10 % fluorescein sodium vs 1 % isosulfan blue in breast sentinel lymph node biopsy.

PubMed

Ren, Lidong; Liu, Zhao; Liang, Mengdi; Wang, Li; Song, Xingli; Wang, Shui

2016-11-03

Sentinel lymph node biopsy (SLNB) is well accepted to be a standard procedure in breast cancer surgery with clinically negative lymph nodes. Isosulfan blue is the first dye approved by the USA Food and Drug Administration for the localization of the lymphatic system. Few alternative tracers have been investigated. In this study, we aimed to compare the differences between 10 % fluorescein sodium and 1 % isosulfan blue in breast sentinel lymph node biopsy and to investigate the feasibility of using 10 % fluorescein sodium as a new dye for breast sentinel lymph node biopsy. A total of 30 New Zealand rabbits were randomly divided into the fluorescein sodium group and the isosulfan blue group (15 rabbits per group). Fluorescein sodium or isosulfan blue was injected subcutaneously into the second pair of mammary areolas. The average fading time of the second lymph nodes in the isosulfan blue group was significantly shorter than that in the fluorescein sodium group. Moreover, the detection rates of SLNs were higher in the fluorescein sodium group than in the isosulfan blue group. No significant differences between the fluorescein sodium group and isosulfan blue group were observed regarding the distances between the detected sentinel lymph nodes and second pair of mammary areolas, the distances between the second lymph nodes and second pair of mammary areolas, the number of detected sentinel lymph nodes and second lymph nodes, the average dyeing time of the sentinel and the second lymph nodes, and the average fading time of the second lymph nodes. In summary, we first reported that fluorescein sodium is a potential new tracer for breast sentinel lymph node biopsy.

Optimal time following fluorescein instillation to evaluate rigid gas permeable contact lens fit.

PubMed

Wolffsohn, James S; Tharoo, Ali; Lakhlani, Nikita

2015-04-01

To examine the optimum time at which fluorescein patterns of gas permeable lenses (GPs) should be evaluated. Aligned, 0.2mm steep and 0.2mm flat GPs were fitted to 17 patients (aged 20.6 ± 1.1 years, 10 male). Fluorescein was applied to their upper temporal bulbar conjunctiva with a moistened fluorescein strip. Digital slit lamp images (CSO, Italy) at 10× magnification of the fluorescein pattern viewed with blue light through a yellow filter were captured every 15s. Fluorescein intensity in central, mid peripheral and edge regions of the superior, inferior, temporal and nasal quadrants of the lens were graded subjectively using a +2 to -2 scale and using ImageJ software on the simultaneously captured images. Subjectively graded and objectively image analysed fluorescein intensity changed with time (p < 0.001), lens region (centre, mid-periphery and edge: p < 0.05) and there was interaction between lens region with lens fit (p < 0.001). For edge band width, there was a significant effect of time (F = 118.503, p < 0.001) and lens fit (F = 5.1249, p = 0.012). The expected alignment, flat and steep fitting patterns could be seen from approximately after 30 to 180 s subjectively and 15 to 105 s in captured images. Although the stability of fluorescein intensity can start to decline in as little as 45 s post fluorescein instillation, the diagnostic pattern of alignment, steep or flat fit is seen in each meridian by subjective observation from about 30s to 3 min indicating this is the most appropriate time window to evaluate GP lenses in clinical practice. Copyright © 2014 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

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

Colotelo, Alison HA; Smokorowski, Karen; Haxton, Tim

Current methods of fish epithelial injury detection are limited to gross macroscopic examination that has a subjective bias as well as an inability to reliably quantify the degree of injury. Fluorescein, a presumptive test for blood, has been shown to have the capability to detect and quantify fish epithelial injury. However, there are several other presumptive tests for blood (Bluestar*, phenolphthalein, and HemastixH) that may have benefits over the use of fluorescein, particularly for field research on wild fish. This study investigated the capabilities of these four tests to detect and quantify a variety of injuries commonly encountered by fishmore » (abrasion, cuts, fin frays, and punctures) using the freshwater bluegill Lepomis macrochirus as a model. Fluorescein was consistently found to be the most reliable (i.e., detected the highest proportion of true positive results and rarely detected false positive reactions) of the four presumptive tests for blood compared. Further testing was conducted to examine the reliability of fluorescein. By 24 h after an injury was inflicted, the injury was no longer detectable by fluorescein, and when fluorescein was applied to an injured fish, the fluorescein was no longer detectable 3 h after application. In a comparison of two common anaesthetics used in fisheries research, there was no significant difference in the proportion of injury detected when 3- aminobenzoic acid ethyl ester methanesulfate (tricaine) was used compared with a clove oil and ethanol (1:9) solution. In summary, fluorescein was the most reliable presumptive test for blood examined in this study for the detection and quantification of recent (hours) fish epithelial injury.« less

In vivo imaging of human retinal microvasculature using adaptive optics scanning light ophthalmoscope fluorescein angiography

PubMed Central

Pinhas, Alexander; Dubow, Michael; Shah, Nishit; Chui, Toco Y.; Scoles, Drew; Sulai, Yusufu N.; Weitz, Rishard; Walsh, Joseph B.; Carroll, Joseph; Dubra, Alfredo; Rosen, Richard B.

2013-01-01

The adaptive optics scanning light ophthalmoscope (AOSLO) allows visualization of microscopic structures of the human retina in vivo. In this work, we demonstrate its application in combination with oral and intravenous (IV) fluorescein angiography (FA) to the in vivo visualization of the human retinal microvasculature. Ten healthy subjects ages 20 to 38 years were imaged using oral (7 and/or 20 mg/kg) and/or IV (500 mg) fluorescein. In agreement with current literature, there were no adverse effects among the patients receiving oral fluorescein while one patient receiving IV fluorescein experienced some nausea and heaving. We determined that all retinal capillary beds can be imaged using clinically accepted fluorescein dosages and safe light levels according to the ANSI Z136.1-2000 maximum permissible exposure. As expected, the 20 mg/kg oral dose showed higher image intensity for a longer period of time than did the 7 mg/kg oral and the 500 mg IV doses. The increased resolution of AOSLO FA, compared to conventional FA, offers great opportunity for studying physiological and pathological vascular processes. PMID:24009994

Ultra-wide-field angiography improves the detection and classification of diabetic retinopathy.

