2, and 7 and HAdV55 HVR1 and 2, respectively. After a booster immunization, cross-neutralizing fiber-specific nAbs became detectable. These results indicated that hexon elicits type-specific nAbs whereas fiber induces cross-neutralizing nAbs to HAdV14 and HAdV55, which are of significance in vaccine development. Copyright © 2018 Elsevier Inc. All rights reserved.

Carbon dots based immunosorbent assay for the determination of GFAP in human serum

NASA Astrophysics Data System (ADS)

Ma, Yunsu; Xu, Guanhong; Wei, Fangdi; Cen, Yao; Song, Yueyue; Ma, Yujie; Xu, Xiaoman; Shi, Menglan; Sohail, Muhammad; Hu, Qin

2018-04-01

Glial fibrillary acidic protein (GFAP) is expressed in the central nervous system and the level of GFAP normally rises with brain injury and astroglial tumors. So, serum GFAP is used as a marker for diagnosing various types of brain damage and astroglial tumors. In this study, a new sensor based on carbon dots (CDs) linked with antibodies to specifically detect GFAP in human serum was developed. Anti-GFAP (Ab1) linked with protein A/G agarose resin (PA/G) as a capture antibody (PA/G-Ab1) and anti-GFAP (Ab2) labeled with CDs as a detection antibody (CDs-Ab2) were prepared firstly. Then the CD-linked antibody immunosorbent assay (CLAISA) method was constructed based on the sandwich conjunction reaction among PA/G-Ab1, GFAP, and CDs-Ab2. CLAISA, using the fluorescence of PA/G-Ab1-GFAP-Ab2-CDs as the direct signal, enabled the proposed immunosensor to detect GFAP sensitively with a linear range of 0.10-8.00 ng ml-1 and a detection limit of 25 pg ml-1. This method was applied to the determination of GFAP in human serum by the standard addition method, and the results showed high accuracy and precision. Considering the easy synthetic process and excellent performance of CLAISA, this method has great potential to be used to monitor GFAP in the clinic.

Small-angle neutron scattering study of a monoclonal antibody using free-energy constraints.

PubMed

Clark, Nicholas J; Zhang, Hailiang; Krueger, Susan; Lee, Hyo Jin; Ketchem, Randal R; Kerwin, Bruce; Kanapuram, Sekhar R; Treuheit, Michael J; McAuley, Arnold; Curtis, Joseph E

2013-11-14

Monoclonal antibodies (mAbs) contain hinge-like regions that enable structural flexibility of globular domains that have a direct effect on biological function. A subclass of mAbs, IgG2, have several interchain disulfide bonds in the hinge region that could potentially limit structural flexibility of the globular domains and affect the overall configuration space available to the mAb. We have characterized human IgG2 mAb in solution via small-angle neutron scattering (SANS) and interpreted the scattering data using atomistic models. Molecular Monte Carlo combined with molecular dynamics simulations of a model mAb indicate that a wide range of structural configurations are plausible, spanning radius of gyration values from ∼39 to ∼55 Å. Structural ensembles and representative single structure solutions were derived by comparison of theoretical SANS profiles of mAb models to experimental SANS data. Additionally, molecular mechanical and solvation free-energy calculations were carried out on the ensemble of best-fitting mAb structures. The results of this study indicate that low-resolution techniques like small-angle scattering combined with atomistic molecular simulations with free-energy analysis may be helpful to determine the types of intramolecular interactions that influence function and could lead to deleterious changes to mAb structure. This methodology will be useful to analyze small-angle scattering data of many macromolecular systems.

Anti-GM2 ganglioside antibodies are a biomarker for acute canine polyradiculoneuritis.

PubMed

Rupp, Angie; Galban-Horcajo, Francesc; Bianchi, Ezio; Dondi, Maurizio; Penderis, Jacques; Cappell, Joanna; Burgess, Karl; Matiasek, Kaspar; McGonigal, Rhona; Willison, Hugh J

2013-03-01

Acute canine polyradiculoneuritis (ACP) is considered to be the canine equivalent of the human peripheral nerve disorder Guillain-Barré syndrome (GBS); an aetiological relationship, however, remains to be demonstrated. In GBS, anti-glycolipid antibodies (Abs) are considered as important disease mediators. To address the possibility of common Ab biomarkers, the sera of 25 ACP dogs, 19 non-neurological, and 15 epileptic control dogs were screened for IgG Abs to 10 glycolipids and their 1 : 1 heteromeric complexes using combinatorial glycoarrays. Anti-GM2 ganglioside Abs were detected in 14/25 ACP dogs, and anti-GA1 Abs in one further dog. All controls except for one were negative for anti-glycolipid Abs. In this cohort of cases and controls, the glycoarray screen reached a diagnostic sensitivity of 60% and a specificity of 97%; a lower sensitivity (32%) was reported using a conventional glycolipid ELISA. To address the possible pathogenic role for anti-GM2 Abs in ACP, we identified GM2 in canine sciatic nerve by both mass spectrometry and thin layer chromatography overlay. In immunohistological studies, GM2 was localized predominantly to the abaxonal Schwann cell membrane. The presence of anti-GM2 Abs in ACP suggests that it may share a similar pathophysiology with GBS, for which it could thus be considered a naturally occurring animal model. © 2013 Peripheral Nerve Society.

Poly(ethylene oxide)-Assisted Macromolecular Self-Assembly of Lignin in ABS Matrix for Sustainable Composite Applications

DOE PAGES

Akato, Kokouvi M.; Tran, Chau D.; Chen, Jihua; ...

2015-11-05

Here we report the compatibilization of biomass-derived lignin polymer in acrylonitrile butadiene styrene (ABS) thermoplastic matrix without loss of mechanical properties via poly(ethylene oxide) (PEO)-mediated macromolecular self-assembly. ABS was blended with lignin in different concentrations, and blends with 10 wt % PEO (relative to lignin) were prepared. The relative tensile strength improved slightly at low lignin content but diminished rapidly as the lignin content was increased. However, the inclusion of PEO as an interfacial adhesion promoter helped avoid deleterious effects. Dynamic mechanical analysis showed that PEO plasticized the hard phase and thus lowered the activation energy (E a) for itsmore » relaxation but caused stiffening of the soft phase and increased its E a. Microscopy revealed that incorporating lignin in ABS led to the statistical dispersion of discrete lignin domains (300–1000 nm) which, after PEO addition, were reduced to smaller interconnected particles (200–500 nm). The lignin-extended partially renewable ABS resins showed shear-thinning behavior and reduced viscosity compared to neat ABS. The preferred lignin-loaded compositions reinforced with 20 vol % chopped carbon fibers exhibited mechanical performances (77–80 MPa) equivalent to those of reinforced ABS materials reportedly used in 3D printing applications. In conclusion, this approach could lower the cost of ABS while reducing its carbon footprint.« less

Anti-GM2 ganglioside antibodies are a biomarker for acute canine polyradiculoneuritis

PubMed Central

Bianchi, Ezio; Dondi, Maurizio; Penderis, Jacques; Cappell, Joanna; Burgess, Karl; Matiasek, Kaspar; McGonigal, Rhona; Willison, Hugh J.

2016-01-01

Acute canine polyradiculoneuritis (ACP) is considered to be the canine equivalent of the human peripheral nerve disorder Guillain-Barré syndrome (GBS); an aetiological relationship, however, remains to be demonstrated. In GBS, anti-glycolipid antibodies (Abs) are considered as important disease mediators. To address the possibility of common Ab biomarkers, the sera of 25 ACP dogs, 19 non-neurological, and 15 epileptic control dogs were screened for IgG Abs to 10 glycolipids and their 1 : 1 heteromeric complexes using combinatorial glycoarrays. Anti-GM2 ganglioside Abs were detected in 14/25 ACP dogs, and anti-GA1 Abs in one further dog. All controls except for one were negative for anti-glycolipid Abs. In this cohort of cases and controls, the glycoarray screen reached a diagnostic sensitivity of 60% and a specificity of 97%; a lower sensitivity (32%) was reported using a conventional glycolipid ELISA. To address the possible pathogenic role for anti-GM2 Abs in ACP, we identified GM2 in canine sciatic nerve by both mass spectrometry and thin layer chromatography overlay. In immunohistological studies, GM2 was localized predominantly to the abaxonal Schwann cell membrane. The presence of anti-GM2 Abs in ACP suggests that it may share a similar pathophysiology with GBS, for which it could thus be considered a naturally occurring animal model. PMID:23521648

Occurrence, leaching, and degradation of Cry1Ab protein from transgenic maize detritus in agricultural streams

DOE PAGES

Griffiths, Natalie A.; Tank, Jennifer L.; Royer, Todd V.; ...

2017-03-15

The insecticidal Cry1Ab protein expressed by transgenic (Bt) maize can enter adjacent water bodies via multiple pathways, but its fate in stream ecosystems is not as well studied as in terrestrial systems. In this study, we used a combination of field sampling and laboratory experiments to examine the occurrence, leaching, and degradation of soluble Cry1Ab protein derived from Bt maize in agricultural streams. We surveyed 11 agricultural streams in northwestern Indiana, USA, on 6 dates that encompassed the growing season, crop harvest, and snowmelt/spring flooding, and detected Cry1Ab protein in the water column and in flowing subsurface tile drains atmore » concentrations of 3–60 ng/L. In a series of laboratory experiments, submerged Bt maize leaves leached Cry1Ab into stream water with 1% of the protein remaining in leaves after 70 d. Laboratory experiments suggested that dissolved Cry1Ab protein degraded rapidly in microcosms containing water-column microorganisms, and light did not enhance breakdown by stimulating assimilatory uptake of the protein by autotrophs. Here, the common detection of Cry1Ab protein in streams sampled across an agricultural landscape, combined with laboratory studies showing rapid leaching and degradation, suggests that Cry1Ab may be pseudo-persistent at the watershed scale due to the multiple input pathways from the surrounding terrestrial environment.« less

Detection of anti-isoniazid and anti-cytochrome P450 antibodies in patients with isoniazid-induced liver failure.

PubMed

Metushi, Imir G; Sanders, Corron; Lee, William M; Uetrecht, Jack

2014-03-01

Isoniazid (INH)-induced hepatotoxicity remains one of the most common causes of drug-induced idiosyncratic liver injury and liver failure. This form of liver injury is not believed to be immune-mediated because it is not usually associated with fever or rash, does not recur more rapidly on rechallenge, and previous studies have failed to identify anti-INH antibodies (Abs). In this study, we found Abs present in sera of 15 of 19 cases of INH-induced liver failure. Anti-INH Abs were present in 8 sera; 11 had anti-cytochrome P450 (CYP)2E1 Abs, 14 had Abs against CYP2E1 modified by INH, 14 had anti-CYP3A4 antibodies, and 10 had anti-CYP2C9 Abs. INH was found to form covalent adducts with CYP2E1, CYP3A4, and CYP2C9. None of these Abs were detected in sera from INH-treated controls without significant liver injury. The presence of a range of antidrug and autoAbs has been observed in other drug-induced liver injury that is presumed to be immune mediated. These data provide strong evidence that INH induces an immune response that causes INH-induced liver injury. © 2014 by the American Association for the Study of Liver Diseases.

Occurrence, leaching, and degradation of Cry1Ab protein from transgenic maize detritus in agricultural streams

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

Griffiths, Natalie A.; Tank, Jennifer L.; Royer, Todd V.

The insecticidal Cry1Ab protein expressed by transgenic (Bt) maize can enter adjacent water bodies via multiple pathways, but its fate in stream ecosystems is not as well studied as in terrestrial systems. In this study, we used a combination of field sampling and laboratory experiments to examine the occurrence, leaching, and degradation of soluble Cry1Ab protein derived from Bt maize in agricultural streams. We surveyed 11 agricultural streams in northwestern Indiana, USA, on 6 dates that encompassed the growing season, crop harvest, and snowmelt/spring flooding, and detected Cry1Ab protein in the water column and in flowing subsurface tile drains atmore » concentrations of 3–60 ng/L. In a series of laboratory experiments, submerged Bt maize leaves leached Cry1Ab into stream water with 1% of the protein remaining in leaves after 70 d. Laboratory experiments suggested that dissolved Cry1Ab protein degraded rapidly in microcosms containing water-column microorganisms, and light did not enhance breakdown by stimulating assimilatory uptake of the protein by autotrophs. Here, the common detection of Cry1Ab protein in streams sampled across an agricultural landscape, combined with laboratory studies showing rapid leaching and degradation, suggests that Cry1Ab may be pseudo-persistent at the watershed scale due to the multiple input pathways from the surrounding terrestrial environment.« less

Flow shear stress differentially regulates endothelial uptake of nanocarriers targeted to distinct epitopes of PECAM-1.

PubMed

Han, Jingyan; Shuvaev, Vladimir V; Davies, Peter F; Eckmann, David M; Muro, Silvia; Muzykantov, Vladimir R

2015-07-28

Targeting nanocarriers (NC) to endothelial cell adhesion molecules including Platelet-Endothelial Cell Adhesion Molecule-1 (PECAM-1 or CD31) improves drug delivery and pharmacotherapy of inflammation, oxidative stress, thrombosis and ischemia in animal models. Recent studies unveiled that hydrodynamic conditions modulate endothelial endocytosis of NC targeted to PECAM-1, but the specificity and mechanism of effects of flow remain unknown. Here we studied the effect of flow on endocytosis by human endothelial cells of NC targeted by monoclonal antibodies Ab62 and Ab37 to distinct epitopes on the distal extracellular domain of PECAM. Flow in the range of 1-8dyn/cm(2), typical for venous vasculature, stimulated the uptake of spherical Ab/NC (~180nm diameter) carrying ~50 vs 200 Ab62 and Ab37 per NC, respectively. Effect of flow was inhibited by disruption of cholesterol-rich plasmalemma domains and deletion of PECAM-1 cytosolic tail. Flow stimulated endocytosis of Ab62/NC and Ab37/NC via eliciting distinct signaling pathways mediated by RhoA/ROCK and Src Family Kinases, respectively. Therefore, flow stimulates endothelial endocytosis of Ab/NC in a PECAM-1 epitope specific manner. Using ligands of binding to distinct epitopes on the same target molecule may enable fine-tuning of intracellular delivery based on the hemodynamic conditions in the vascular area of interest. Copyright © 2015 Elsevier B.V. All rights reserved.

Protective B-cell epitopes of Francisella tularensis O-polysaccharide in a mouse model of respiratory tularaemia.

PubMed

Lu, Zhaohua; Madico, Guillermo; Roche, Marly I; Wang, Qi; Hui, Julia H; Perkins, Hillary M; Zaia, Joseph; Costello, Catherine E; Sharon, Jacqueline

2012-07-01

Antibodies to the lipopolysaccharide (LPS) of Francisella tularensis have been shown to be protective against respiratory tularaemia in mouse models, and we have previously described mouse monoclonal antibodies (mAbs) to non-overlapping terminal and internal epitopes of the F. tularensis LPS O-polysaccharide (OAg). In the current study, we used F. tularensis LPS oligosaccharides of defined OAg repeat length as molecular rulers in competition ELISA to demonstrate that the epitope targeted by the terminal OAg-binding mAb FB11 is contained within one tetrasaccharide repeat whereas the epitope targeted by the internal OAg-binding mAb Ab52 spans two tetrasaccharide repeats. Both mAbs conferred survival to BALB/c mice infected intranasally with the F. tularensis type B live vaccine strain and prolonged survival of BALB/c mice infected intranasally with the highly virulent F. tularensis type A strain SchuS4. The protective effects correlated with reduced bacterial burden in mAb-treated infected mice. These results indicate that an oligosaccharide with two OAg tetrasaccharide repeats covers both terminal and internal protective OAg epitopes, which may inform the design of vaccines for tularaemia. Furthermore, the FB11 and Ab52 mAbs could serve as reporters to monitor the response of vaccine recipients to protective B-cell epitopes of F. tularensis OAg. © 2012 The Authors. Immunology © 2012 Blackwell Publishing Ltd.

A novel impedance-based cellular assay for the detection of anti-calcium channel autoantibodies in type 1 diabetes.

PubMed

Jackson, Michael W; Gordon, Tom P

2010-09-30

We have recently postulated that functional autoantibodies (Abs) against L-type voltage-gated calcium channels (VGCCs) contribute to autonomic dysfunction in type 1 diabetes (T1D). Previous studies based on whole-organ assays have proven valuable in establishing the mechanism of anti-VGCC Ab activity, but are complex and unsuitable for screening large patient cohorts. In the current study, we used real-time dynamic monitoring of cell impedance to demonstrate that anti-VGCC Abs from patients with T1D inhibit the adherence of Rin A12 cells. The functional effect of the anti-VGCC Abs was mimicked by the dihydropyridine agonist, Bay K8644, and reversed by the antagonist, nicardipine, providing a pharmacological link to the whole-organ studies. IVIg neutralized the effect on cell adhesion of the anti-VGCC Abs, consistent with the presence of anti-idiotypic Abs in IVIg that may prevent the emergence of pathogenic Abs in healthy individuals. The cell impedance assay can be performed in a 96 well plate format, and represents a simple method for detecting the presence of anti-VGCC activity in patient immunoglobulin (IgG). The new cell assay should prove useful for further studies to determine the prevalence of the Ab and its association with symptoms of autonomic dysfunction in patients with T1D. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Co-targeting of EGFR and autophagy signaling is an emerging treatment strategy in metastatic colorectal cancer.

PubMed

Koustas, Evangelos; Karamouzis, Michalis V; Mihailidou, Chrysovalantou; Schizas, Dimitrios; Papavassiliou, Athanasios G

2017-06-28

The epidermal growth factor receptor (EGFR) and its associated pathway is a critical key regulator of CRC development and progression. The monoclonal antibodies (MoAbs) cetuximab and panitumumab, directed against EGFR, represent a major step forward in the treatment of metastatic colorectal cancer (mCRC), in terms of progression-free survival and overall survival in several clinical trials. However, the activity of anti-EGFR MoAbs appears to be limited to a subset of patients with mCRC. Studies have highlighted that acquired-resistance to anti-EGFR MoAbs biochemically converge into Ras/Raf/Mek/Erk and PI3K/Akt/mTOR pathways. Recent data also suggest that acquired-resistance to anti-EGFR MoAbs is accompanied by inhibition of EGFR internalization, ubiqutinization, degradation and prolonged downregulation. It is well established that autophagy, a self-cannibalization process, is considered to be associated with resistance to the anti-EGFR MoAbs therapy. Additionally, autophagy induced by anti-EGFR MoAbs acts as a protective response in cancer cells. Thus, inhibition of autophagy after treatment with EGFR MoAbs can result in autophagic cell death. A combination therapy comprising of anti-EGFR MoAbs and autophagy inhibitors would represent a multi-pronged approach that could be evolved into an active therapeutic strategy in mCRC patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Antibodies causing thrombocytopenia in patients treated with RGD-mimetic platelet inhibitors recognize ligand-specific conformers of αIIb/β3 integrin

PubMed Central

Rasmussen, Mark; Zhu, Jieqing; Aster, Richard H.

2012-01-01

Arginine-glycine-aspartic acid (RGD)–mimetic platelet inhibitors act by occupying the RGD recognition site of αIIb/β3 integrin (GPIIb/IIIa), thereby preventing the activated integrin from reacting with fibrinogen. Thrombocytopenia is a well-known side effect of treatment with this class of drugs and is caused by Abs, often naturally occurring, that recognize αIIb/β3 in a complex with the drug being administered. RGD peptide and RGD-mimetic drugs are known to induce epitopes (ligand-induced binding sites [LIBS]) in αIIb/β3 that are recognized by certain mAbs. It has been speculated, but not shown experimentally, that Abs from patients who develop thrombocytopenia when treated with an RGD-mimetic inhibitor similarly recognize LIBS determinants. We addressed this question by comparing the reactions of patient Abs and LIBS-specific mAbs against αIIb/β3 in a complex with RGD and RGD-mimetic drugs, and by examining the ability of selected non-LIBS mAbs to block binding of patient Abs to the liganded integrin. Findings made provide evidence that the patient Abs recognize subtle, drug-induced structural changes in the integrin head region that are clustered about the RGD recognition site. The target epitopes differ from classic LIBS determinants, however, both in their location and by virtue of being largely drug-specific. PMID:22490676

Delivery of Flightless I Neutralizing Antibody from Porous Silicon Nanoparticles Improves Wound Healing in Diabetic Mice.

PubMed

Turner, Christopher T; McInnes, Steven J P; Melville, Elizabeth; Cowin, Allison J; Voelcker, Nicolas H

2017-01-01

Flightless I (Flii) is elevated in human chronic wounds and is a negative regulator of wound repair. Decreasing its activity improves healing responses. Flii neutralizing antibodies (FnAbs) decrease Flii activity in vivo and hold significant promise as healing agents. However, to avoid the need for repeated application in a clinical setting and to protect the therapeutic antibody from the hostile environment of the wound, suitable delivery vehicles are required. In this study, the use of porous silicon nanoparticles (pSi NPs) is demonstrated for the controlled release of FnAb to diabetic wounds. We achieve FnAb loading regimens exceeding 250 µg antibody per mg of vehicle. FnAb-loaded pSi NPs increase keratinocyte proliferation and enhance migration in scratch wound assays. Release studies confirm the functionality of the FnAb in terms of Flii binding. Using a streptozotocin-induced model of diabetic wound healing, a significant improvement in healing is observed for mice treated with FnAb-loaded pSi NPs compared to controls, including FnAb alone. FnAb-loaded pSi NPs treated with proteases show intact and functional antibody for up to 7 d post-treatment, suggesting protection of the antibodies from proteolytic degradation in wound fluid. pSi NPs may therefore enable new therapeutic approaches for the treatment of diabetic ulcers. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Inhibitory Monoclonal Antibodies against Mouse Proteases Raised in Gene-Deficient Mice Block Proteolytic Functions in vivo

PubMed Central

Lund, Ida K.; Rasch, Morten G.; Ingvarsen, Signe; Pass, Jesper; Madsen, Daniel H.; Engelholm, Lars H.; Behrendt, Niels; Høyer-Hansen, Gunilla

2012-01-01

Identification of targets for cancer therapy requires the understanding of the in vivo roles of proteins, which can be derived from studies using gene-targeted mice. An alternative strategy is the administration of inhibitory monoclonal antibodies (mAbs), causing acute disruption of the target protein function(s). This approach has the advantage of being a model for therapeutic targeting. mAbs for use in mouse models can be obtained through immunization of gene-deficient mice with the autologous protein. Such mAbs react with both species-specific epitopes and epitopes conserved between species. mAbs against proteins involved in extracellular proteolysis, including plasminogen activators urokinase plasminogen activator (uPA), tissue-type plasminogen activator (tPA), their inhibitor PAI-1, the uPA receptor (uPAR), two matrix metalloproteinases (MMP9 and MMP14), as well as the collagen internalization receptor uPARAP, have been developed. The inhibitory mAbs against uPA and uPAR block plasminogen activation and thereby hepatic fibrinolysis in vivo. Wound healing, another plasmin-dependent process, is delayed by an inhibitory mAb against uPA in the adult mouse. Thromboembolism can be inhibited by anti-PAI-1 mAbs in vivo. In conclusion, function-blocking mAbs are well-suited for targeted therapy in mouse models of different diseases, including cancer. PMID:22754528

Molecular characterization and functions of fatty acid and retinoid binding protein gene (Ab-far-1) in Aphelenchoides besseyi.

PubMed

Cheng, Xi; Xiang, Yu; Xie, Hui; Xu, Chun-Ling; Xie, Teng-Fei; Zhang, Chao; Li, Yu

2013-01-01

Rice white tip nematode, Aphelenchoides besseyi, is a kind of plant parasitic nematodes that cause serious losses in rice and many other crops. Fatty acid and retinoid binding protein (FAR) is a specific protein in nematodes and is related to development, reproduction, infection to the host, and disruption of plant defense reactions, so the inhibition of FAR function is the potential approach to control A. besseyi. The full-length of Ab-far-1 cDNA is 805 bp, including 546 bp of ORF that encodes 181 amino acids. Software analysis revealed that the Ab-FAR-1 was rich in α-helix structure, contained a predicted consensus casein kinase II phosphorylation site and a hydrophobic secretory signal peptide, but did not have glycosylation sites. The Ab-FAR-1 had 52% homology to Gp-FAR-1 protein. The Ab-FAR-1 and Gp-FAR-1 were grouped in the same branch according to the phylogenetic tree. Fluorescence-based ligand binding analysis confirmed that the recombinant Ab-FAR-1 (rAb-FAR-1) bound with the fluorescent analogues 11-((5-dimethylaminonaphthalene-1-sulfonyl) amino) undecannoic acid (DAUDA), cis-parinaric acid and retinol, but the oleic acid would compete with the binding site. Quantitative PCR (qPCR) was used to assess the expression level of Ab-far-1 at different development stages of A. besseyi, the highest expression was found in the females, followed by eggs, juveniles and males. Using in situ hybridization technique, Ab-far-1 mRNA was present in the hypodermis of juveniles and adults, the ovaries of females and the testes of males. When A. besseyi was treated with Ab-far-1 dsRNA for 48 h, the silencing efficiency of Ab-far-1 was the best and the number of nematodes on the carrot was the least. Thus FAR plays important roles in the development and reproduction of nematodes. This study is useful and helpful to figure out a new way to control the plant parasitic nematodes.

B-cell subsets in the joint compartments of seropositive and seronegative rheumatoid arthritis (RA) and No-RA arthritides express memory markers and ZAP70 and characterize the aggregate pattern irrespectively of the autoantibody status.

PubMed

Michelutti, Alessandro; Gremese, Elisa; Morassi, Francesca; Petricca, Luca; Arena, Vincenzo; Tolusso, Barbara; Alivernini, Stefano; Peluso, Giusy; Bosello, Silvia Laura; Ferraccioli, Gianfranco

2011-01-01

The aim of the present study was to determine whether different subsets of B cells characterize synovial fluid (SF) or synovial tissue (ST) of seropositive or seronegative rheumatoid arthritis (RA) with respect to the peripheral blood (PB). PB, SF and ST of 14 autoantibody (AB)-positive (rheumatoid factor [RF]-IgM, RF-IgA, anti-citrullinated peptide [CCP]), 13 negative RA and 13 no-RA chronic arthritides were examined for B-cell subsets (Bm1-Bm5 and IgD-CD27 classifications), zeta-associated protein kinase-70 (ZAP70) expression on B cells and cytokine levels (interleukin [IL]-1β, tumor necrosis factor [TNF]-α, IL-6, IL-8 and monocyte chemotactic protein [MCP]-1). Synovial tissues were classified as aggregate and diffuse patterns. No differences were found in B-cell percentages or in subsets in PB and SF between AB(+) and AB(-) RA and no-RA. In both AB(+) and AB(-) RA (and no-RA), the percentage of CD19(+)/ZAP70(+) was higher in SF than in PB (AB(+): P = 0.03; AB(-): P = 0.01; no-RA: P = 0.01). Moreover, SF of both AB(+) and AB(-) RA (and no-RA) patients was characterized by a higher percentage of IgD-CD27(+) and IgD-CD27(-) B cells and lower percentage of IgD(+)CD27(-) (P < 0.05) B cells compared to PB. In SF, ZAP70 positivity is more represented in B cell CD27(+)/IgD(-)/CD38(-). The aggregate synovitis pattern was characterized by higher percentages of Bm5 cells in SF compared with the diffuse pattern (P = 0.05). These data suggest that no difference exists between AB(+) and AB(-) in B-cell subset compartmentalization. CD27(+)/IgD(-)/ZAP70(+) memory B cells accumulate preferentially in the joints of RA, suggesting a dynamic maturation of the B cells in this compartment.

The Anti-Tumorigenic Mushroom Agaricus blazei Murill Enhances IL-1β Production and Activates the NLRP3 Inflammasome in Human Macrophages

PubMed Central

Huang, Tsung-Teng; Ojcius, David M.; Young, John D.; Wu, Yi-Hui; Ko, Yun-Fei; Wong, Tsui-Yin; Wu, Cheng-Yeu; Lu, Chia-Chen; Lai, Hsin-Chih

2012-01-01

Agaricus blazei Murill (AbM) has been reported to possess immune activity against tumors and infections through stimulation of mononuclear phagocytes. Recently, AbM extract was shown to induce the production of the pro-inflammatory cytokine, interleukin-1β (IL-1β), in human monocytes. IL-1β is a key pro-inflammatory cytokine produced by activated macrophages and monocytes and its secretion is strictly controlled by the inflammasome. The purpose of this study is to investigate the effect of AbM water extracts on the regulation of IL-1β production and activation of the NLRP3 inflammasome in human THP-1 macrophages. The NLRP3 inflammasome consists of an NLRP3 receptor, an adaptor protein called ASC, and the inflammatory protease, caspase-1. Typically, stimulation of immune cells with microbial products results in production of pro-IL-1β, but a second stress-related signal activates the inflammasome and caspase-1, leading to processing and secretion of IL-1β. Our results show that AbM enhances transcription of IL-1β and triggers NLRP3 inflammasome-mediated IL-1β secretion in human THP-1 macrophages. AbM-mediated IL-1β secretion was markedly reduced in macrophages deficient in NLRP3 and ASC, demonstrating that the NLRP3 inflammasome is essential for AbM-induced IL-1β secretion. In addition, caspase-1 was activated and involved in proteolytic cleavage and secretion of IL-1β in AbM-treated macrophages. AbM-mediated IL-1β secretion also decreased in cells treated with cathepsin B inhibitor, suggesting that AbM can induce the release of cathepsin B. Furthermore, our data show that AbM-induced inflammasome activation requires the release of ATP, binding of extracellular ATP to the purinergic receptor P2X7, the generation of reactive oxygen species, and efflux of potassium. Taken together, these findings reveal that AbM activates the NLRP3 inflammasome via multiple mechanisms, resulting in the secretion of IL-1β. PMID:22844468

The anti-tumorigenic mushroom Agaricus blazei Murill enhances IL-1β production and activates the NLRP3 inflammasome in human macrophages.

PubMed

Huang, Tsung-Teng; Ojcius, David M; Young, John D; Wu, Yi-Hui; Ko, Yun-Fei; Wong, Tsui-Yin; Wu, Cheng-Yeu; Lu, Chia-Chen; Lai, Hsin-Chih

2012-01-01

Agaricus blazei Murill (AbM) has been reported to possess immune activity against tumors and infections through stimulation of mononuclear phagocytes. Recently, AbM extract was shown to induce the production of the pro-inflammatory cytokine, interleukin-1β (IL-1β), in human monocytes. IL-1β is a key pro-inflammatory cytokine produced by activated macrophages and monocytes and its secretion is strictly controlled by the inflammasome. The purpose of this study is to investigate the effect of AbM water extracts on the regulation of IL-1β production and activation of the NLRP3 inflammasome in human THP-1 macrophages. The NLRP3 inflammasome consists of an NLRP3 receptor, an adaptor protein called ASC, and the inflammatory protease, caspase-1. Typically, stimulation of immune cells with microbial products results in production of pro-IL-1β, but a second stress-related signal activates the inflammasome and caspase-1, leading to processing and secretion of IL-1β. Our results show that AbM enhances transcription of IL-1β and triggers NLRP3 inflammasome-mediated IL-1β secretion in human THP-1 macrophages. AbM-mediated IL-1β secretion was markedly reduced in macrophages deficient in NLRP3 and ASC, demonstrating that the NLRP3 inflammasome is essential for AbM-induced IL-1β secretion. In addition, caspase-1 was activated and involved in proteolytic cleavage and secretion of IL-1β in AbM-treated macrophages. AbM-mediated IL-1β secretion also decreased in cells treated with cathepsin B inhibitor, suggesting that AbM can induce the release of cathepsin B. Furthermore, our data show that AbM-induced inflammasome activation requires the release of ATP, binding of extracellular ATP to the purinergic receptor P2X(7), the generation of reactive oxygen species, and efflux of potassium. Taken together, these findings reveal that AbM activates the NLRP3 inflammasome via multiple mechanisms, resulting in the secretion of IL-1β.