PubMed

Wessel, Matthew M; Aaker, Grant D; Parlitsis, George; Cho, Minhee; D'Amico, Donald J; Kiss, Szilárd

2012-04-01

To evaluate patients with diabetic retinopathy using ultra-wide-field fluorescein angiography and to compare the visualized retinal pathology with that seen on an overly of conventional 7 standard field (7SF) imaging. Two hundred and eighteen eyes of 118 diabetic patients who underwent diagnostic fluorescein angiography using the Optos Optomap Panoramic 200A imaging system were included. The visualized area of the retina, retinal nonperfusion, retinal neovascularization, and panretinal photocoagulation were quantified by two independent masked graders. The respective areas identified on the ultra-wide-field fluorescein angiography image were compared with an overly of a modified 7SF image as outlined in the Early Treatment Diabetic Retinopathy Study. Ultra-wide-field fluorescein angiograms imaging, on average, demonstrated 3.2 times more total retinal surface area than 7SF. When compared with 7SF, ultra-wide-field fluorescein angiography showed 3.9 times more nonperfusion (P < 0.001), 1.9 times more neovascularization (P = 0.036), and 3.8 times more panretinal photocoagulation (P < 0.001). In 22 eyes (10%), ultra-wide-field fluorescein angiography demonstrated retinal pathology (including nonperfusion and neovascularization) not evident in an 7SF overly. Compared with conventional 7SF imaging, ultra-wide-field fluorescein angiography reveals significantly more retinal vascular pathology in patients with diabetic retinopathy. Improved retinal visualization may alter the classification of diabetic retinopathy and may therefore influence follow-up and treatment of these patients.

Fluorescein as a Visible-Light-Induced Oxidase Mimic for Signal-Amplified Colorimetric Assay of Carboxylesterase by an Enzymatic Cascade Reaction.

PubMed

Liu, Li; Sun, Chaoqun; Yang, Juan; Shi, Ying; Long, Yijuan; Zheng, Huzhi

2018-04-20

We have found that fluorescein possesses high visible-light-induced oxidase mimetic activity and could transform colorless 3,3',5,5'-tetramethylbenzidine (TMB) into blue oxidized TMB (oxTMB) without unstable and destructive H 2 O 2 under visible-light illumination. Instead, fluorescein uses oxygen as a mild and green electron acceptor, and its activity can be easily controlled by the switching "on/off" of visible light. In addition, the visible-light-induced catalytic mechanism was elucidated in detail and, as the main reactive species h + and O 2 .- accounted for TMB oxidation. Based on the fact that fluorescein diacetate (FDA) possessed no activity and generated active fluorescein in situ in the presence of carboxylesterase (CaE), a signal-amplified sensing platform through a cascade reaction for CaE detection was constructed. Our proposed sensing system displayed excellent analytical performance for the detection of CaE in a wide linear range from 0.040 to 20 U L -1 with a low detection limit of 0.013 U L -1 . This work not only changes the conventional concept that fluorescein is generally considered to be photocatalytically inert, but also provides a novel sensing strategy by tailoring the enzyme mimetic activity of fluorescein derivatives with analyte. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

New stroboscopic light source and technique for intraoperative retinal fluorescein angiography during penetrating keratoplasty

NASA Astrophysics Data System (ADS)

Krueger, Ronald R.; Morales, Ronald B.; Chong, Lawrence P.; Smith, Ronald E.

1994-06-01

We report the development of a new stroboscopic light source system and technique for performing intraoperative fluorescein angiography during penetrating keratoplasty for aphakic or pseudophakic bullous keratopathy. A controllable pulse xenon light source system with a fiber optic endoilluminator probe is used to perform high-quality intraoperative fluorescein angiography during penetrating keratoplasty in pigmented rabbits and human subjects. Following corneal trephination and extraction of the intraocular lens, a temporary Cobo keratoprosthesis is secured while a 20-gauge endoilluminator is inserted into the vitreous cavity through a limbal incision. The endoilluminator is advanced to a retinal illumination area of approximately 3 DD and 10% fluorescein is injected intravenously. A microscope camera coupled to a 50:50 beamsplitter photographs the passage of fluorescein dye while the surgeon maintains an unaltered view through the operating microscope. Angiograms through a keratoprosthesis show excellent contrast and resolution, comparable to standard fluorescein angiography. Fine peripapillary vessels are seen reproducibly and with great detail in the rabbits. All the phases of retinal angiography can be seen, including arteriolar constriction and capillary nonperfusion in one of four human subjects examined. High quality intraoperative fluorescein angiography can be performed in patients undergoing penetrating keratoplasty for aphakic/ pseudophakic bullous keratopathy. With this technology, preexisting retinal disorders such as cystoid macular edema might be identified in the perioperative setting allowing for important management decisions to be made intraoperatively.