A new link between transcriptional initiation and pre-mRNA splicing: The RNA binding histone variant H2A.B

PubMed Central

Hart-Smith, Gene; Tay, Ying Jin; Tng, Wei-Quan; Wilkins, Marc; Ryan, Daniel

2017-01-01

The replacement of histone H2A with its variant forms is critical for regulating all aspects of genome organisation and function. The histone variant H2A.B appeared late in evolution and is most highly expressed in the testis followed by the brain in mammals. This raises the question of what new function(s) H2A.B might impart to chromatin in these important tissues. We have immunoprecipitated the mouse orthologue of H2A.B, H2A.B.3 (H2A.Lap1), from testis chromatin and found this variant to be associated with RNA processing factors and RNA Polymerase (Pol) II. Most interestingly, many of these interactions with H2A.B.3 (Sf3b155, Spt6, DDX39A and RNA Pol II) were inhibited by the presence of endogenous RNA. This histone variant can bind to RNA directly in vitro and in vivo, and associates with mRNA at intron—exon boundaries. This suggests that the ability of H2A.B to bind to RNA negatively regulates its capacity to bind to these factors (Sf3b155, Spt6, DDX39A and RNA Pol II). Unexpectedly, H2A.B.3 forms highly decompacted nuclear subdomains of active chromatin that co-localizes with splicing speckles in male germ cells. H2A.B.3 ChIP-Seq experiments revealed a unique chromatin organization at active genes being not only enriched at the transcription start site (TSS), but also at the beginning of the gene body (but being excluded from the +1 nucleosome) compared to the end of the gene. We also uncover a general histone variant replacement process whereby H2A.B.3 replaces H2A.Z at intron-exon boundaries in the testis and the brain, which positively correlates with expression and exon inclusion. Taken together, we propose that a special mechanism of splicing may occur in the testis and brain whereby H2A.B.3 recruits RNA processing factors from splicing speckles to active genes following its replacement of H2A.Z. PMID:28234895

Immunogenicity and therapeutic effects of recombinant Ag85AB fusion protein vaccines in mice infected with Mycobacterium tuberculosis.

PubMed

Liang, Yan; Zhang, Junxian; Yang, Yourong; Bai, Xuejuan; Yu, Qi; Li, Ning; Hou, Ying; Shi, Yingchang; Wang, Lan; Wu, Xueqiong

2017-07-13

The immune function of tuberculosis (TB) patients is disordered. By using immune regulators to assist chemotherapy for TB the curative effect might be improved. In this study, a vaccine containing Mycobacterium tuberculosis (M. tuberculosis) recombinant Ag85AB fusion protein (rAg85AB) was constructed and evaluated. The mice were immunized intramuscularly three times at two-week intervals with Ag85AB fusion protein combined with Corynebacterium parvum adjuvant (rAg85AB+CP). In comparison to control mice that received either CP alone or saline, the mice that received rAg85AB+CP had significantly higher number of T cells secreting IFN-γ and higher levels of specific antibodies of IgG, IgG1 and IgG2a isotypes in sera. The specific antibodies also had higher ratios of IgG2a to IgG1, indicating a predominant Th1 immune response. To test for immunotherapy of TB, M. tuberculosis infected mice were given three intramuscular doses of 20μg, 40μg or 60μg of rAg85AB in rAg85AB+CP, or phosphate-buffered saline (PBS), or CP or Mycobacterium phlei (M. Phlei) F.U.36. Compared with the PBS group, 20µg, 40µg and 60µg rAg85AB+CP and M. phlei F.U.36 groups reduced the pulmonary bacterial loads by 0.13, 0.15, 0.42 and 0.40 log 10 , and the liver bacterial loads by 0.64, 0.64, 0.53 and 0.61 log 10 , respectively. Pathological changes of lungs were less, and the lesions were limited to a certain extent in 40µg and 60µg rAg85AB+CP and M. phlei F.U.36 groups. These results showed that rAg85AB+CP had immunotherapeutic effect on TB, significantly increasing the cellular immune response, and inhibiting the growth of M. tuberculosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hepatitis B-core antibody positive donors in liver transplantation and their impact on graft survival: evidence from the Liver Match cohort study.

PubMed

Angelico, Mario; Nardi, Alessandra; Marianelli, Tania; Caccamo, Lucio; Romagnoli, Renato; Tisone, Giuseppe; Pinna, Antonio D; Avolio, Alfonso W; Fagiuoli, Stefano; Burra, Patrizia; Strazzabosco, Mario; Costa, Alessandro Nanni

2013-04-01

The appropriate allocation of grafts from HBcAb positive donors in liver transplantation is crucial, yet a consensus is still lacking. We evaluated this issue within Liver Match, a prospective observational Italian study. Data from 1437 consecutive, first transplants performed in 2007-2009 using grafts from deceased heart beating donors were analyzed (median follow-up: 1040 days). Of these, 219 (15.2%) were HBcAb positive. Sixty-six HBcAb positive grafts were allocated to HBsAg positive and 153 to HBsAg negative recipients. 329 graft losses occurred (22.9%): 66 (30.1%) among 219 recipients of HBcAb positive grafts, and 263 (21.6%) among 1218 recipients of HBcAb negative grafts. Graft survival was lower in recipients of HBcAb positive compared to HBcAb negative donors, with unadjusted 3-year graft survival of 0.69 (s.e. 0.032) and 0.77 (0.013), respectively (log-rank, p=0.0047). After stratifying for recipient HBsAg status, this difference was only observed among HBsAg negative recipients (log rank, p=0.0007), 3-year graft survival being excellent (0.88, s.e. 0.020) among HBsAg positive recipients, regardless of the HBcAb donor status (log rank, p=0.4478). Graft loss due to de novo HBV hepatitis occurred only in one patient. At Cox regression, hazard ratios for graft loss were: MELD (1.30 per 10 units, p=0.0002), donor HBcAb positivity (1.56, p=0.0015), recipient HBsAg positivity (0.43, p <0.0001), portal vein thrombosis (1.99, p=0.0156), and DRI (1.41 per unit, p=0.0325). HBcAb positive donor grafts have better outcomes when transplanted into HBsAg positive than HBsAg negative recipients. These findings suggest that donor HBcAb positivity requires more stringent allocation strategies. Copyright © 2012 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Performance comparison of the 4th generation Bio-Rad Laboratories GS HIV Combo Ag/Ab EIA on the EVOLIS™ automated system versus Abbott ARCHITECT HIV Ag/Ab Combo, Ortho Anti-HIV 1+2 EIA on Vitros ECi and Siemens HIV-1/O/2 enhanced on Advia Centaur.

PubMed

Mitchell, Elizabeth O; Stewart, Greg; Bajzik, Olivier; Ferret, Mathieu; Bentsen, Christopher; Shriver, M Kathleen

2013-12-01

A multisite study was conducted to evaluate the performance of the Bio-Rad 4th generation GS HIV Combo Ag/Ab EIA versus Abbott 4th generation ARCHITECT HIV Ag/Ab Combo. The performance of two 3rd generation EIAs, Ortho Diagnostics Anti-HIV 1+2 EIA and Siemens HIV 1/O/2 was also evaluated. Study objective was comparison of analytical HIV-1 p24 antigen detection, sensitivity in HIV-1 seroconversion panels, specificity in blood donors and two HIV false reactive panels. Analytical sensitivity was evaluated with International HIV-1 p24 antigen standards, the AFFSAPS (pg/mL) and WHO 90/636 (IU/mL) standards; sensitivity in acute infection was compared on 55 seroconversion samples, and specificity was evaluated on 1000 negative blood donors and two false reactive panels. GS HIV Combo Ag/Ab demonstrated better analytical HIV antigen sensitivity compared to ARCHITECT HIV Ag/Ab Combo: 0.41 IU/mL versus 1.2 IU/mL (WHO) and 12.7 pg/mL versus 20.1 pg/mL (AFSSAPS); GS HIV Combo Ag/Ab EIA also demonstrated slightly better specificity compared to ARCHITECT HIV Ag/Ab Combo (100% versus 99.7%). The 4th generation HIV Combo tests detected seroconversion 7-11 days earlier than the 3rd generation HIV antibody only EIAs. Both 4th generation immunoassays demonstrated excellent performance in sensitivity, with the reduction of the serological window period (7-11 days earlier detection than the 3rd generation HIV tests). However, GS HIV Combo Ag/Ab demonstrated improved HIV antigen analytical sensitivity and slightly better specificity when compared to ARCHITECT HIV Ag/Ab Combo assay, with higher positive predictive values (PPV) for low prevalence populations. Copyright © 2013 Elsevier B.V. All rights reserved.

Military and academic programs outperform community programs on the American Board of Surgery Examinations.

PubMed

Falcone, John L; Charles, Anthony G

2013-01-01

There is a paucity of American Board of Surgery (ABS) Qualifying Examination (QE) and Certifying Examination (CE) outcomes comparing residency programs by academic, community, or military affiliation. We hypothesize that the larger academic programs will outperform the smaller community programs. In this retrospective study from 2002 to 2012, examination performance on the ABS QE and CE were obtained from the ABS for all of the general surgery residency programs. Programs were categorized by academic, community, and military affiliation. Both nonparametric and parametric statistics were used for comparison, using an α = 0.05. There were 137/235 (58.3%) academic programs, 90/235 (38.3%) community programs, and 8/235 (3.4%) military programs that satisfied inclusion criteria for this study. The Mann-Whitney U tests showed that the military programs outperformed academic and community programs on the ABS QE and the ABS CE, and had a higher proportion of examinees passing both examinations on the first attempt (all p≤0.02). One-tailed Student t-tests showed that academic programs had higher pass rates than community programs on the ABS QE (85.4%±9.5% vs. 81.9%±11.5%), higher pass rates on the ABS CE (83.6%±8.3% vs. 80.6%±11.0%), and a higher proportion of examinees passing both examinations on the first attempt (0.73±0.12 vs. 0.68±0.15) (all p≤0.01). The chi-square and Fisher exact tests showed that examinees performed highest in military programs, followed by academic programs, and lowest in community programs on the ABS QE and ABS CE (all p≤ 0.01). Military programs have the highest degrees of success on all of the ABS examinations. Academic programs outperform community programs. These results have the potential to affect application patterns to established general surgery residency programs. Copyright © 2013 Association of Program Directors in Surgery. Published by Elsevier Inc. All rights reserved.

Positivity rates of antithyroid antibody, antinuclear antibody and thyroid peroxidase antibody in different types of vitiligo.

PubMed

Lim, H K; Bae, M I; Jeong, K H; Shin, M K; Lee, M-H

2016-04-01

Vitiligo is associated with various autoimmune disorders, and organ-specific autoantibodies are frequently found in patients with this disorder. Vitiligo is classically divided into segmental vitiligo (SV) and nonsegmental vitiligo (NSV), and it is believed that the pathogenesis differs between these two types. As the NSV type is related to an autoimmune mechanism, autoantibody detection rates are likely to be higher in the NSV type than in the segmental type; however, no comparative studies have been performed. To analyse the rates of autoantibody positivity according to the clinical features in patients with vitiligo. Rates of antithyroid antibody (Tg Ab), antinuclear antibody (ANA) and thyroid peroxidase antibody (TPO Ab) positivity were analysed and compared according to the sex, clinical type and age of onset of 807 patients with vitiligo. There were 106 patients with SV (13.1%) and 701 patients with NSV (86.9%). Tg Ab and ANA positivity did not differ between the SV and NSV types. A positive TPO Ab result was obtained in 16 patients with SV (15.1%) and 173 patients with NSV (24.7%). The TPO Ab positivity rate was significantly higher in NSV (χ² = 4.14, P < 0.05). The positivity rates of the three autoantibodies differed significantly according to age of onset (P = 0.001, P = 0.02 and P < 0.001 for Tg Ab, ANA and TPO Ab positivity, respectively). The TPO Ab positivity rate also showed a sex difference (P < 0.001). The positivity rates for the three autoantibodies showed differences according to age of onset and sex. The rates of Tg Ab and ANA positivity showed no significant differences according to clinical type, but the TPO Ab positivity rate was significantly different between SV and NSV. It appears likely that an autoimmune mechanism contributes to the pathogenesis of SV. © 2015 British Association of Dermatologists.

Mapping of binding epitopes of a human decay-accelerating factor monoclonal antibody capable of enhancing rituximab-mediated complement-dependent cytotoxicity.

PubMed

Guo, Bo; Ma, Zheng-wei; Li, Hua; Xu, Gui-lian; Zheng, Ping; Zhu, Bo; Wu, Yu-Zhang; Zou, Qiang

2008-08-01

Complement-dependent cytotoxicity (CDC) is thought to be one of the most important mechanisms of action of therapeutic monoclonal antibodies (mAbs). The decay-accelerating factor (DAF) overexpressed in certain tumors limits the CDC effect of the therapeutic anticancer antibodies. The use of DAF blocking antibodies targeted specifically at cancer cells in combination with immunotherapeutic mAbs of cancer may improve the therapeutic effect in cancer patients. In this study, the lysis of Raji cells mediated by CDC was determined after blocking DAF function by anti-DAF polyclonal antibody and 3 mAbs (DG3, DG9, DA11) prepared in our laboratory, respectively, in the presence of the anti-CD20 chimeric mAb rituximab. The binding domains of the three anti-DAF mAbs were identified using yeast surface display technique, and the mimic epitopes of mAb DG3 were screened from a random phage-display nonapeptide library. The results showed that blocking DAF function by anti-DAF polyclonal antibody enhanced complement-mediated killing of Raji cells. Among the 3 mAbs against DAF, only DG3 was found to be able to remarkably enhance the CDC effect of the therapeutic mAb rituximab. DG3 bound to the third short consensus repeat (SCR) of DAF. Binding of DG3 to immobilized DAF was inhibited by mimic epitope peptides screened from the peptide library. Our results suggest that a higher level of DAF expressed by certain tumor cells is significant to abolish the CDC effect of therapeutic anticancer antibodies, and mAbs binding to SCR3 can enhance the complement-mediated killing of Raji cells. It is of significance to identify the DAF epitopes required in inhibiting CDC not only for better understanding of the relationship between the structure and function of DAF, but also for designing and developing anti-DAF mAbs capable of enhancing CDC.

Effects of Transcranial Direct Current Stimulation over Left Dorsolateral pFC on the Attentional Blink Depend on Individual Baseline Performance.

PubMed

London, Raquel E; Slagter, Heleen A

2015-12-01

Selection mechanisms that dynamically gate only relevant perceptual information for further processing and sustained representation in working memory are critical for goal-directed behavior. We examined whether this gating process can be modulated by anodal transcranial direct current stimulation (tDCS) over left dorsolateral pFC (DLPFC)--a region known to play a key role in working memory and conscious access. Specifically, we examined the effects of tDCS on the magnitude of the so-called "attentional blink" (AB), a deficit in identifying the second of two targets presented in rapid succession. Thirty-four participants performed a standard AB task before (baseline), during, and after 20 min of 1-mA anodal and cathodal tDCS in two separate sessions. On the basis of previous reports linking individual differences in AB magnitude to individual differences in DLPFC activity and on suggestions that effects of tDCS depend on baseline brain activity levels, we hypothesized that anodal tDCS over left DLPFC would modulate the magnitude of the AB as a function of individual baseline AB magnitude. Indeed, individual differences analyses revealed that anodal tDCS decreased the AB in participants with a large baseline AB but increased the AB in participants with a small baseline AB. This effect was only observed during (but not after) stimulation, was not found for cathodal tDCS, and could not be explained by regression to the mean. Notably, the effects of tDCS were not apparent at the group level, highlighting the importance of taking individual variability in performance into account when evaluating the effectiveness of tDCS. These findings support the idea that left DLPFC plays a critical role in the AB and in conscious access more generally. They are also in line with the notion that there is an optimal level of prefrontal activity for cognitive function, with both too little and too much activity hurting performance.

A cysteine protease (cathepsin Z) from disk abalone, Haliotis discus discus: Genomic characterization and transcriptional profiling during bacterial infections.

PubMed

Godahewa, G I; Perera, N C N; Lee, Sukkyoung; Kim, Myoung-Jin; Lee, Jehee

2017-09-05

Cathepsin Z (CTSZ) is lysosomal cysteine protease of the papain superfamily. It participates in the host immune defense via phagocytosis, signal transduction, cell-cell communication, proliferation, and migration of immune cells such as monocytes, macrophages, and dendritic cells. Hence, CTSZ is also acknowledged as an acute-phase protein in host immunity. In this study, we sought to identify the CTSZ homolog from disk abalone (AbCTSZ) and characterize it at the molecular, genomic, and transcriptional levels. AbCTSZ encodes a protein with 318 amino acids and a molecular mass of 36kDa. The structure of AbCTSZ reveals amino acid sequences that are characteristic of the signal sequence, pro-peptide, peptidase-C1 papain family cysteine protease domain, mini-loop, HIP motif, N-linked glycosylation sites, active sites, and conserved Cys residues. A pairwise comparison revealed that AbCTSZ shared the highest amino acid homology with its molluscan counterpart from Crassostrea gigas. A multiple alignment analysis revealed the conservation of functionally crucial elements of AbCTSZ, and a phylogenetic study further confirmed a proximal evolutionary relationship with its invertebrate counterparts. Further, an analysis of AbCTSZ genomic structure revealed seven exons separated by six introns, which differs from that of its vertebrate counterparts. Quantitative real time PCR (qPCR) detected the transcripts of AbCTSZ in early developmental stages and in eight different tissues. Higher levels of AbCTSZ transcripts were found in trochophore, gill, and hemocytes, highlighting its importance in the early development and immunity of disk abalone. In addition, we found that viable bacteria (Vibrio parahaemolyticus and Listeria monocytogenes) and bacterial lipopolysaccharides significantly modulated AbCTSZ transcription. Collectively, these lines of evidences suggest that AbCTSZ plays an indispensable role in the innate immunity of disk abalone. Copyright © 2017. Published by Elsevier B.V.

Asymptomatic bacteriuria in women with autoimmune rheumatic disease: prevalence, risk factors, and clinical significance.

PubMed

Georgiadou, Sarah P; Gamaletsou, Maria N; Mpanaka, Ioanna; Vlachou, Aggeliki; Goules, Andreas V; Ziogas, Dimitrios C; Syriou, Vassiliki; Tektonidou, Maria G; Kaltsas, Gregory; Manoussakis, Menelaos N; Sipsas, Nikolaos V

2015-03-15

Data regarding the prevalence and clinical significance of asymptomatic bacteriuria (AB) in women with autoimmune rheumatic disease (ARD) are scarce. In this prospective, case-control study, consecutive female outpatients with ARD were screened for AB. For each patient, demographics, type, duration, and treatment of underlying ARD, and risk factors for urinary tract infection (UTI), were recorded. Age-matched women with endocrine disease, without any autoimmune disease, not receiving immunosuppressive agents were used as controls. Subjects were followed up for 1 year for the development of symptomatic UTI. Two hundred sixty patients with ARD (mean age, 52.4 [standard deviation {SD}, 14.6] years) and 238 controls (mean age, 51.2 [SD, 16.5] years) were enrolled. The majority of patients with ARD (93.5%; 95% confidence interval [CI], 89.7%-95.9%) were receiving immunosuppressive agents. AB was detected in 24 patients with ARD (9.2%; 95% CI, 6.2%-13.4%) and in 22 controls (9.2%; 95% CI, 5.5%-12.9%) (P = 1.000). The most prevalent pathogen was Escherichia coli (16/24 [66%]). Independent predictors for AB among patients were diabetes (odds ratio [OR], 6.6; P = .008) and a longer ARD duration (>84 months; OR, 4.3; P = .018). During the 1-year follow-up, 9 patients with baseline AB remained persistently bacteriuric, whereas 11 were intermittently bacteriuric. Symptomatic UTI developed in 4 of 24 patients (16.7%; 95% CI, 6.1%-36.5%) with baseline AB vs 29 of 236 (12.3%; 95% CI, 8.6%-17.1%) without AB (P = .522). In our study, the prevalence of AB among women with ARD was not higher than that of controls, and AB was not associated with higher risk for symptomatic UTI. Risk factors for AB were longer duration of ARD and diabetes. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Optimal combinations of broadly neutralizing antibodies for prevention and treatments of HIV-1 clade C infection

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

Wagh, Kshitij; Bhattacharya, Tanmoy; Williamson, Carolyn

In this study, the identification of a new generation of potent broadly neutralizing HIV-1 antibodies (bnAbs) has generated substantial interest in their potential use for the prevention and/or treatment of HIV-1 infection. While combinations of bnAbs targeting distinct epitopes on the viral envelope (Env) will likely be required to overcome the extraordinary diversity of HIV-1, a key outstanding question is which bnAbs, and how many, will be needed to achieve optimal clinical benefit. We assessed the neutralizing activity of 15 bnAbs targeting four distinct epitopes of Env, including the CD4-binding site (CD4bs), the V1/V2-glycan region, the V3-glycan region, and themore » gp41 membrane proximal external region (MPER), against a panel of 200 acute/early clade C HIV-1 Env pseudoviruses. A mathematical model was developed that predicted neutralization by a subset of experimentally evaluated bnAb combinations with high accuracy. Using this model, we performed a comprehensive and systematic comparison of the predicted neutralizing activity of over 1,600 possible double, triple, and quadruple bnAb combinations. The most promising bnAb combinations were identified based not only on breadth and potency of neutralization, but also other relevant measures, such as the extent of complete neutralization and instantaneous inhibitory potential (IIP). By this set of criteria, triple and quadruple combinations of bnAbs were identified that were significantly more effective than the best double combinations, and further improved the probability of having multiple bnAbs simultaneously active against a given virus, a requirement that may be critical for countering escape in vivo. These results provide a rationale for advancing bnAb combinations with the best in vitro predictors of success into clinical trials for both the prevention and treatment of HIV-1 infection.« less

Guided bone regeneration with local zoledronic acid and titanium barrier: An experimental study.

PubMed

Dundar, Serkan; Ozgur, Cem; Yaman, Ferhan; Cakmak, Omer; Saybak, Arif; Ozercan, Ibrahim Hanifi; Alan, Hilal; Artas, Gokhan; Nacakgedigi, Onur

2016-10-01

The aim of this study was to evaluate the effects on new bone formation of autogenous blood alone or in combination with zoledronic acid (ZA), a β-tricalcium phosphate (β-TCP) graft or ZA plus a β-TCP graft placed under titanium barriers. For this purpose, eight adult male New Zealand white rabbits were used in the study, each with four titanium barriers fixed around four sets of nine holes drilled in the calvarial bones. The study included four groups, each containing 2 rabbits. In the autogenous blood (AB group), only autogeneous blood was placed under the titanium barriers. The three experimental groups were the AB+ZA group, with autogenous blood plus ZA, the AB+β-TCP group, with autogeneous blood plus a β-TCP graft, and the AB+β-TCP+ZA group, with autogeneous blood plus a β-TCP graft and ZA mixture under the titanium barriers. The animals were sacrificed after 3 months. The amounts of new bone formation identified histomorphometrically were found to be higher after 3 months than at the time of surgery in all groups. The differences between the groups were examined with histomorphometric analysis, and statistically significant differences were identified at the end of the 3 months. The bone formation rate in the AB+β-TCP+ZA group was determined to be significantly higher than that in the other groups (P<0.05). In the AB+ZA and AB+β-TCP groups, the bone formation rate was determined to be significantly higher than that in the AB group (P<0.05). No statistically significant difference in bone formation rate was observed between the AB+β-TCP and AB+ZA groups. Local ZA used with autogeneous blood and/or graft material appears to be a more effective method than the use of autogeneous blood or graft alone in bone augmentation executed with a titanium barrier.

Stellar Origin of {sup 15}N-rich Presolar SiC Grains of Type AB: Supernovae with Explosive Hydrogen Burning

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

Liu, Nan; Nittler, Larry R.; Alexander, Conel M. O’D.

We report C, N, and Si isotopic data for 59 highly {sup 13}C-enriched presolar submicron- to micron-sized SiC grains from the Murchison meteorite, including eight putative nova grains (PNGs) and 29 {sup 15}N-rich ({sup 14}N/{sup 15}N ≤ solar) AB grains, and their Mg–Al, S, and Ca–Ti isotope data when available. These 37 grains are enriched in {sup 13}C, {sup 15}N, and {sup 26}Al with the PNGs showing more extreme enhancements. The {sup 15}N-rich AB grains show systematically higher {sup 26}Al and {sup 30}Si excesses than the {sup 14}N-rich AB grains. Thus, we propose to divide the AB grains into groupsmore » 1 ({sup 14}N/{sup 15}N < solar) and 2 ({sup 14}N/{sup 15}N ≥ solar). For the first time, we have obtained both S and Ti isotopic data for five AB1 grains and one PNG and found {sup 32}S and/or {sup 50}Ti enhancements. Interestingly, one AB1 grain had the largest {sup 32}S and {sup 50}Ti excesses, strongly suggesting a neutron-capture nucleosynthetic origin of the {sup 32}S excess and thus the initial presence of radiogenic {sup 32}Si ( t {sub 1/2} = 153 years). More importantly, we found that the {sup 15}N and {sup 26}Al excesses of AB1 grains form a trend that extends to the region in the N–Al isotope plot occupied by C2 grains, strongly indicating a common stellar origin for both AB1 and C2 grains. Comparison of supernova models with the AB1 and C2 grain data indicates that these grains came from supernovae that experienced H ingestion into the He/C zones of their progenitors.« less

Quantum calculations of the IR spectrum of liquid water using ab initio and model potential and dipole moment surfaces and comparison with experiment.

PubMed

Liu, Hanchao; Wang, Yimin; Bowman, Joel M

2015-05-21

The calculation and characterization of the IR spectrum of liquid water have remained a challenge for theory. In this paper, we address this challenge using a combination of ab initio approaches, namely, a quantum treatment of IR spectrum using the ab initio WHBB water potential energy surface and a refined ab initio dipole moment surface. The quantum treatment is based on the embedded local monomer method, in which the three intramolecular modes of each embedded H2O monomer are fully coupled and also coupled singly to each of six intermolecular modes. The new dipole moment surface consists of a previous spectroscopically accurate 1-body dipole moment surface and a newly fitted ab initio intrinsic 2-body dipole moment. A detailed analysis of the new dipole moment surface in terms of the coordinate dependence of the effective atomic charges is done along with tests of it for the water dimer and prism hexamer double-harmonic spectra against direct ab initio calculations. The liquid configurations are taken from previous molecular dynamics calculations of Skinner and co-workers, using the TIP4P plus E3B rigid monomer water potential. The IR spectrum of water at 300 K in the range of 0-4000 cm(-1) is calculated and compared with experiment, using the ab initio WHBB potential and new ab initio dipole moment, the q-TIP4P/F potential, which has a fixed-charged description of the dipole moment, and the TTM3-F potential and dipole moment surfaces. The newly calculated ab initio spectrum is in very good agreement with experiment throughout the above spectral range, both in band positions and intensities. This contrasts to results with the other potentials and dipole moments, especially the fixed-charge q-TIP4P/F model, which gives unrealistic intensities. The calculated ab initio spectrum is analyzed by examining the contribution of various transitions to each band.

Presence, location, type and size of denuded areas of subchondral bone in the knee as a function of radiographic stage of OA - data from the OA initiative.

PubMed

Frobell, R B; Wirth, W; Nevitt, M; Wyman, B T; Benichou, O; Dreher, D; Davies, R Y; Lee, J H; Baribaud, F; Gimona, A; Hudelmaier, M; Cotofana, S; Eckstein, F

2010-05-01

To assess the presence, location, type and size of denuded areas of subchondral bone (dAB) in the femorotibial joint, measured quantitatively with 3T MRI, in a large subset of OAI participants. One knee of 633 subjects (250 men, 383 women, aged 61.7+/-9.6 y) were studied, spanning all radiographic osteoarthritis (OA) stages. dABs were determined quantitatively using segmentations of coronal FLASHwe images, representing areas where the subchondral bone was not covered by cartilage. Post hoc visual examination of segmented images determined whether dABs represented full thickness cartilage loss or internal osteophyte. 7% Of the knees were Kellgren & Lawrence (KL) grade 0, 6% grade 1, 41% grade 2, 41% grade 3, and 5% grade 4. 39% Of the participants (48% of the men and 33% of the women) displayed dABs; 61% of the dABs represented internal osteophytes. 1/47 Participants with KL grade 0 displayed 'any' dAB whereas 29/32 of the KL grade 4 knees were affected. Even as early as KL grade 1, 29% of the participants showed dABs. There were significant relationships of dAB with increasing KL grades (P<0.001) and with ipsi-compartimental JSN (P< or =0.001). Internal osteophytes were more frequent laterally (mainly posterior tibia and internal femur) whereas full thickness cartilage loss was more frequent medially (mainly external tibia and femur). dABs occur already at earliest stages of radiographic OA (KL grades 1 and 2) and become more common (and larger) with increasing disease severity. Almost all KL grade 4 knees exhibited dABs, with cartilage loss being more frequent than internal osteophytes. Copyright 2010 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Sclerostin Antibody Treatment Enhances Rotator Cuff Tendon-to-Bone Healing in an Animal Model.

PubMed

Shah, Shivam A; Kormpakis, Ioannis; Havlioglu, Necat; Ominsky, Michael S; Galatz, Leesa M; Thomopoulos, Stavros

2017-05-17

Rotator cuff tears are a common source of pain and disability, and poor healing after repair leads to high retear rates. Bone loss in the humeral head before and after repair has been associated with poor healing. The purpose of the current study was to mitigate bone loss near the repaired cuff and improve healing outcomes. Sclerostin antibody (Scl-Ab) treatment, previously shown to increase bone formation and strength in the setting of osteoporosis, was used in the current study to address bone loss and enhance rotator cuff healing in an animal model. Scl-Ab was administered subcutaneously at the time of rotator cuff repair and every 2 weeks until the animals were sacrificed. The effect of Scl-Ab treatment was evaluated after 2, 4, and 8 weeks of healing, using bone morphometric analysis, biomechanical evaluation, histological analysis, and gene expression outcomes. Injury and repair led to a reduction in bone mineral density after 2 and 4 weeks of healing in the control and Scl-Ab treatment groups. After 8 weeks of healing, animals receiving Scl-Ab treatment had 30% greater bone mineral density than the controls. A decrease in biomechanical properties was observed in both groups after 4 weeks of healing compared with healthy tendon-to-bone attachments. After 8 weeks of healing, Scl-Ab-treated animals had improved strength (38%) and stiffness (43%) compared with control animals. Histological assessment showed that Scl-Ab promoted better integration of tendon and bone by 8 weeks of healing. Scl-Ab had significant effects on gene expression in bone, indicative of enhanced bone formation, and no effect on the expression of genes in tendon. This study provides evidence that Scl-Ab treatment improves tendon-to-bone healing at the rotator cuff by increasing attachment-site bone mineral density, leading to improved biomechanical properties. Scl-Ab treatment may improve outcomes after rotator cuff repair.

Immune recognition of botulinum neurotoxin B: antibody-binding regions on the heavy chain of the toxin.