Clinical applications of in vivo fluorescence confocal laser scanning microscopy

NASA Astrophysics Data System (ADS)

Oh, Chilhwan; Park, Sangyong; Kim, Junhyung; Ha, Seunghan; Park, Gyuman; Lee, Gunwoo; Lee, Onseok; Chun, Byungseon; Gweon, Daegab

2008-02-01

Living skin for basic and clinical research can be evaluated by Confocal Laser Scanning Microscope (CLSM) non-invasively. CLSM imaging system can achieve skin image its native state either "in vivo" or "fresh biopsy (ex vivo)" without fixation, sectioning and staining that is necessary for routine histology. This study examines the potential fluorescent CLSM with a various exogenous fluorescent contrast agent, to provide with more resolution images in skin. In addition, in vivo fluorescent CLSM researchers will be extended a range of potential clinical application. The prototype of our CLSM system has been developed by Prof. Gweon's group. The operating parameters are composed of some units, such as illuminated wavelength 488 nm, argon illumination power up to 20mW on the skin, objective lens, 0.9NA oil immersion, axial resolution 1.0μm, field of view 200μm x 100μm (lateral resolution , 0.3μm). In human volunteer, fluorescein sodium was administrated topically and intradermally. Animal studies were done in GFP transgenic mouse, IRC mouse and pig skin. For imaging of animal skin, fluorescein sodium, acridine orange, and curcumine were used for fluorescein contrast agent. We also used the GFP transgenic mouse for fluorescein CLSM imaging. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. Acridin Orange can be highlight nuclei in viable keratinocyte. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, hair and eccrine gland. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. In papillary dermis, fluorescein distribution is more homogeneous. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, skin appendage and blood vessels. In conclusion, this study demonstrates the usefulness of CLSM as technique for imaging skin in vivo. In addition, CLSM is non-invasive, the same tissue site may be imaged over a period of time to monitor the various change such as wound healing, severity of skin diseases and effect of therapeutic management.

NI-16INTRA-OPERATIVE USE OF FLUORESCEIN FOR MALIGNANT GLIOMA RESECTION DIFFERENTIATES TUMOR FROM NORMAL BRAIN TISSUE BASED ON HISTOPATHOLOGIC ANALYSIS

PubMed Central

Decker, Matthew; Kresak, Jesse; Yachnis, Anthony; Bova, Frank; Rahman, Maryam

2014-01-01

OBJECTIVES: To determine whether the use of IV fluorescein during surgery for malignant glioma can reliably be used to differentiate between infiltrative tumor and normal brain tissue. BACKGROUND: Fluorescein sodium is a molecular compound with fluorescent capabilities between light wavelengths of 520-530nm, appearing yellow-green (1). Neurosurgical application of fluorescein has been studied primarily for increasing intra-operative visibility of malignant gliomas (1). The mechanism of action has been hypothesized to involve disruption of the blood brain barrier (BBB) (2). Cells in areas with disrupted BBB take up fluorescein with a sensitivity of 94% and specificity of 89% for high-grade gliomas (2). We performed histopathologic analysis on tissue obtained during fluorescein-guided tumor resections to evaluate the differences between fluorescent and non-fluorescent tissue. METHODS: Two adult patients with suspected high-grade gliomas underwent surgical resection. Prior to opening of the dura 3mg/kg of IV fluorescein was given. A Zeiss OPMI Pentero microscope (Carl Zeiss Meditech Inc.) with a yellow 560nm filter was used to visualize the tumor. At the tumor margins, tissue was identified as "bright" and "dark" and sent as separate specimens for histopathological analysis. RESULTS: Histological sections of specimens labeled "bright" contained infiltrating glioma with focal microvascular proliferation. Histological sections of specimens labeled "dark" contained gray matter and focal subcortical white matter with no high-grade glioma identified. Final grading for both patients was WHO Grade IV, glioblastoma. CONCLUSION: Intra-operative use of fluorescein in surgical resection of malignant gliomas can help to distinguish between infiltrating tumor and normal brain tissue based on histopathological analysis. Further evaluation of the utility of flurorescein during high and low-grade glioma surgery is necessary.

Acute Toxicity of Sodium Fluorescein to Ashy Pebblesnails Fluminicola fuscus

USGS Publications Warehouse

Stockton, Kelly A.; Moffitt, Christine M.; Blew, David L.; Farmer, C. Neil

2011-01-01

Water resource agencies and groundwater scientists use fluorescein dyes to trace ground water flows that supply surface waters that may contain threatened or endangered mollusk species. Since little is known of the toxicity of sodium fluorescein to mollusks, we tested the toxicity of sodium fluorescein to the ashy pebblesnail Fluminicola fuscus. The pebblesnail was selected as a surrogate test species for the threatened Bliss Rapid snail Taylorcocha serpenticola that is endemic to the Snake River and its tributaries in the Hagerman Valley, Idaho. In laboratory tests, we expose replicated groups of snails to a series of concentrations of fluorescein in a static 24 h exposure at 15 degrees C. Following the exposure, we removed snails, rinsed them, and allowed a 48 h recovery in clean water before recording mortality. We estimated 377 mg/L as the median lethal dose. Mortality to snails occurred at concentrations well above those expected in test wells during the monitoring efforts.

[Quenched fluorescein: a reference dye for instrument response function of TCSPC].

PubMed

Pan, Hai-feng; Ding, Jing-xin; Liang, Rong-rong; Tao, Zhan-dong; Liu, Meng-wei; Zhang, San-jun; Xu, Jian-hua

2014-08-01

Measuring the instrument response function (IRF) and fitting by reconvolution algorithms are routines to improve time resolution in fluorescence lifetime measurements. Iodide ions were successfully used to quench the fluorescence of fluorescein in this study. By systematically adding saturated NaI water solution in basic fluorescein solution, the lifetimes of fluorescein were reduced from 4 ns to 24 ps. The quenched lifetime of fluorescein obtained from the analysis of Time-Correlated Single Photon Counting (TCSPC) measurement agrees well with that from femtosecond frequency up-conversion measurement. In time resolved excitation spectra measurements, the IRF should be measured at various detection wavelengths providing scattring materials are used. This study could not only reduce the complexity of IRF measurement, but also avoid the existing color effect in system. This study should have wide applications in time resolved fluorescence spectroscopy and fluorescence lifetime imaging.

Use of vital dyes to assess embryonic viability in the hamster, Mesocricetus auratus

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

Hutz, R.J.; DeMayo, F.J.; Dukelow, W.R.