PubMed

Dolimbek, Behzod Z; Steward, Lance E; Aoki, K Roger; Atassi, M Zouhair

2008-02-01

The purpose of this work was to map the continuous regions recognized by human, horse and mouse anti-botulinum neurotoxin B (BoNT/B) antibodies (Abs). We synthesized a panel of sixty 19-residue peptides (peptide C31 was 24 residues) that overlapped consecutively by 5 residues and together encompassed the entire heavy chain of BoNT/B (H/B, residues 442-1291). Abs from the three host species recognized similar, but not identical, peptides. There were also peptides recognized by two or only by one host species. Where a peptide was recognized by Abs of more than one host species, these Abs were at different levels among the species. Human, horse and mouse Abs bound, although in different amounts, to regions within peptides 736-754, 778-796, 848-866, 932-950, 974-992, 1058-1076 and 1128-1146. Human and horse Abs bound to peptides 890-908 and 1170-1188. Human and mouse Abs recognized peptides 470-488/484-502 overlap, 638-656, 722-740, 862-880, 1030-1048, 1072-1090, 1240-1258 and 1268-1291. We concluded that the antigenic regions localized with the three antisera are quite similar, exhibiting in some cases a small shift to the left or to the right. This is consistent with what is known about protein immune recognition. In the three-dimensional structure, the regions recognized on H/B by anti-BoNT/B Abs occupied surface locations and analysis revealed no correlation between these surface locations and surface electrostatic potential, hydrophilicity, hydrophobicity, or temperature factor. A region that bound mouse Abs overlapped with a recently defined site on BoNT/B that binds to mouse and rat synaptotagmin II, thus providing a molecular explanation for the blocking (protecting) activity of these Abs. The regions thus localized afford candidates for incorporation into a synthetic vaccine design.

Optimal combinations of broadly neutralizing antibodies for prevention and treatments of HIV-1 clade C infection

DOE PAGES

Wagh, Kshitij; Bhattacharya, Tanmoy; Williamson, Carolyn; ...

2016-03-30

In this study, the identification of a new generation of potent broadly neutralizing HIV-1 antibodies (bnAbs) has generated substantial interest in their potential use for the prevention and/or treatment of HIV-1 infection. While combinations of bnAbs targeting distinct epitopes on the viral envelope (Env) will likely be required to overcome the extraordinary diversity of HIV-1, a key outstanding question is which bnAbs, and how many, will be needed to achieve optimal clinical benefit. We assessed the neutralizing activity of 15 bnAbs targeting four distinct epitopes of Env, including the CD4-binding site (CD4bs), the V1/V2-glycan region, the V3-glycan region, and themore » gp41 membrane proximal external region (MPER), against a panel of 200 acute/early clade C HIV-1 Env pseudoviruses. A mathematical model was developed that predicted neutralization by a subset of experimentally evaluated bnAb combinations with high accuracy. Using this model, we performed a comprehensive and systematic comparison of the predicted neutralizing activity of over 1,600 possible double, triple, and quadruple bnAb combinations. The most promising bnAb combinations were identified based not only on breadth and potency of neutralization, but also other relevant measures, such as the extent of complete neutralization and instantaneous inhibitory potential (IIP). By this set of criteria, triple and quadruple combinations of bnAbs were identified that were significantly more effective than the best double combinations, and further improved the probability of having multiple bnAbs simultaneously active against a given virus, a requirement that may be critical for countering escape in vivo. These results provide a rationale for advancing bnAb combinations with the best in vitro predictors of success into clinical trials for both the prevention and treatment of HIV-1 infection.« less

Quantum calculations of the IR spectrum of liquid water using ab initio and model potential and dipole moment surfaces and comparison with experiment

NASA Astrophysics Data System (ADS)

Liu, Hanchao; Wang, Yimin; Bowman, Joel M.

2015-05-01

The calculation and characterization of the IR spectrum of liquid water have remained a challenge for theory. In this paper, we address this challenge using a combination of ab initio approaches, namely, a quantum treatment of IR spectrum using the ab initio WHBB water potential energy surface and a refined ab initio dipole moment surface. The quantum treatment is based on the embedded local monomer method, in which the three intramolecular modes of each embedded H2O monomer are fully coupled and also coupled singly to each of six intermolecular modes. The new dipole moment surface consists of a previous spectroscopically accurate 1-body dipole moment surface and a newly fitted ab initio intrinsic 2-body dipole moment. A detailed analysis of the new dipole moment surface in terms of the coordinate dependence of the effective atomic charges is done along with tests of it for the water dimer and prism hexamer double-harmonic spectra against direct ab initio calculations. The liquid configurations are taken from previous molecular dynamics calculations of Skinner and co-workers, using the TIP4P plus E3B rigid monomer water potential. The IR spectrum of water at 300 K in the range of 0-4000 cm-1 is calculated and compared with experiment, using the ab initio WHBB potential and new ab initio dipole moment, the q-TIP4P/F potential, which has a fixed-charged description of the dipole moment, and the TTM3-F potential and dipole moment surfaces. The newly calculated ab initio spectrum is in very good agreement with experiment throughout the above spectral range, both in band positions and intensities. This contrasts to results with the other potentials and dipole moments, especially the fixed-charge q-TIP4P/F model, which gives unrealistic intensities. The calculated ab initio spectrum is analyzed by examining the contribution of various transitions to each band.

Evaluation of a fully automated treponemal test and comparison with conventional VDRL and FTA-ABS tests.

PubMed

Park, Yongjung; Park, Younhee; Joo, Shin Young; Park, Myoung Hee; Kim, Hyon-Suk

2011-11-01

We evaluated analytic performances of an automated treponemal test and compared this test with the Venereal Disease Research Laboratory test (VDRL) and fluorescent treponemal antibody absorption test (FTA-ABS). Precision performance of the Architect Syphilis TP assay (TP; Abbott Japan, Tokyo, Japan) was assessed, and 150 serum samples were assayed with the TP before and after heat inactivation to estimate the effect of heat inactivation. A total of 616 specimens were tested with the FTA-ABS and TP, and 400 were examined with the VDRL. The TP showed good precision performance with total imprecision of less than a 10% coefficient of variation. An excellent linear relationship between results before and after heat inactivation was observed (R(2) = 0.9961). The FTA-ABS and TP agreed well with a κ coefficient of 0.981. The concordance rate between the FTA-ABS and TP was the highest (99.0%), followed by the rates between FTA-ABS and VDRL (85.0%) and between TP and VDRL (83.8%). The automated TP assay may be adequate for screening for syphilis in a large volume of samples and can be an alternative to FTA-ABS.

Attention Bias Modification (ABM): Review of Effects of Multisession ABM Training on Anxiety and Threat-Related Attention in High-Anxious Individuals

PubMed Central

Mogg, Karin; Waters, Allison M.; Bradley, Brendan P.

2017-01-01

Attention bias modification (ABM) aims to reduce anxiety by reducing attention bias (AB) to threat; however, effects on anxiety and AB are variable. This review examines 34 studies assessing effects of multisession-ABM on both anxiety and AB in high-anxious individuals. Methods include ABM-threat-avoidance (promoting attention-orienting away from threat), ABM-positive-search (promoting explicit, goal-directed attention-search for positive/nonthreat targets among negative/threat distractors), and comparison conditions (e.g., control-attention training combining threat-cue exposure and attention-task practice without AB-modification). Findings indicate anxiety reduction often occurs during both ABM-threat-avoidance and control-attention training; anxiety reduction is not consistently accompanied by AB reduction; anxious individuals often show no pretraining AB in orienting toward threat; and ABM-positive-search training appears promising in reducing anxiety. Methodological and theoretical issues are discussed concerning ABM paradigms, comparison conditions, and AB assessment. ABM methods combining explicit goal-directed attention-search for nonthreat/positive information and effortful threat-distractor inhibition (promoting top-down cognitive control during threat-cue exposure) warrant further evaluation. PMID:28752017

Evaluation of protection by two endotoxin-neutralizing IgM monoclonal antibodies in different peritonitis models.

PubMed

Hustinx, W N; Benaissa-Trouw, B J; Harmsen, T; Klein, S; Verhoef, J; Hoepelman, A I; Kraaijeveld, K

1997-10-01

Two anti-core glycolipid (CGL) IgM monoclonal antibodies (mAbs 8-2 and 26-20), previously shown to display cross-reactivity with heterologous lipopolysaccharide (LPS) in vitro and to provide cross-protectivity against endotoxin challenge in vivo, were evaluated for their potential to protect mice against death from peritonitis caused by heterologous bacterial challenge. Without concurrent antibiotic treatment neither antibody was protective. Compared with a control mAb, prophylactic treatment with mAb 8-2 significantly increased the survival of gentamicin-treated mice challenged with the rough strain Salmonella minnesota Re595. Both mAb 8-2 and a control mAb, in combination with a suboptimal dose of ceftazidime, increased survival following challenge with the clinical isolate Escherichia coli O7:K1. In a model of mucin-enhanced peritonitis, neither mAb was protective against challenge with inocula of E. coli O7:K1, ranging from 10(2) to 10(4) bacteria. We conclude that protection of mice by anti-CGL mAb 8-2 against heterologous challenge is vitally dependent on concurrent treatment with antibiotics and that protection may not be attributable to the anti-CGL specificity of these antibodies.

Attitudes towards antimicrobial drugs among general population in Croatia, Fyrom, Greece, Hungary, Serbia and Slovenia.

PubMed

Radosević, Nives; Vlahović-Palcevski, Vera; Benko, Ria; Peklar, Jure; Miskulin, Inka; Matuz, Maria; Papaioannidou, Paraskevi; Sabo, Ana; Palcevska-Koceska, Snezana

2009-08-01

A pilot study to assess patients' attitudes towards antimicrobials (ABs) in six European countries (Croatia, Former Yugoslav Republic of Macedonia (FYROM), Greece, Hungary, Slovenia and Serbia), as a step preceding educational intervention on the importance of patients' compliance with instructions on taking ABs and consequences of their inappropriate use. Patients' knowledge, emotions and behaviour regarding ABs were assessed using a structured questionnaire, constructed by a psychologist and intended for general population in six European countries. Questionnaires were filled out by individuals who visited pharmacies and general practitioners. A total of 838 questionnaires were filled in. Respondents from Slovenia showed the best knowledge about ABs, followed by Croatians. The highest willingness for self-medication reported respondents from FYROM. The most positive emotions about ABs were expressed by respondents in Greece and Hungary, and the most negative in Slovenia. All components of attitudes towards antibiotics were influenced by country and level of education. Behaviour regarding ABs complied with emotions and knowledge in all countries. The results of this study may lay a basis for conducting national public campaigns, as a step forward in education of patients on rational AB use. Copyright 2009 John Wiley & Sons, Ltd.

The content of compound conditioning.

PubMed

Harris, Justin A; Andrew, Benjamin J; Livesey, Evan J

2012-04-01

In three experiments using Pavlovian conditioning of magazine approach, rats were trained with a compound stimulus, AB, and were concurrently trained with stimulus B on its own. The reinforcement rate of B, rB, was either 1/2, 2/3, or 2/5 of rAB. After extended training, the conditioning strength of A was assessed using probe trials in which A was presented alone. Responding during A was compared with that during AB, B, and a third stimulus, C, for which rC = rAB - rB. In each experiment, the rats' response rate during A was almost identical to that during C (and during B, when rB = 1/2rAB). This suggests that, during AB conditioning, the rats had learned about rA as being equal to [rAB - rB], and implies that the content of their learning was a linear function of r. The findings provide strong support for rate-based models of conditioning (e.g., Gallistel & Gibbon, 2000). They are also consistent with the associative account of learning defined in the Rescorla and Wagner (1972) model, but only if the learning rate during reinforcement equals that during nonreinforcement. (c) 2012 APA, all rights reserved.

Natural additives to preserve quality and improve nutritional value of fresh-cut nectarine.

PubMed

Falagán, Natalia; Artés, Francisco; Aguayo, Encarna

2016-07-01

There is currently a high demand for natural and fresh-cut fruits. In this study, natural additives were applied to fresh-cut nectarines: (1) control, nontreated; (2) wedges were dipped in an antibrowning solution containing calcium ascorbate (AB); (3) and (4) wedges were dipped in an AB plus natural antimicrobial agents (vanillin or cinnamic acid, respectively). After these treatments, fresh-cut nectarines were packed and stored at 5 ℃ for eight days. The treatments AB+Vanillin and AB+Cinnamic inhibited microbial counts when compared with control and AB-only samples. The application of these solutions did not impart any aromas or off-flavors to nectarines and maintained firmness during the shelf-life period. AB solutions inhibited polyophenol oxidase action and reduced browning while stabilizing the soluble phenolic content, increasing consumer's acceptance. Nectarine wedges assimilated the ascorbic acid from the AB solution and retained it during the shelf-life period. The combination of an antibrowning agent and natural antimicrobials helped to control microbiological growth while maintaining high-quality parameters. They can be an attractive "green" alternative for organic fresh-cut products to other chemical sanitizers such as chlorine. © The Author(s) 2015.

Bridging the gap: facilities and technologies for development of early stage therapeutic mAb candidates.

PubMed

Munro, Trent P; Mahler, Stephen M; Huang, Edwin P; Chin, David Y; Gray, Peter P

2011-01-01

Therapeutic monoclonal antibodies (mAbs) currently dominate the biologics marketplace. Development of a new therapeutic mAb candidate is a complex, multistep process and early stages of development typically begin in an academic research environment. Recently, a number of facilities and initiatives have been launched to aid researchers along this difficult path and facilitate progression of the next mAb blockbuster. Complementing this, there has been a renewed interest from the pharmaceutical industry to reconnect with academia in order to boost dwindling pipelines and encourage innovation. In this review, we examine the steps required to take a therapeutic mAb from discovery through early stage preclinical development and toward becoming a feasible clinical candidate. Discussion of the technologies used for mAb discovery, production in mammalian cells and innovations in single-use bioprocessing is included. We also examine regulatory requirements for product quality and characterization that should be considered at the earliest stages of mAb development. We provide details on the facilities available to help researchers and small-biotech build value into early stage product development, and include examples from within our own facility of how technologies are utilized and an analysis of our client base.

mAbs

PubMed Central

2009-01-01

The twenty two monoclonal antibodies (mAbs) currently marketed in the U.S. have captured almost half of the top-20 U.S. therapeutic biotechnology sales for 2007. Eight of these products have annual sales each of more than $1 B, were developed in the relatively short average period of six years, qualified for FDA programs designed to accelerate drug approval, and their cost has been reimbursed liberally by payers. With growth of the product class driven primarily by advancements in protein engineering and the low probability of generic threats, mAbs are now the largest class of biological therapies under development. The high cost of these drugs and the lack of generic competition conflict with a financially stressed health system, setting reimbursement by payers as the major limiting factor to growth. Advances in mAb engineering are likely to result in more effective mAb drugs and an expansion of the therapeutic indications covered by the class. The parallel development of biomarkers for identifying the patient subpopulations most likely to respond to treatment may lead to a more cost-effective use of these drugs. To achieve the success of the current top-tier mAbs, companies developing new mAb products must adapt to a significantly more challenging commercial environment. PMID:20061824

46 CFR 175.600 - Incorporation by reference.

Code of Federal Regulations, 2011 CFR

2011-10-01

... Classing Steel Vessels, 1995 (“ABS Steel Vessel Rules”) 183.360. Rules for Building and Classing Steel Vessels Under 61 Meters (200 feet) in Length, 1983 (“ABS Steel Vessel Rules (Steel Vessels for Service on Rivers and Intracoastal Waterways, 1995 (“ABS...

46 CFR 175.600 - Incorporation by reference.

Code of Federal Regulations, 2012 CFR

2012-10-01

... Classing Steel Vessels, 1995 (“ABS Steel Vessel Rules”) 183.360. Rules for Building and Classing Steel Vessels Under 61 Meters (200 feet) in Length, 1983 (“ABS Steel Vessel Rules (Steel Vessels for Service on Rivers and Intracoastal Waterways, 1995 (“ABS...

75 FR 57778 - Issuance of an Experimental Use Permit

Federal Register 2010, 2011, 2012, 2013, 2014

2010-09-22

...) necessary for its production in Event TC1507 corn. There are 10.213 kilograms (kg) of eCry3.1Ab, 11.14 grams of Cry1Ab, 8,145 grams of Cry34Ab1, 112.1 grams of Cry35Ab1, 280.6 grams of Vip3Aa20, 217.9 grams of mCry3A, and 177.5 grams of Cry1F proteins in 69,473 kg seeds on 7,311 acres (3,830 PIP and 3,481 non...

Single-ion 4f element magnetism: an ab-initio look at Ln(COT)2(-).

PubMed

Gendron, Frédéric; Pritchard, Benjamin; Bolvin, Hélène; Autschbach, Jochen

2015-12-14

The electron densities associated with the Ln 4f shell, and spin and orbital magnetizations ('magnetic moment densities'), are investigated for the Ln(COT)2(-) series. The densities are obtained from ab-initio calculations including spin-orbit coupling. For Ln = Ce, Pr the magnetizations are also derived from crystal field models and shown to agree with the ab-initio results. Analysis of magnetizations from ab-initio calculations may be useful in assisting research on single molecule magnets.

[Find your way in the jungle of mAbs].

PubMed

Watier, H

2017-09-01

The rapidly increasing number of approved monoclonal antibodies (mAbs) and the huge number of mAbs in clinical development are a matter of concern for who wants to easily identify targets, indications, mechanisms of action and possible adverse effects. The current nomenclature being of limited interest, simple rationales will be presented for helping practitioners in rapidly classify mAbs depending on their structure-pharmacology relationship and in evaluating their potential effects, particularly in transfusion medicine. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Prostate Specific Expression of Maspin in a Transgenic Mouse Model: Implications in Prostate Carcinogenesis

DTIC Science & Technology

2004-05-01

should not be construed as an official Department of the Army position, policy or decision unless so designated by other documentation. 20050603 257 r...such material. Where material from documents designated for limited distribution is quoted, permission has been obtained to use the material...amino acid peptide located in the reactive site loop of maspin, a region designated as AbS4A. The antibody was purified using an AbS4A sulfo-linked

[Kidney allotransplantation from the AB0-incompatible donors].

PubMed

Goriaĭnov, V A; Kaabak, M M; Babenko, N N; Shishlo, L A; Morozova, M M; Ragimov, A A; Dazhkova, N G; Salimov, E L

2013-01-01

The experience of 28 kidney allotransplantations from the AB0-incompatible donors was analyzed. The comparative group consisted of 38 patients, who received the AB0-compatible organ. The results were assessed using the following parameters: renal function, morphology of the biopsy samples of the transplanted kidney and actuary survival of the recipients with functioning transplants in both groups. The comparative analysis showed no significant difference between the two groups, giving the right to consider the kidney allotransplantation from the AB0-incompatible donors safe and effective.

Clinical laboratory verification of thyroglobulin concentrations in the presence of autoantibodies to thyroglobulin: comparison of EIA, radioimmunoassay and LC MS/MS measurements in an Urban Hospital.

PubMed

Wheeler, Sarah E; Liu, Li; Blair, Harry C; Sivak, Richard; Longo, Nancy; Tischler, Jeffery; Mulvey, Kathryn; Palmer, Octavia M Peck

2017-12-08

Thyroglobulin (Tg) measurements assess recurrence in post-thyroidectomy thyroid cancer patients. Tg measurements by enzyme immunoassays (EIA) can be falsely elevated by interference from Tg autoantibodies (TgAb). Radioimmunoassay (RIA) is less susceptible to TgAb interference and has been the standard-of-care test for TgAb positive patients. Recently developed liquid chromatography tandem mass spectrometry (LC-MS/MS) methods may eliminate TgAb interference. We assessed the performance of Tg measurements by EIA, RIA and LC-MS/MS to evaluate TgAb interference differences. We measured TgAb and Tg in 50 plasma samples from 40 patients in whom Tg measurement was part of their routine follow-up and 10 healthy volunteers. Discrepancy between EIA and both LC-MS/MS and RIA was observed at low Tg concentrations (≤ 7.55 ng/mL) in TgAb positive specimens (LC-MS/MS = 1.9 * EIA - 0.03, r = 0.68). RIA and LC-MS/MS Tg measurements in TgAb positive specimens with low Tg concentrations had improved correlation but demonstrated bias (LC MS/MS = 0.6 * RIA - 1.4, r = 0.90). Disagreement between methods may be attributed to LC-MS/MS reported Tg concentrations as undetectable compared to RIA. It seems likely that most discrepant cases are falsely elevated in RIA due to TgAb interference, however, some cases appear below the detection limit of LC-MS/MS; implementation of LC-MS/MS by clinicians will require lower detection limits.

Individual Differences in Temporal Selective Attention as Reflected in Pupil Dilation.

PubMed

Willems, Charlotte; Herdzin, Johannes; Martens, Sander

2015-01-01

Attention is restricted for the second of two targets when it is presented within 200-500 ms of the first target. This attentional blink (AB) phenomenon allows one to study the dynamics of temporal selective attention by varying the interval between the two targets (T1 and T2). Whereas the AB has long been considered as a robust and universal cognitive limitation, several studies have demonstrated that AB task performance greatly differs between individuals, with some individuals showing no AB whatsoever. Here, we studied these individual differences in AB task performance in relation to differences in attentional timing. Furthermore, we investigated whether AB magnitude is predictive for the amount of attention allocated to T1. For both these purposes pupil dilation was measured, and analyzed with our recently developed deconvolution method. We found that the dynamics of temporal attention in small versus large blinkers differ in a number of ways. Individuals with a relatively small AB magnitude seem better able to preserve temporal order information. In addition, they are quicker to allocate attention to both T1 and T2 than large blinkers. Although a popular explanation of the AB is that it is caused by an unnecessary overinvestment of attention allocated to T1, a more complex picture emerged from our data, suggesting that this may depend on whether one is a small or a large blinker. The use of pupil dilation deconvolution seems to be a powerful approach to study the temporal dynamics of attention, bringing us a step closer to understanding the elusive nature of the AB. We conclude that the timing of attention to targets may be more important than the amount of allocated attention in accounting for individual differences.

Clinical performance of the novel DiaSorin LIAISON(®) XL murex: HBsAg Quant, HCV-Ab, HIV-Ab/Ag assays.

PubMed

Krawczyk, Adalbert; Hintze, Christian; Ackermann, Jessica; Goitowski, Birgit; Trippler, Martin; Grüner, Nico; Neumann-Fraune, Maria; Verheyen, Jens; Fiedler, Melanie

2014-01-01

The fully automated and closed LIAISON(®)XL platform was developed for reliable detection of infection markers like hepatitis B virus (HBV) surface antigen (HBsAg), hepatitis C virus (HCV) antibodies (Ab) or human immunodeficiency virus (HIV)-Ag/Ab. To date, less is known about the diagnostic performance of this system in direct comparison to the common Abbott ARCHITECT(®) platform. We compared the diagnostic performance and usability of the DiaSorin LIAISON(®)XL with the commonly used Abbott ARCHITECT(®) system. The qualitative performance of the above mentioned assays was compared in about 500 sera. Quantitative tests were performed for HBsAg-positive samples from patients under therapy (n=289) and in vitro expressed mutants (n=37). For HCV-Ab, a total number of 155 selected samples from patients chronically infected with different HCV genotypes were tested. The concordance between both systems was 99.4% for HBsAg, 98.81% for HCV-Ab, and 99.6% for HIV-Ab/Ag. The quantitative LIAISON(®)XL murex HBsAg assay detected all mutants in comparable amounts to the HBsAg wild type and yielded highly reliable HBsAg kinetics in patients treated with antiviral drugs. Dilution experiments using the 2nd International Standard for HBsAg (WHO) showed a high accuracy of this test. HCV-Ab from patients infected with genotypes 1-3 were equally detected in both systems. Interestingly, S/CO levels of HCV-Ab from patients infected with genotype 3 seem to be relatively low using both systems. The LIAISON(®)XL platform proved to be an excellent system for diagnostics of HBV, HCV, and HIV with equal performance compared to the ARCHITECT(®) system. Copyright © 2013 Elsevier B.V. All rights reserved.

Bmi-1-targeting suppresses osteosarcoma aggressiveness through the NF-κB signaling pathway

PubMed Central

Liu, Jiaguo; Luo, Bin; Zhao, Meng

2017-01-01

Bone cancer is one of the most lethal malignancies and the specific causes of tumor initiation are not well understood. B-cell-specific Moloney murine leukemia virus integration site 1 protein (Bmi-1) has been reported to be associated with the initiation and progression of osteosarcoma, and as a prognostic indicator in the clinic. In the current study, a full-length antibody targeting Bmi-1 (AbBmi-1) was produced and the preclinical value of Bmi-1-targeted therapy was evaluated in bone carcinoma cells and tumor xenograft mice. The results indicated that the Bmi-1 expression level was markedly upregulated in bone cancer cell lines, and inhibition of Bmi-1 by AbBmi-1 reduced the invasiveness and migration of osteosarcoma cells. Overexpression of Bmi-1 promoted proliferation and angiogenesis, and increased apoptosis resistance induced by cisplatin via the nuclear factor-κB (NF-κB) signal pathway. In addition, AbBmi-1 treatment inhibited the tumorigenicity of osteosarcoma cells in vivo. Furthermore, AbBmi-1 blocked NF-κB signaling and reduced MMP-9 expression. Furthermore, Bmi-1 promoted osteosarcoma tumor growth, whereas AbBmi-1 significantly inhibited osteosarcoma tumor growth in vitro and in vivo. Notably, AbBmi-1 decreased the percentages of Ki67-positive cells and terminal deoxynucleotidyl transferase dUTP nick end labeling-positive cells in tumors compared with Bmi-1-treated and PBS controls. Notably, MMP-9 and NF-κB expression were downregulated by treatment with AbBmi-1 in MG-63 osteosarcoma cells. In conclusion, the data provides evidence that AbBmi-1 inhibited the progression of osteosarcoma, suggesting that AbBmi-1 may be a novel anti-cancer agent through the inhibition of Bmi-1 via activating the NF-κB pathway in osteosarcoma. PMID:28983587

Ethanolic Extract of Agaricus blazei Fermentation Product Inhibits the Growth and Invasion of Human Hepatoma HA22T/VGH and SK-Hep-1 Cells

PubMed Central

Tung, Yen-Chen; Su, Zheng-Yuan; Kuo, Min-Liang; Sheen, Lee-Yan

2012-01-01

Hepatoma is a leading cause of death in the world. SK-Hep-1 and HA22T/VGH cells are poorly differentiated human hepatocellular carcinoma cell lines with invasive and migratory abilities. Agaricus blazei (AB) is a mushroom with many biological effects and active ingredients, and the ethanolic extract of AB fermentation product (AB-pE) was demonstrated to inhibit the growth of hepatoma Hep3B and HepG2 cells in our previous study. In this study, we further investigated the anticancer and anti-invasive abctivities of the AB-pE. Results showed that the AB-pE inhibited the growth of SK-Hep1 and HA22T/VGH cells (with IC50 values of 26.8 and 28.7 μg/mL, respectively) and led cells toward apoptosis after 48 h of treatment. Activation of caspase-3 by AB-pE (12.5~200 μg/mL) in a dose-dependent manner was observed in both cell lines using fluorescence microscopy and flow cytometry. The apoptosis triggered by the AB-pE was regulated by the increased expression of Bax, the activation of caspase-3, caspase-9, and PARP, and the decreased expression of Bcl-2. Additionally, the AB-pE showed the potential ability to inhibit invasion of SK-Hep1 and HA22T/VGH cells according to the results of a Matrigel invasion assay. Our results suggested that the AB-pE may be a further developed for its potential against hepatoma due to its antiproliferative (via apoptosis) and anti-invasive activities in hepatoma cells. PMID:24716127

The interplay of non-specific binding, target-mediated clearance and FcRn interactions on the pharmacokinetics of humanized antibodies

PubMed Central

Datta-Mannan, Amita; Lu, Jirong; Witcher, Derrick R; Leung, Donmienne; Tang, Ying; Wroblewski, Victor J

2015-01-01

The application of protein engineering technologies toward successfully improving antibody pharmacokinetics has been challenging due to the multiplicity of biochemical factors that influence monoclonal antibody (mAb) disposition in vivo. Physiological factors including interactions with the neonatal Fc receptor (FcRn) and specific antigen binding properties of mAbs, along with biophysical properties of the mAbs themselves play a critical role. It has become evident that applying an integrated approach to understand the relative contribution of these factors is critical to rationally guide and apply engineering strategies to optimize mAb pharmacokinetics. The study presented here evaluated the influence of unintended non-specific interactions on the disposition of mAbs whose clearance rates are governed predominantly by either non-specific (FcRn) or target-mediated processes. The pharmacokinetics of 8 mAbs representing a diverse range of these properties was evaluated in cynomolgus monkeys. Results revealed complementarity-determining region (CDR) charge patch engineering to decrease charge-related non-specific binding can have a significant impact on improving the clearance. In contrast, the influence of enhanced in vitro FcRn binding was mixed, and related to both the strength of charge interaction and the general mechanism predominant in governing the clearance of the particular mAb. Overall, improved pharmacokinetics through enhanced FcRn interactions were apparent for a CDR charge-patch normalized mAb which was affected by non-specific clearance. The findings in this report are an important demonstration that mAb pharmacokinetics requires optimization on a case-by-case basis to improve the design of molecules with increased therapeutic application. PMID:26337808

Adaptive optics imaging of healthy and abnormal regions of retinal nerve fiber bundles of patients with glaucoma.

PubMed

Chen, Monica F; Chui, Toco Y P; Alhadeff, Paula; Rosen, Richard B; Ritch, Robert; Dubra, Alfredo; Hood, Donald C

2015-01-08

To better understand the nature of glaucomatous damage of the macula, especially the structural changes seen between relatively healthy and clearly abnormal (AB) retinal regions, using an adaptive optics scanning light ophthalmoscope (AO-SLO). Adaptive optics SLO images and optical coherence tomography (OCT) vertical line scans were obtained on one eye of seven glaucoma patients, with relatively deep local arcuate defects on the 10-2 visual field test in one (six eyes) or both hemifields (one eye). Based on the OCT images, the retinal nerve fiber (RNF) layer was divided into two regions: (1) within normal limits (WNL), relative RNF layer thickness within mean control values ±2 SD; and (2) AB, relative thickness less than -2 SD value. As seen on AO-SLO, the pattern of AB RNF bundles near the border of the WNL and AB regions differed across eyes. There were normal-appearing bundles in the WNL region of all eyes and AB-appearing bundles near the border with the AB region. This region with AB bundles ranged in extent from a few bundles to the entire AB region in the case of one eye. All other eyes had a large AB region without bundles. However, in two of these eyes, a few bundles were seen within this region of otherwise missing bundles. The AO-SLO images revealed details of glaucomatous damage that are difficult, if not impossible, to see with current OCT technology. Adaptive optics SLO may prove useful in following progression in clinical trials, or in disease management, if AO-SLO becomes widely available and easy to use. Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

Role of EfrAB efflux pump in biocide tolerance and antibiotic resistance of Enterococcus faecalis and Enterococcus faecium isolated from traditional fermented foods and the effect of EDTA as EfrAB inhibitor.

PubMed

Lavilla Lerma, Leyre; Benomar, Nabil; Valenzuela, Antonio Sánchez; Casado Muñoz, María del Carmen; Gálvez, Antonio; Abriouel, Hikmate

2014-12-01

Enterococcus faecalis and Enterococcus faecium isolated from various traditional fermented foods of both animal and vegetable origins have shown multidrug resistance to several antibiotics and tolerance to biocides. Reduced susceptibility was intra and inter-species dependent and was due to specific and unspecific mechanisms such as efflux pumps. EfrAB, a heterodimeric ABC transporter efflux pump, was detected in 100% of multidrug resistant (MDR) E. faecalis strains and only in 12% of MDR E. faecium strains. EfrAB expression was induced by half of minimum inhibitory concentration (MIC) of gentamicin, streptomycin and chloramphenicol. However, expression of efrA and efrB genes was highly dependent on the strain tested and on the antimicrobial used. Our results indicated that 3 mM EDTA highly reduced the MICs of almost all drugs tested. Nevertheless, the higher reductions (>8 folds) were obtained with gentamicin, streptomycin, chlorhexidine and triclosan. Reductions of MICs were correlated with down-regulation of EfrAB expression (10-140 folds) in all three MDR enterococci strains. This is the first report describing the role of EfrAB in the efflux of antibiotics and biocides which reflect also the importance of EfrAB in multidrug resistance in enterococci. EDTA used at low concentration as food preservative could be one of the best choices to prevent spread of multidrug resistant enterococci throughout food chain by decreasing EfrAB expression. EfrAB could be an attractive target not only in enterococci present in food matrix but also those causing infections as well by using EDTA as therapeutic agent in combination with low doses of antibiotics. Copyright © 2014 Elsevier Ltd. All rights reserved.