1985-05-01

Experiments were designed to assess the use of the vital dyes trypan blue and fluorescein diacetate as indicators of the viability of hamster ova and embryos. Exclusion of trypan blue and fluorescence with fluorescein diacetate showed high correlations with uptake of (/sup 3/H)uridine by ova and further development of embryos in vitro. Ova killed by freezing and thawing incorporated (/sup 3/H)uridine at background levels. Trypan blue exclusion and fluorescein diacetate uptake were highly correlated with each other (r = 0.99). Trypan blue and fluorescein diacetate serve as excellent indices of viability in ova and early embryos of hamsters.

Quantification of intracellular payload release from polymersome nanoparticles

NASA Astrophysics Data System (ADS)

Scarpa, Edoardo; Bailey, Joanne L.; Janeczek, Agnieszka A.; Stumpf, Patrick S.; Johnston, Alexander H.; Oreffo, Richard O. C.; Woo, Yin L.; Cheong, Ying C.; Evans, Nicholas D.; Newman, Tracey A.

2016-07-01

Polymersome nanoparticles (PMs) are attractive candidates for spatio-temporal controlled delivery of therapeutic agents. Although many studies have addressed cellular uptake of solid nanoparticles, there is very little data available on intracellular release of molecules encapsulated in membranous carriers, such as polymersomes. Here, we addressed this by developing a quantitative assay based on the hydrophilic dye, fluorescein. Fluorescein was encapsulated stably in PMs of mean diameter 85 nm, with minimal leakage after sustained dialysis. No fluorescence was detectable from fluorescein PMs, indicating quenching. Following incubation of L929 cells with fluorescein PMs, there was a gradual increase in intracellular fluorescence, indicating PM disruption and cytosolic release of fluorescein. By combining absorbance measurements with flow cytometry, we quantified the real-time intracellular release of a fluorescein at a single-cell resolution. We found that 173 ± 38 polymersomes released their payload per cell, with significant heterogeneity in uptake, despite controlled synchronisation of cell cycle. This novel method for quantification of the release of compounds from nanoparticles provides fundamental information on cellular uptake of nanoparticle-encapsulated compounds. It also illustrates the stochastic nature of population distribution in homogeneous cell populations, a factor that must be taken into account in clinical use of this technology.

Effects of topical anaesthetics and fluorescein on the real‐time PCR used for the diagnosis of Herpesviruses and Acanthamoeba keratitis

PubMed Central

Goldschmidt, P; Rostane, H; Saint‐Jean, C; Batellier, L; Alouch, C; Zito, E; Bourcier, T; Laroche, L; Chaumeil, C

2006-01-01

Background The early microbiological diagnosis of corneal infections may prevent the condition from worsening. Aim To study the potential interferences of oxybuprocain and fluorescein solutions used by ophthalmologists on the performances of the real‐time polymerase chain reaction (PCR) carried out as routine test for diagnosis of keratitis. Methods Quantified suspensions of Herpes simplex virus (HSV1), Varicella zoster virus (VZV), Cytomegalovirus (CMV) and Acanthamoeba with and without oxybuprocain or fluorescein added before DNA extraction were tested by real‐time PCR. Results The capacities of the real‐time PCR to detect HSV, VZV, CMV and Acanthamoeba were reduced by oxybuprocain and fluorescein. Both products diluted to 1/16 reduced the PCR detection capacities for more than 2 logs (DNA copies/sample). Conclusions The simultaneous introduction of fluorescein or topical anaesthetics into the tubes containing the specimens to be tested by PCR may lead to false negative results. Because corneal specimens for microbiological diagnosis of keratitis are obtained after topical administration of anaesthetics and corneal staining with fluorescein, ophthalmologists should be aware to rinse the eye surface intensively with appropriate eye solutions to minimise the risks of misdiagnosis. PMID:16899529

Effects of topical anaesthetics and fluorescein on the real-time PCR used for the diagnosis of Herpesviruses and Acanthamoeba keratitis.

PubMed

Goldschmidt, P; Rostane, H; Saint-Jean, C; Batellier, L; Alouch, C; Zito, E; Bourcier, T; Laroche, L; Chaumeil, C

2006-11-01

The early microbiological diagnosis of corneal infections may prevent the condition from worsening. To study the potential interferences of oxybuprocain and fluorescein solutions used by ophthalmologists on the performances of the real-time polymerase chain reaction (PCR) carried out as routine test for diagnosis of keratitis. Quantified suspensions of Herpes simplex virus (HSV1), Varicella zoster virus (VZV), Cytomegalovirus (CMV) and Acanthamoeba with and without oxybuprocain or fluorescein added before DNA extraction were tested by real-time PCR. The capacities of the real-time PCR to detect HSV, VZV, CMV and Acanthamoeba were reduced by oxybuprocain and fluorescein. Both products diluted to 1/16 reduced the PCR detection capacities for more than 2 logs (DNA copies/sample). The simultaneous introduction of fluorescein or topical anaesthetics into the tubes containing the specimens to be tested by PCR may lead to false negative results. Because corneal specimens for microbiological diagnosis of keratitis are obtained after topical administration of anaesthetics and corneal staining with fluorescein, ophthalmologists should be aware to rinse the eye surface intensively with appropriate eye solutions to minimise the risks of misdiagnosis.

Functional expression of the 11 human Organic Anion Transporting Polypeptides in insect cells reveals that sodium fluorescein is a general OATP substrate.

PubMed

Patik, Izabel; Kovacsics, Daniella; Német, Orsolya; Gera, Melinda; Várady, György; Stieger, Bruno; Hagenbuch, Bruno; Szakács, Gergely; Özvegy-Laczka, Csilla

2015-12-15

Organic Anion Transporting Polypeptides (OATPs), encoded by genes of the Solute Carrier Organic Anion (SLCO) family, are transmembrane proteins involved in the uptake of various compounds of endogenous or exogenous origin. In addition to their physiological roles, OATPs influence the pharmacokinetics and drug-drug interactions of several clinically relevant compounds. To examine the function and molecular interactions of human OATPs, including several poorly characterized family members, we expressed all 11 human OATPs at high levels in the baculovirus-Sf9 cell system. We measured the temperature- and inhibitor-sensitive cellular accumulation of sodium fluorescein and fluorescein-methotrexate, two fluorescent substrates of the OATPs, OATP1B1 and 1B3. OATP1B1 and 1B3 were functional in Sf9 cells, showing rapid uptake (t1/2(fluorescein-methotrexate) 2.64 and 4.16 min, and t1/2(fluorescein) 6.71 and 5.58 min for OATP1B1 and 1B3, respectively) and high-affinity transport (Km(fluorescein-methotrexate) 0.23 and 0.53 μM, and Km(fluorescein) 25.73 and 38.55 μM for OATP1B1 and 1B3, respectively) of both substrates. We found that sodium fluorescein is a general substrate of all human OATPs: 1A2, 1B1, 1B3, 1C1, 2A1, 2B1, 3A1, 4A1, 4C1, 5A1 and 6A1, while fluorescein-methotrexate is only transported by 1B1, 1B3, 1A2 and 2B1. Acidic extracellular pH greatly facilitated fluorescein uptake by all OATPs, and new molecular interactions were detected (between OATP2B1 and Imatinib, OATP3A1, 5A1 and 6A1 and estradiol 17-β-d-glucuronide, and OATP1C1 and 4C1 and prostaglandin E2). These studies demonstrate, for the first time, that the insect cell system is suitable for the functional analysis of the entire human OATP family, and for drug-OATP interaction screening. Copyright © 2015 Elsevier Inc. All rights reserved.

Environmental and body contamination from cleaning vomitus in a health care setting: A simulation study.

PubMed

Phan, Linh; Su, Yu-Min; Weber, Rachel; Fritzen-Pedicini, Charissa; Edomwande, Osayuwamen; Jones, Rachael M

2018-04-01

Environmental service workers may be exposed to pathogens during the cleaning of pathogen-containing bodily fluids. Participants with experience cleaning hospital environments were asked to clean simulated, fluorescein-containing vomitus using normal practices in a simulated patient room. Fluorescein was visualized in the environment and on participants under black lights. Fluorescein was quantitatively measured on the floor, in the air, and on gloves and shoe covers. In all 21 trials involving 7 participants, fluorescein was found on the floor after cleaning and on participants' gloves. Lower levels of floor contamination were associated with the use of towels to remove bulk fluid (ρ = -0.56, P = .01). Glove contamination was not associated with the number or frequency of contacts with environmental surfaces, suggesting contamination occurs with specific events, such as picking up contaminated towels. Fluorescein contamination on shoe covers was measured in 19 trials. Fluorescein was not observed on participants' facial personal protective equipment, if worn, or faces. Contamination on other body parts, primarily the legs, was observed in 8 trials. Fluorescein was infrequently quantified in the air. Using towels to remove bulk fluid prior to mopping is part of the recommended cleaning protocol and should be used to minimize residual contamination. Contamination on shoes and the floor may serve as reservoirs for pathogens. Copyright © 2018 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.

Fluorescein angiography of the bladder: technique and relevance to bladder cancer and interstitial cystitis patients.

PubMed

Zimmern, P E; Laub, D; Leach, G E

1995-07-01

Fluorescein angiography has been used in the study of bleeding vessels, neovascularity, tumors and ischemic tissues in a variety of disorders. This pilot study was designed to evaluate the feasibility, safety and relevance of this interesting technology for the evaluation of bladder wall vessels in patients with interstitial cystitis and bladder cancer. Five patients with National Institutes of Health defined interstitial cystitis symptoms and 10 with bladder cancer were studied during cytoscopy while they were under general anesthesia. A yellow-green barrier filter (520 nm.) was placed over the cystoscope eyepiece and a blue exciter filter (465 nm.) was attached to the light source. Patients received a 5 ml. bolus of 10% fluorescein intravenously. After hydrodistension, glomerulations in interstitial cystitis patients were more prominent with fluorescein angiography and occurred in the venule phase. Areas of papillary transitional cell tumor and carcinoma in situ developed a brilliant yellow-green fluorescence. Adjacent normal urothelium was nonfluorescent and provided a contrasting dark background facilitating the detection of all lesions. No allergic reaction or other adverse effect related to the fluorescein injection was observed. These unique observations in a limited number of patients suggest that fluorescein angiography of the bladder is a safe and simple procedure. This preliminary report underscores the relevance of fluorescein angiography in the detection of bladder tumor and offers a new approach to the evaluation of bladder wall vessels in interstitial cystitis patients.

Fluress, fluorescein and benoxinate: recovery from bacterial contamination.

PubMed

Yolton, D P; German, C J

1980-05-01

The ability to recover from bacterial contaminations with Staphylococcus auresus and Pseudomonas aeruginosa was determined for three optometric DPAs: Fluress, fluorescein and benoxinate. Results show that Fluress recovers from contamination more rapidly than benoxinate or fluorescein. This ability to recover from contamination and the relative ease of use of Fluress may make it the DPA choice for a number of optometric procedures including applanation tonometry.

Sibling rivalry: intrinsic luminescence from two xanthene dye monoanions, resorufin and fluorescein, provides evidence for excited-state proton transfer in the latter.

PubMed

Kjær, Christina; Brøndsted Nielsen, Steen; Stockett, Mark H

2017-09-20

While the emission spectrum of fluorescein monoanions isolated in vacuo displays a broad and featureless band, that of resorufin, also belonging to the xanthene family, has a sharp band maximum, clear vibronic structure, and experiences a small Stokes shift. Excited-state proton transfer in fluorescein can account for the differences.