Antibody Response Specific to the Capsular Polysaccharide Is Impaired in Streptococcus suis Serotype 2-Infected Animals

PubMed Central

Calzas, Cynthia; Lemire, Paul; Auray, Gael; Gerdts, Volker; Gottschalk, Marcelo

2014-01-01

Streptococcus suis serotype 2 is an extracellular encapsulated bacterium that causes severe septicemia and meningitis in swine and humans. Albeit crucial in the fight against encapsulated bacteria, the nature of the capsular polysaccharide (CPS)-specific antibody (Ab) response during S. suis type 2 infection is unknown. We compared for the first time the features of CPS-specific versus protein-specific Ab responses during experimental infections with live virulent S. suis type 2 in mice. The primary protein-specific Ab response was dominated by both type 1 and 2 IgG subclasses, whereas IgM titers were more modest. The secondary protein-specific Ab response showed all of the features of a memory response with faster kinetics and boosted the titers of all Ig isotypes. In contrast, the primary CPS-specific Ab response was either inexistent or had titers only slightly higher than those in noninfected animals and was essentially composed of IgM. A poor CPS-specific memory response was observed, with only a moderate boost in IgM titers and no IgG. Both protein- and CPS-specific Ab responses were Toll-like receptor 2 independent. By using S. suis type 2 strains of European or North American origin, the poor CPS-specific Ab response was demonstrated to be independent of the genotypic/phenotypic diversity of the strain within serotype 2. Finally, the CPS-specific Ab response was also impaired and lacked isotype switching in S. suis-infected pigs, the natural host of the bacterium. The better resistance of preinfected animals to reinfection with the same strain of S. suis type 2 might thus more likely be related to the development of a protein rather than CPS Ab response. PMID:25385801

Anti-neuropeptide Y plasma immunoglobulins in relation to mood and appetite in depressive disorder.

PubMed

Garcia, Frederico D; Coquerel, Quentin; do Rego, Jean-Claude; Cravezic, Aurore; Bole-Feysot, Christine; Kiive, Evelyn; Déchelotte, Pierre; Harro, Jaanus; Fetissov, Sergueï O

2012-09-01

Depression and eating disorders are frequently associated, but the molecular pathways responsible for co-occurrence of altered mood, appetite and body weight are not yet fully understood. Neuropeptide Y (NPY) has potent antidepressant and orexigenic properties and low central NPY levels have been reported in major depression. In the present study, we hypothesized that in patients with major depression alteration of mood, appetite and body weight may be related to NPY-reactive autoantibodies (autoAbs). To test this hypothesis, we compared plasma levels and affinities of NPY-reactive autoAbs between patients with major depression and healthy controls. Then, to evaluate if changes of NPY autoAb properties can be causally related to altered mood and appetite, we developed central and peripheral passive transfer models of human autoAbs in mice and studied depressive-like behavior in forced-swim test and food intake. We found that plasma levels of NPY IgG autoAbs were lower in patients with moderate but not with mild depression correlating negatively with the Montgomery-Åsberg Depression Rating Scale scores and with immobility time of the forced-swim test in mice after peripheral injection of autoAbs. No significant differences in NPY IgG autoAb affinities between patients with depression and controls were found, but higher affinity of IgG autoAbs for NPY was associated with lower body mass index and prevented NPY-induced orexigenic response in mice after their central injection. These data suggest that changes of plasma levels of anti-NPY autoAbs are relevant to altered mood, while changes of their affinity may participate in altered appetite and body weight in patients with depressive disorder. Copyright © 2012 Elsevier Ltd. All rights reserved.

Differences in incidence and co-occurrence of vaccine and nonvaccine human papillomavirus types in Finnish population before human papillomavirus mass vaccination suggest competitive advantage for HPV33.

PubMed

Merikukka, Marko; Kaasila, Marjo; Namujju, Proscovia B; Palmroth, Johanna; Kirnbauer, Reinhard; Paavonen, Jorma; Surcel, Heljä-Marja; Lehtinen, Matti

2011-03-01

To understand likelihood of type replacement after vaccination against the high-risk human papillomavirus (HPV) types, we evaluated competition of the seven most common genital HPV types in a population sample of unvaccinated, fertile-aged Finnish women. First trimester sera from two consecutive pregnancies were retrieved from 3,183 Finnish women (mean age, 23.1 years) of whom 42.3% had antibodies to at least one HPV type (6/11/16/18/31/33/45) at the baseline. Antibody positivity to more than one HPV types by the second pregnancy was common among the baseline HPV seropositives. However, compared to baseline HPV-seronegative women, significantly increased incidence rate ratios (IRRs), indicating an increased risk to seroconvert for another HPV type, were consistently noted only for HPV33 among baseline HPV16 or HPV18 antibody (ab)-positive women: HPV(16ab only) (→) (16&33ab) IRR 2.9 [95% confidence interval (CI) 1.6-5.4] and HPV(18ab only) (→) (18&33ab) IRR 2.5 (95% CI 1.1-6.0), irrespectively of the presence of antibodies to other HPV types at baseline: HPV(16ab) (→) (16&33ab) IRR 3.2 (95% CI 2.0-5.2) and HPV(18ab) (→) (18&33ab) IRR 3.6 (95% CI 2.1-5.9). Our findings suggest a possible competitive advantage for HPV33 over other genital HPV types in the unvaccinated population. HPV33 should be monitored for type replacement after HPV mass vaccination. Copyright © 2010 UICC.

The Novel Bacterial N-Demethylase PdmAB Is Responsible for the Initial Step of N,N-Dimethyl-Substituted Phenylurea Herbicide Degradation

PubMed Central

Gu, Tao; Zhou, Chaoyang; Sørensen, Sebastian R.; Zhang, Ji; He, Jian; Yu, Peiwen; Li, Shunpeng

2013-01-01

The environmental fate of phenylurea herbicides has received considerable attention in recent decades. The microbial metabolism of N,N-dimethyl-substituted phenylurea herbicides can generally be initiated by mono-N-demethylation. In this study, the molecular basis for this process was revealed. The pdmAB genes in Sphingobium sp. strain YBL2 were shown to be responsible for the initial mono-N-demethylation of commonly used N,N-dimethyl-substituted phenylurea herbicides. PdmAB is the oxygenase component of a bacterial Rieske non-heme iron oxygenase (RO) system. The genes pdmAB, encoding the α subunit PdmA and the β subunit PdmB, are organized in a transposable element flanked by two direct repeats of an insertion element resembling ISRh1. Furthermore, this transposable element is highly conserved among phenylurea herbicide-degrading sphingomonads originating from different areas of the world. However, there was no evidence of a gene for an electron carrier (a ferredoxin or a reductase) located in the immediate vicinity of pdmAB. Without its cognate electron transport components, expression of PdmAB in Escherichia coli, Pseudomonas putida, and other sphingomonads resulted in a functional enzyme. Moreover, coexpression of a putative [3Fe-4S]-type ferredoxin from Sphingomonas sp. strain RW1 greatly enhanced the catalytic activity of PdmAB in E. coli. These data suggested that PdmAB has a low specificity for electron transport components and that its optimal ferredoxin may be the [3Fe-4S] type. PdmA exhibited low homology to the α subunits of previously characterized ROs (less than 37% identity) and did not cluster with the RO group involved in O- or N-demethylation reactions, indicating that PdmAB is a distinct bacterial RO N-demethylase. PMID:24123738

Uncoupling the widespread occurrence of anti-NMDAR1 autoantibodies from neuropsychiatric disease in a novel autoimmune model.

PubMed

Pan, Hong; Oliveira, Bárbara; Saher, Gesine; Dere, Ekrem; Tapken, Daniel; Mitjans, Marina; Seidel, Jan; Wesolowski, Janina; Wakhloo, Debia; Klein-Schmidt, Christina; Ronnenberg, Anja; Schwabe, Kerstin; Trippe, Ralf; Mätz-Rensing, Kerstin; Berghoff, Stefan; Al-Krinawe, Yazeed; Martens, Henrik; Begemann, Martin; Stöcker, Winfried; Kaup, Franz-Josef; Mischke, Reinhard; Boretius, Susann; Nave, Klaus-Armin; Krauss, Joachim K; Hollmann, Michael; Lühder, Fred; Ehrenreich, Hannelore

2018-02-09

Autoantibodies of the IgG class against N-methyl-D-aspartate-receptor subunit-NR1 (NMDAR1-AB) were considered pathognomonic for anti-NMDAR encephalitis. This view has been challenged by the age-dependent seroprevalence (up to >20%) of functional NMDAR1-AB of all immunoglobulin classes found in >5000 individuals, healthy or affected by different diseases. These findings question a merely encephalitogenic role of NMDAR1-AB. Here, we show that NMDAR1-AB belong to the normal autoimmune repertoire of dogs, cats, rats, mice, baboons, and rhesus macaques, and are functional in the NMDAR1 internalization assay based on human IPSC-derived cortical neurons. The age dependence of seroprevalence is lost in nonhuman primates in captivity and in human migrants, raising the intriguing possibility that chronic life stress may be related to NMDAR1-AB formation, predominantly of the IgA class. Active immunization of ApoE -/- and ApoE +/+ mice against four peptides of the extracellular NMDAR1 domain or ovalbumin (control) leads to high circulating levels of specific AB. After 4 weeks, the endogenously formed NMDAR1-AB (IgG) induce psychosis-like symptoms upon MK-801 challenge in ApoE -/- mice, characterized by an open blood-brain barrier, but not in their ApoE +/+ littermates, which are indistinguishable from ovalbumin controls. Importantly, NMDAR1-AB do not induce any sign of inflammation in the brain. Immunohistochemical staining for microglial activation markers and T lymphocytes in the hippocampus yields comparable results in ApoE -/- and ApoE +/+ mice, irrespective of immunization against NMDAR1 or ovalbumin. These data suggest that NMDAR1-AB of the IgG class shape behavioral phenotypes upon access to the brain but do not cause brain inflammation on their own.

The fate and transport of the Cry1Ab protein in an agricultural field and laboratory aquatic microcosms.

PubMed

Strain, Katherine E; Lydy, Michael J

2015-08-01

Genetically engineered crops expressing insecticidal crystalline proteins derived from Bacillus thuringiensis (Bt), were commercialized almost two decades ago as a means to manage agricultural pests. The Bt proteins are highly specific and only lethal upon ingestion, limiting the scope of toxicity to target insects. However, concern of exposure to non-target organisms and negative public perceptions regarding Bt crops has caused controversy surrounding their use. The objective of this research was to monitor the fate and transport of a Bt protein, Cry1Ab, in a large-scale agricultural field containing maize expressing the Cry1Ab protein and a non-Bt near isoline, and in aquatic microcosms. The highest environmental concentrations of the Cry1Ab protein were found in runoff water and sediment, up to 130ngL(-1) and 143ngg(-1) dry weight, respectively, with the Cry1Ab protein detected in both Bt and non-Bt maize fields. As surface runoff and residual crop debris can transport Bt proteins to waterways adjacent to agricultural fields, a series of laboratory experiments were conducted to determine the potential fate of the Cry1Ab protein under different conditions. The results showed that sediment type and temperature can influence the degradation of the Cry1Ab protein in an aquatic system and that the Cry1Ab protein can persist for up to two months. Although Cry1Ab protein concentrations measured in the field soil indicate little exposure to terrestrial organisms, the consistent input of Bt-contaminated runoff and crop debris into agricultural waterways is relevant to understanding potential consequences to aquatic species. Copyright © 2015 Elsevier Ltd. All rights reserved.

Erythrocyte Saturation with IgG Is Required for Inducing Antibody-Mediated Immune Suppression and Impacts Both Erythrocyte Clearance and Antigen-Modulation Mechanisms.

PubMed

Cruz-Leal, Yoelys; Marjoram, Danielle; Lazarus, Alan H

2018-02-15

Anti-D prevents hemolytic disease of the fetus and newborn, and this mechanism has been referred to as Ab-mediated immune suppression (AMIS). Anti-D, as well as other polyclonal AMIS-inducing Abs, most often induce both epitope masking and erythrocyte clearance mechanisms. We have previously observed that some Abs that successfully induce AMIS effects could be split into those that mediate epitope masking versus those that induce erythrocyte clearance, allowing the ability to analyze these mechanisms separately. In addition, AMIS-inducing activity has recently been shown to induce Ag modulation (Ag loss from the erythrocyte surface). To assess these mechanisms, we immunized mice with transgenic murine RBCs expressing a single Ag protein comprising a recombinant Ag composed of hen egg lysozyme, OVA sequences comprising aa 251-349, and the human Duffy transmembrane protein (HOD-Ag) with serial doses of polyclonal anti-OVA IgG as the AMIS-inducing Ab. The anti-OVA Ab induced AMIS in the absence of apparent epitope masking. AMIS occurred only when the erythrocytes appeared saturated with IgG. This Ab was capable of inducing HOD-RBC clearance, as well as loss of the OVA epitope at doses of Ab that caused AMIS effects. HOD-RBCs also lost reactivity with Abs specific for the hen egg lysozyme and Duffy portions of the Ag consistent with the initiation of Ag modulation and/or trogocytosis mechanisms. These data support the concept that an AMIS-inducing Ab that does not cause epitope masking can induce AMIS effects in a manner consistent with RBC clearance and/or Ag modulation. Copyright © 2018 by The American Association of Immunologists, Inc.

Conformational Changes of Blood ACE in Chronic Uremia

PubMed Central

Petrov, Maxim N.; Shilo, Valery Y.; Tarasov, Alexandr V.; Schwartz, David E.; Garcia, Joe G. N.; Kost, Olga A.; Danilov, Sergei M.

2012-01-01

Background The pattern of binding of monoclonal antibodies (mAbs) to 16 epitopes on human angiotensin I-converting enzyme (ACE) comprise a conformational ACE fingerprint and is a sensitive marker of subtle protein conformational changes. Hypothesis Toxic substances in the blood of patients with uremia due to End Stage Renal Disease (ESRD) can induce local conformational changes in the ACE protein globule and alter the efficacy of ACE inhibitors. Methodology/Principal Findings The recognition of ACE by 16 mAbs to the epitopes on the N and C domains of ACE was estimated using an immune-capture enzymatic plate precipitation assay. The precipitation pattern of blood ACE by a set of mAbs was substantially influenced by the presence of ACE inhibitors with the most dramatic local conformational change noted in the N-domain region recognized by mAb 1G12. The “short” ACE inhibitor enalaprilat (tripeptide analog) and “long” inhibitor teprotide (nonapeptide) produced strikingly different mAb 1G12 binding with enalaprilat strongly increasing mAb 1G12 binding and teprotide decreasing binding. Reduction in S-S bonds via glutathione and dithiothreitol treatment increased 1G12 binding to blood ACE in a manner comparable to enalaprilat. Some patients with uremia due to ESRD exhibited significantly increased mAb 1G12 binding to blood ACE and increased ACE activity towards angiotensin I accompanied by reduced ACE inhibition by inhibitory mAbs and ACE inhibitors. Conclusions/Significance The estimation of relative mAb 1G12 binding to blood ACE detects a subpopulation of ESRD patients with conformationally changed ACE, which activity is less suppressible by ACE inhibitors. This parameter may potentially serve as a biomarker for those patients who may need higher concentrations of ACE inhibitors upon anti-hypertensive therapy. PMID:23166630

MicroRNA-20a/b regulates cholesterol efflux through post-transcriptional repression of ATP-binding cassette transporter A1.