In Vitro Investigation of Influences of Chitosan Nanoparticles on Fluorescein Permeation into Alveolar Macrophages.

PubMed

Chachuli, Siti Haziyah Mohd; Nawaz, Asif; Shah, Kifayatullah; Naharudin, Idanawati; Wong, Tin Wui

2016-06-01

Pulmonary infection namely tuberculosis is characterized by alveolar macrophages harboring a large microbe population. The chitosan nanoparticles exhibit fast extracellular drug release in aqueous biological milieu. This study investigated the matrix effects of chitosan nanoparticles on extracellular drug diffusion into macrophages. Oligo, low, medium and high molecular weight chitosan nanoparticles were prepared by nanospray drying technique. These nanoparticles were incubated with alveolar macrophages in vitro and had model drug sodium fluorescein added into the same cell culture. The diffusion characteristics of sodium fluorescein and nanoparticle behavior were investigated using fluorescence microscopy, scanning electron microscopy, differential scanning calorimetry and Fourier transform infrared spectroscopy techniques. The oligochitosan nanoparticles enabled macrophage membrane fluidization with the extent of sodium fluorescein entry into macrophages being directly governed by the nanoparticle loading. Using nanoparticles made of higher molecular weight chitosan, sodium fluorescein permeation into macrophages was delayed due to viscous chitosan diffusion barrier at membrane boundary. Macrophage-chitosan nanoparticle interaction at membrane interface dictates drug migration into cellular domains.

Graphene-based biosensors

NASA Astrophysics Data System (ADS)

Lebedev, A. A.; Davydov, V. Yu.; Novikov, S. N.; Litvin, D. P.; Makarov, Yu. N.; Klimovich, V. B.; Samoilovich, M. P.

2016-07-01

Results of developing and testing graphene-based sensors capable of detecting protein molecules are presented. The biosensor operation was checked using an immunochemical system comprising fluorescein dye and monoclonal antifluorescein antibodies. The sensor detects fluorescein concentration on a level of 1-10 ng/mL and bovine serum albumin-fluorescein conjugate on a level of 1-5 ng/mL. The proposed device has good prospects for use for early diagnostics of various diseases.

Rasch analysis of three dry eye questionnaires and correlates with objective clinical tests.

PubMed

McAlinden, Colm; Gao, Rongrong; Wang, Qinmei; Zhu, Senmiao; Yang, Jing; Yu, Ayong; Bron, Anthony J; Huang, Jinhai

2017-04-01

To assess the psychometric properties of Chinese versions of the Ocular Comfort Index (OCI), Ocular Surface Disease Index (OSDI) and McMonnies questionnaires. Further, to assess the correlation between questionnaire scores and objective dry eye disease (DED) clinical tests. Translated versions of the OCI, OSDI and McMonnies questionnaires were completed in a random order by 238 participants with DED. Objective clinical tests included visual acuity (VA), fluorescein tear film break-up time (TBUT), corneal fluorescein staining, Schirmer I testing and meibomian gland grading. Rasch analysis was used to assess questionnaire psychometrics and spearman rank for correlations. For the OCI, the person separation was 2.31, item infit and outfit statistics ranged from 0.74-1.14 and 0.75-1.32, respectively, and targeting 1.54 logits. For the OSDI, person separation was 0.94. None of the three subscales provided valid measurements based on Rasch analysis. For the McMonnies questionnaire, person separation was 1.17, item infit and outfit statistics ranged from 0.7 to 1.21 and 0.51-3.49, respectively. There were weak correlations between questionnaire scores and clinical tests. There were weak correlations between OSDI scores and VA, fluorescein TBUT, Schirmer I testing and corneal fluorescein staining. There were weak correlations between McMonnies scores and VA, fluorescein TBUT, Schirmer I testing, and corneal fluorescein staining and meibomian gland grading. The OCI questionnaire was the only questionnaire that provided valid measurement on the basis of Rasch analysis, although slight multidimensionality was found. There were weak correlations between OCI scores and fluorescein TBUT, Schirmer I testing, and corneal fluorescein staining. Due to this paradoxical disconnect between symptoms and signs and the repeatability of tests, the use of both subjective and objective markers in the clinical management of patients or as endpoints in clinical trials would appear prudent. Copyright © 2017 Elsevier Inc. All rights reserved.

Method and compositions for detecting of bloodstains using fluorescin-fluorescein reaction

DOEpatents

Di Benedetto, John; Kyle, Kevin; Boan, Terry; Marie, Charlene

2004-02-17

A method, compositions and kit are set forth for detecting blood stains. A reactant solution includes fluorescin solubilized (reduced) in acetic acid in ethanol. The solution may be buffered to a pH of approximately 9. After spraying the reactant solution on the suspected area an oxidizer is applied to promote the fluorescin to fluorescein reaction with the blood. The reacted fluorescein is then detected through luminescence for capture by photography.

Precise correlation of histopathological and fluorescein angiographic morphology using retinal vascular casting.

PubMed

Bek, T; Prause, J U

1996-12-01

The histopathology of three eyes obtained post mortem from 2 patients with age-related macular degeneration was correlated with the pre mortem fluorescein angiographic morphology. A precise point-by-point correlation between histopathology and the corresponding angiographic appearance was ensured by using the cast retinal vascular system as a pattern of reference. The study showed that both the photoreceptors, the pigment epithelium, and substances accumulated between the retinal and the choroidal vascular systems, may have a blocking effect on choroidal background fluorescence as seen on fluorescein angiograms. Furthermore, it is confirmed that fluorescein angiographic hyperfluorescence may be due to a lack of blocking of the choroidal fluorescence because of a window defect in the retinal photoreceptor layer and/or the pigment epithelium.

Fluorescein angiography basic science and engineering.

PubMed

Wolfe, D R

1986-12-01

Fluorescein angiography is an application of the physical phenomenon of fluorescence, which is phosphorescence in which the quantum mechanical decay curve is so rapid that it appears instantaneous, and it consequently has no afterglow. Sodium fluorescein is excited by light energy between 465 and 490 nm, and it decays into a lower state emitting light energy between 520 and 530 nm as fluorescent radiation. The free electrons available for excitation are reduced by chemical bonding between the fluorescein dye and plasma proteins to which up to 80% of the dye is bound in the bloodstream, thus reducing overall fluorescence. Optimalization of the observed and recorded fluorescence is afforded by providing exciter and barrier filters with as little overlap as possible to reduce or eliminate contrast reducing pseudofluorescence.

Dual-labeling with 5-aminolevulinic acid and fluorescein for fluorescence-guided resection of high-grade gliomas: technical note.

PubMed

Suero Molina, Eric; Wölfer, Johannes; Ewelt, Christian; Ehrhardt, André; Brokinkel, Benjamin; Stummer, Walter

2018-02-01

OBJECTIVE Fluorescence guidance with 5-aminolevulinic acid (5-ALA) helps improve resections of malignant gliomas. However, one limitation is the low intensity of blue light for background illumination. Fluorescein has recently been reintroduced into neurosurgery, and novel microscope systems are available for visualizing this fluorochrome, which highlights all perfused tissues but has limited selectivity for tumor detection. Here, the authors investigate a combination of both fluorochromes: 5-ALA for distinguishing tumor and fluorescein for providing tissue fluorescence of adjacent brain tissue. METHODS The authors evaluated 6 patients who harbored cerebral lesions suggestive of high-grade glioma. Patients received 5-ALA (20 mg/kg) orally 4 hours before induction of anesthesia. Low-dose fluorescein (3 mg/kg intravenous) was injected immediately after anesthesia induction. Pentero microscopes (equipped either with Yellow 560 or Blue 400 filters) were used to visualize fluorescence. To simultaneously visualize both fluorochromes, the Yellow 560 module was combined with external blue light illumination (D-light C System). RESULTS Fluorescein-induced fluorescence created a useful background for protoporphyrin IX (PPIX) fluorescence, which appeared orange to red, surrounded by greenly fluorescent normal brain and edematous tissue. Green brain-tissue fluorescence was helpful in augmenting background. Levels of blue illumination that were too strong obscured PPIX fluorescence. Unspecific extravasation of fluorescein was noted at resection margins, which did not interfere with PPIX fluorescence detection. CONCLUSIONS Dual labeling with both PPIX and fluorescein fluorescence is feasible and gives superior background information during fluorescence-guided resections. The authors believe that this technique carries potential as a next step in fluorescence-guided resections if it is completely integrated into the surgical microscope.

Fluorescein-guided surgery for grade IV gliomas with a dedicated filter on the surgical microscope: preliminary results in 12 cases.

PubMed

Acerbi, Francesco; Broggi, Morgan; Eoli, Marica; Anghileri, Elena; Cuppini, Lucia; Pollo, Bianca; Schiariti, Marco; Visintini, Sergio; Orsi, Chiara; Franzini, Angelo; Broggi, Giovanni; Ferroli, Paolo

2013-07-01

Fluorescein is widely used as a fluorescent tracer for many applications. Its capability to accumulate in cerebral areas with blood-brain barrier damage makes it an ideal dye for intraoperative visualization of malignant gliomas (MG). We report our preliminary experience in fluorescein-guided removal of grade IV gliomas using a dedicated filter on the surgical microscope. In September 2011 we started a prospective phase II trial (FLUOGLIO) to evaluate the safety and obtain initial indications about the efficacy of fluorescein-guided surgery for MG. Patients with suspected MG amenable to complete resection of contrast-enhancing areas were eligible to participate in this study. This report is based on a preliminary analysis of the results of 12 patients with grade IV gliomas out of 15 consecutive cases (age range 48-72 years) enrolled since September 2011. Fluorescein was injected intravenously (i.v.) after intubation (5-10 mg/kg). The tumor was removed using a microsurgical technique and fluorescence visualization by BLU 400 or YELLOW 560 filters on a Pentero microscope (Carl Zeiss, Germany). The study was approved by our ethics committee and registered on the European Regulatory Authorities website (EudraCT no. 2011-002527-18). Histological analysis confirmed grade IV gliomas in 12/15 cases. Median preoperative tumor volume was 33.15 cm(3) (9.6-87.8 cm(3)). No adverse reaction related to the administration of fluorescein was registered. Contrast-enhanced tumor was completely removed in 75 % of the patients. This preliminary analysis suggested that the use of intravenous fluorescein during surgery on grade IV gliomas is safe and allows a high rate of complete resection of contrast-enhanced tumor at the early postoperative MRI.

Case Report of Bullous Pemphigoid following Fundus Fluorescein Angiography

PubMed Central

Demirci, Goktug; Demirci, Gulsen Tukenmez; Gulkilik, Gokhan

2010-01-01

Purpose To report a first case of bullous pemphigoid (BP) following intravenous fluorescein for fundus angiography. Clinical Features: A 70-year-old male patient was admitted to the intensive care unit with BP and sepsis. He reported a history of fundus fluorescein angiography with a pre-diagnosis of senile macular degeneration 2 months prior to presentation. At that time, fluorescein extravasated at the antecubital region. Following the procedure, pruritus and erythema began at the wrists bilaterally, and quickly spread to the entire body. The patient also reported a history of allergy to human albumin solution (Plamasteril®; Abbott) 15 years before, during bypass surgery. On dermatologic examination, erythematous patches were present on the scalp, chest and anogenital region. Vesicles and bullous lesions were present on upper and lower extremities. On day 2 of hospitalization, tense bullae appeared on the upper and lower extremities. The patient was treated with oral methylprednisolone 48 mg (Prednol®; Mustafa Nevzat), topical clobetasol dipropionate 0.05% cream (Dermovate®; Glaxo SmithKline), and topical 4% urea lotion (Excipial Lipo®; Orva) for presumptive bullous pemphigoid. Skin punch biopsy provided tissue for histopathology, direct immunofluorescence examination, and salt extraction, which were all consistent with BP. After 1 month, the patient was transferred to the intensive care unit with sepsis secondary to urinary tract infection; he died 2 weeks later from sepsis and cardiac failure. Conclusions To our knowledge, this is the first reported case of BP following fundus fluorescein angiography in a patient with known human albumin solution allergy. Consideration should be made to avoid fluorescein angiography, change administration route, or premedicate with antihistamines in patients with known human albumin solution allergy. The association between fundus fluorescein angiography and BP should be further investigated. PMID:20737052