PubMed

Liang, Bin; Wang, Xin; Song, Xiaosu; Bai, Rui; Yang, Huiyu; Yang, Zhiming; Xiao, Chuanshi; Bian, Yunfei

2017-09-01

ATP-binding cassette transporter A1 (ABCA1) plays a crucial role in reverse cholesterol transport and exhibits anti-atherosclerosis effects. Some microRNAs (miRs) regulate ABCA1 expression, and recent studies have shown that miR-20a/b might play a critical role in atherosclerotic diseases. Here, we attempted to clarify the potential contribution of miR-20a/b in post-transcriptional regulation of ABCA1, cholesterol efflux, and atherosclerosis. We performed bioinformatics analysis and found that miR-20a/b was highly conserved and directly bound to ABCA1 mRNA with low binding free energy. Luciferase-reporter assay also confirmed that miR-20a/b significantly reduced luciferase activity associated with the ABCA1 3' untranslated region reporter construct. Additionally, miR-20a/b decreased ABCA1 expression, which, in turn, decreased cholesterol efflux and increased cholesterol content in THP-1 and RAW 264.7 macrophage-derived foam cells. In contrast, miR-20a/b inhibitors increased ABCA1 expression and cholesterol efflux, decreased cholesterol content, and inhibited foam-cell formation. Consistent with our in vitro results, miR-20a/b-treated ApoE -/- mice showed decreased ABCA1expression in the liver and reductions of reverse cholesterol transport in vivo. Furthermore, miR-20a/b regulated the formation of nascent high-density lipoprotein and promoted atherosclerotic development, whereas miR-20a/b knockdown attenuated atherosclerotic formation. miR-20 is a new miRNA capable of targeting ABCA1 and regulating ABCA1 expression. Therefore, miR-20 inhibition constitutes a new strategy for ABCA1-based treatment of atherosclerosis. Copyright © 2017 Elsevier B.V. All rights reserved.

Scale-up of a physiologically-based pharmacokinetic model to predict the disposition of monoclonal antibodies in monkeys.

PubMed

Glassman, Patrick M; Chen, Yang; Balthasar, Joseph P

2015-10-01

Preclinical assessment of monoclonal antibody (mAb) disposition during drug development often includes investigations in non-human primate models. In many cases, mAb exhibit non-linear disposition that relates to mAb-target binding [i.e., target-mediated disposition (TMD)]. The goal of this work was to develop a physiologically-based pharmacokinetic (PBPK) model to predict non-linear mAb disposition in plasma and in tissues in monkeys. Physiological parameters for monkeys were collected from several sources, and plasma data for several mAbs associated with linear pharmacokinetics were digitized from prior literature reports. The digitized data displayed great variability; therefore, parameters describing inter-antibody variability in the rates of pinocytosis and convection were estimated. For prediction of the disposition of individual antibodies, we incorporated tissue concentrations of target proteins, where concentrations were estimated based on categorical immunohistochemistry scores, and with assumed localization of target within the interstitial space of each organ. Kinetics of target-mAb binding and target turnover, in the presence or absence of mAb, were implemented. The model was then employed to predict concentration versus time data, via Monte Carlo simulation, for two mAb that have been shown to exhibit TMD (2F8 and tocilizumab). Model predictions, performed a priori with no parameter fitting, were found to provide good prediction of dose-dependencies in plasma clearance, the areas under plasma concentration versu time curves, and the time-course of plasma concentration data. This PBPK model may find utility in predicting plasma and tissue concentration versus time data and, potentially, the time-course of receptor occupancy (i.e., mAb-target binding) to support the design and interpretation of preclinical pharmacokinetic-pharmacodynamic investigations in non-human primates.

Structural Analysis of Der p 1–Antibody Complexes and Comparison with Complexes of Proteins or Peptides with Monoclonal Antibodies

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

Osinski, Tomasz; Pomés, Anna; Majorek, Karolina A.

Der p 1 is a major allergen from the house dust mite, Dermatophagoides pteronyssinus, that belongs to the papain-like cysteine protease family. To investigate the antigenic determinants of Der p 1, we determined two crystal structures of Der p 1 in complex with the Fab fragments of mAbs 5H8 or 10B9. Epitopes for these two Der p 1–specific Abs are located in different, nonoverlapping parts of the Der p 1 molecule. Nevertheless, surface area and identity of the amino acid residues involved in hydrogen bonds between allergen and Ab are similar. The epitope for mAb 10B9 only showed a partialmore » overlap with the previously reported epitope for mAb 4C1, a cross-reactive mAb that binds Der p 1 and its homolog Der f 1 from Dermatophagoides farinae. Upon binding to Der p 1, the Fab fragment of mAb 10B9 was found to form a very rare α helix in its third CDR of the H chain. To provide an overview of the surface properties of the interfaces formed by the complexes of Der p 1–10B9 and Der p 1–5H8, along with the complexes of 4C1 with Der p 1 and Der f 1, a broad analysis of the surfaces and hydrogen bonds of all complexes of Fab–protein or Fab–peptide was performed. This work provides detailed insight into the cross-reactive and specific allergen–Ab interactions in group 1 mite allergens. The surface data of Fab–protein and Fab–peptide interfaces can be used in the design of conformational epitopes with reduced Ab binding for immunotherapy.« less

Cross-resistance to purified Bt proteins, Bt corn and Bt cotton in a Cry2Ab2-corn resistant strain of Spodoptera frugiperda.

PubMed

Yang, Fei; Kerns, David L; Head, Graham P; Price, Paula; Huang, Fangneng

2017-12-01

Gene-pyramiding by combining two or more dissimilar Bacillus thuringiensis (Bt) proteins into a crop has been used to delay insect resistance. The durability of gene-pyramiding can be reduced by cross-resistance. Fall armyworm, Spodoptera frugiperda, is a major target pest of the Cry2Ab2 protein used in pyramided Bt corn and cotton. Here, we provide the first experimental evaluation of cross-resistance in S. frugiperda selected with Cry2Ab2 corn to multiple Bt sources including purified Bt proteins, Bt corn and Bt cotton. Concentration - response bioassays showed that resistance ratios for Cry2Ab2-resistant (RR) relative to Cry2Ab2-susceptible (SS) S. frugiperda were -1.4 for Cry1F, 1.2 for Cry1A.105, >26.7 for Cry2Ab2, >10.0 for Cry2Ae and -1.1 for Vip3A. Larvae of Cry2Ab2-heterozygous (RS), SS and RR S. frugiperda were all susceptible to Bt corn and Bt cotton containing Cry1 (Cry1F or Cry1A.105) and/or Vip3A proteins. Pyramided Bt cotton containing Cry1Ac + Cry2Ab2 or Cry1Ab + Cry2Ae were also effective against SS and RS, but not RR. These findings suggest that Cry2Ab2-corn-selected S. frugiperda is not cross-resistant to Cry1F, Cry1A.105 or Vip3A protein, or corn and cotton plants containing these Bt proteins, but it can cause strong cross-resistance to Cry2Ae and Bt crops expressing similar Bt proteins. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Production and characterization of a monoclonal antibody against enrofloxacin.

PubMed

Chusri, Manaspong; Wongphanit, Pitikarn; Palaga, Tanapat; Puthong, Songchan; Sooksai, Sarintip; Komolpis, Kittinan

2013-01-01

Enrofloxacin is a fluoroquinolone antibiotic approved for the treatment of infections in animals. Because of the side effects to consumers of animal products, the maximum residue limits (MRLs) of enrofloxacin in animal tissues for consumption are regulated. In this study, a monoclonal antibody (mAb) against enrofloxacin was prepared and characterized for the development of a direct competitive enzyme-linked immunosorbent assay (ELISA). The obtained mAb, Enro44, was highly specific for enrofloxacin and had a 50% inhibition concentration (IC(50)) of 1.99 ng/ml in a competitive ELISA, and the limit of detection (LOD) was 0.50 ng/ml. The cross-reactivity of the mAb with other quinolones and fluoroquinolones was lower than 0.01%. The subclass of the mAb Enro44 was identified as IgG1. The antigen (Ag)-captured direct competitive ELISA using the mAb Enro44 was tested on different spiked samples, including chicken muscle, cattle milk, and cattle urine, and the assay demonstrated recoveries of 82-112%, 80-125%, and 78-124%, respectively. Furthermore, the quantitation of enrofloxacin obtained from the ELISA and from high-performance liquid chromatography (HPLC) was in good agreement, with the linear regression coefficient between 0.933 and 1.056. The cDNAs encoding a heavy-chain Fd fragment (VH and CH1) and a light chain of the mAb Enro44 were cloned and sequenced. Taken together, the results obtained reveal a potential use of this mAb in an ELISA for the detection of enrofloxacin in food samples. The information of amino acid sequence of this mAb will be useful for further modification and production of the mAb in a bioreactor.

Clinical and biological characteristics of diabetic patients under age 40 in Cameroon: relation to autoantibody status and comparison with Belgian patients

PubMed Central

Asanghanwa, Milca; Gorus, Frans K.; Weets, Ilse; der Auwera, Bart V.; Aminkeng, Folefac; Mbunwe, Eric; Goubert, Patrick; Verhaeghen, Katrijn; Sobngwi, Eugene; Wenzlau, Janet M.; Hutton, John C.; Pipeleers, Daniel G.; Keymeulen, Bart; Mbanya, Jean-Claude N.; Schravendijk, Chris v.

2014-01-01

Aims We investigated the prevalence of diabetes autoantibodies (Abs) in Cameroonian patients and controls, assessed their contribution in disease classification and compared results with data from Belgium. Methods Abs against GAD (GADA), IA-2 (IA-2A) and zinc transporter 8 (ZnT8A) were assessed in 302 recently diagnosed Cameroonian patients with diabetes and 184 control subjects without diabetes aged below 40 years. Results Only 27 (9%) Cameroonian patients were younger than 15 years. Overall, 29% of patients presented at least one diabetes-associated antibody versus 9% in healthy controls (24% versus 7% for GADA (p < 0.001), 10% versus 3% for IA-2A (p < 0.006); 4% versus 2% for ZnT8A). Ab+ patients had lower C-peptide levels (p < 0.001), were more often insulin-treated (p < 0.002) and were as frequently diagnosed with type 1 diabetes as Ab− patients. Only 43% of Ab+ patients aged 15–39 years were clinically classified as having type 1 diabetes in Cameroon vs 96% in Belgium (p < 0.001). Not one Ab+ Cameroonian patient carried HLA-DQ2/DQ8 genotype versus 23% of Belgian Ab+ patients (p < 0.001). Younger age at diagnosis and antibody positivity were independent predictors of insulin therapy. Ab+ Cameroonian patients were older (p < 0.001), had higher BMI (p < 0.001) and lower Ab titers than Belgian Ab+ patients. In ketonuric patients, prevalence of autoantibodies was similar as in non-ketonuric patients. Conclusions In Cameroonian patients with diabetes aged under 40 years, antibody-positivity is not clearly related to disease phenotype, but may help predict the need for insulin treatment. PMID:24332797

Conformational stability and aggregation of therapeutic monoclonal antibodies studied with ANS and Thioflavin T binding.

PubMed

Kayser, Veysel; Chennamsetty, Naresh; Voynov, Vladimir; Helk, Bernhard; Trout, Bernhardt L

2011-01-01

Characterization of aggregation profiles of monoclonal antibodies (mAb) is gaining importance because an increasing number of mAb-based therapeutics are entering clinical studies and gaining marketing approval. To develop a successful formulation, it is imperative to identify the critical biochemical properties of each potential mAb drug candidate. We investigated the conformational change and aggregation of a human IgG1 using external dye-binding experiments with fluorescence spectroscopy and compared the aggregation profiles obtained to the results of size-exclusion chromatography. We show that using an appropriate dye at selected mAb concentration, unfolding or aggregation can be studied. In addition, dye-binding experiments may be used as conventional assays to study therapeutic mAb stability.

Modeling study of the ABS relay valve

NASA Astrophysics Data System (ADS)

Lei, Ming; Lin, Min; Guo, Bin; Luo, Zai; Xu, Weidong

2011-05-01

The ABS (anti-lock braking system) relay valve is the key component of anti-lock braking system in most commercial vehicles such as trucks, tractor-trailers, etc. In this paper, structure of ABS relay valve and its work theory were analyzed. Then a mathematical model of ABS relay valve, which was investigated by dividing into electronic part, magnetic part, pneumatic part and mechanical part, was set up. The displacement of spools and the response of pressure increasing, holding, releasing of ABS relay valve were simulated and analyzed under conditions of control pressure 500 KPa, braking pressure 600 KPa, atmospheric pressure 100 KPa and air temperature 310 K. Thisarticle provides reliable theory for improving the performance and efficiency of anti-lock braking system of vehicles.

Structure determination and total synthesis of a novel antibacterial substance, AB0022A, produced by a cellular slime mold.

PubMed

Sawada, T; Aono, M; Asakawa, S; Ito, A; Awano, K

2000-09-01

A novel antibacterial substance, AB0022A, was isolated from the cellular slime mold Dictyostelium purpureum K1001. It inhibited the growth of Gram-positive bacteria, and its MICs ranged from 0.39 to 50 microg/ml. Because AB0022A was a highly substituted aromatic compound, we could not determine its structure based on only its physico-chemical and spectral data. We therefore used a dehalogenated derivative from AB0022A and deduced that its structure was 1,9-dihydroxy-3,7-dimethoxy-2-hexanoyl-4,6,8-trichlorodibenzofuran . To confirm this structure, we synthesized the compound having the deduced structure. The synthetic compound was identical to naturally occurring AB0022A.

Scattering on two Aharonov-Bohm vortices

NASA Astrophysics Data System (ADS)

Bogomolny, E.

2016-12-01

The problem of two Aharonov-Bohm (AB) vortices for the Helmholtz equation is examined in detail. It is demonstrated that the method proposed by Myers (1963 J. Math. Phys. 6 1839) for slit diffraction can be generalised to obtain an explicit solution for AB vortices. Due to the singular nature of AB interaction the Green function and scattering amplitude for two AB vortices obey a series of partial differential equations. Coefficients entering these equations, fulfil ordinary non-linear differential equations whose solutions can be obtained by solving the Painlevé III equation. The asymptotics of necessary functions for very large and very small vortex separations are calculated explicitly. Taken together, this means that the problem of two AB vortices is exactly solvable.

40 CFR 91.123 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2010 CFR

2010-07-01

....203(f), 91.206(d), 91.208(c) or 91.209(g), the Administrator may void such certificate ab initio. (d) When the Administrator denies, revokes, or voids ab initio a certificate of conformity, the engine... makes the certificate void ab initio. ...

40 CFR 86.1850-01 - Denial, suspension or revocation of certificate of conformity.

Code of Federal Regulations, 2010 CFR

2010-07-01

... conditions specified in § 86.1843-01, the Administrator may deem such certificate void ab initio. (e) When the Administrator denies, suspends, revokes, or voids ab initio a certificate, EPA will provide the... that makes the certification void ab initio. ...

40 CFR 86.1850-01 - Denial, suspension or revocation of certificate of conformity.

Code of Federal Regulations, 2014 CFR

2014-07-01

... conditions specified in § 86.1843-01, the Administrator may deem such certificate void ab initio. (e) When the Administrator denies, suspends, revokes, or voids ab initio a certificate, EPA will provide the... that makes the certification void ab initio. ...

40 CFR 91.123 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2013 CFR

2013-07-01

....203(f), 91.206(d), 91.208(c) or 91.209(g), the Administrator may void such certificate ab initio. (d) When the Administrator denies, revokes, or voids ab initio a certificate of conformity, the engine... makes the certificate void ab initio. ...

40 CFR 91.123 - Denial, revocation of certificate of conformity.

Code of Federal Regulations, 2011 CFR

2011-07-01

....203(f), 91.206(d), 91.208(c) or 91.209(g), the Administrator may void such certificate ab initio. (d) When the Administrator denies, revokes, or voids ab initio a certificate of conformity, the engine... makes the certificate void ab initio. ...