Rapid, High Affinity Binding by a Fluorescein Templated Copolymer Combining Covalent, Hydrophobic, and Acid–Base Noncovalent Crosslinks

PubMed Central

Timberman, Anthony; Yang, Rongfang; Papantones, Alex; Seitz, W. Rudolf

2018-01-01

A new type of biomimetic templated copolymer has been prepared by reverse addition fragmentation chain transfer polymerization (RAFT) in dioxane. The initial formulation includes the template fluorescein, N-isopropylacrylamide (NIPAM, 84 mol %), methacrylic acid (MAA, 5-mol %), 4-vinylpyridine (4-VP, 9 mmol %), and N,N′-methylenebis(acrylamide) (MBA, 2 mol %). PolyNIPAM is a thermosensitive polymer that comes out of aqueous solution above its lower critical solution temperature forming hydrophobic ‘crosslinks’. MAA and 4-VP interact in dioxane forming acid–base crosslinks. The excess 4-VP serves as a recognition monomer organizing around the template fluorescein to form a binding site that is held in place by the noncovalent and covalent crosslinks. The MBA is a covalent crosslinker. The RAFT agent in the resulting copolylmer was reduced to a thiol and attached to gold nanoparticles. The gold nanoparticle bound copolymer binds fluorescein completely in less than two seconds with an affinity constant greater than 108 M−1. A reference copolymer prepared with the same monomers by the same procedure binds fluorescein much more weakly. PMID:29693601

Transfer of bacteriophage MS2 and fluorescein from N95 filtering facepiece respirators to hands: Measuring fomite potential

PubMed Central

Brady, Tyler M.; Strauch, Amanda L.; Almaguer, Claudia M.; Niezgoda, George; Shafer, Ronald E.; Yorio, Patrick L.; Fisher, Edward M.

2017-01-01

Contact transmission of pathogens from personal protective equipment is a concern within the healthcare industry. During public health emergency outbreaks, resources become constrained and the reuse of personal protective equipment, such as N95 filtering facepiece respirators, may be needed. This study was designed to characterize the transfer of bacteriophage MS2 and fluorescein between filtering facepiece respirators and the wearer’s hands during three simulated use scenarios. Filtering facepiece respirators were contaminated with MS2 and fluorescein in droplets or droplet nuclei. Thirteen test subjects performed filtering facepiece respirator use scenarios including improper doffing, proper doffing and reuse, and improper doffing and reuse. Fluorescein and MS2 contamination transfer were quantified. The average MS2 transfer from filtering facepiece respirators to the subjects’ hands ranged from 7.6–15.4% and 2.2–2.7% for droplet and droplet nuclei derived contamination, respectively. Handling filtering facepiece respirators contaminated with droplets resulted in higher levels of MS2 transfer compared to droplet nuclei for all use scenarios (p = 0.007). MS2 transfer from droplet contaminated filtering facepiece respirators during improper doffing and reuse was greater than transfer during improper doffing (p = 0.008) and proper doffing and reuse (p = 0.042). Droplet contamination resulted in higher levels of fluorescein transfer compared to droplet nuclei contaminated filtering facepiece respirators for all use scenarios (p = 0.009). Fluorescein transfer was greater for improper doffing and reuse (p = 0.007) from droplet contaminated masks compared to droplet nuclei contaminated filtering facepiece respirators and for improper doffing and reuse when compared improper doffing (p = 0.017) and proper doffing and reuse (p = 0.018) for droplet contaminated filtering facepiece respirators. For droplet nuclei contaminated filtering facepiece respirators, the difference in MS2 and fluorescein transfer did not reach statistical significance when comparing any of the use scenarios. The findings suggest that the results of fluorescein and MS2 transfer were consistent and highly correlated across the conditions of study. The data supports CDC recommendations for using proper doffing techniques and discarding filtering facepiece respirators that are directly contaminated with secretions from a cough or sneeze. PMID:28650715

Spectroscopic studies on the interaction of fluorescein and safranine T in PC liposomes

NASA Astrophysics Data System (ADS)

Bozkurt, Ebru; Bayraktutan, Tuğba; Acar, Murat; Toprak, Mahmut

2013-01-01

In this study, the fluorescence quenching of fluorescein by safranine T in liposome media had been investigated systematically by fluorescence spectroscopy, UV-vis absorption spectroscopy and fluorescence decay lifetime measurements. The spectroscopic data were analyzed using a Stern-Volmer equation to determine the quenching process. The experimental results showed that the intrinsic fluorescence of fluorescein was strongly quenched by safranine T, and that the quenching mechanism was considered as static quenching by forming a ground-complex. The Stern-Volmer quenching constant Ksv, and the bimolecular quenching constant Kq were estimated. The distances between the donor (fluorescein) and the acceptor (safranine T) were calculated according to the Förster non-radiation energy transfer theory. In addition, the partition coefficient of the safranine T (Kp) in the L-egg lecithin phosphatidylcholine liposomes was also calculated by utilizing the fluorescence quenching.

Simple Improvisation to Enhance Utility of Fluorescein Sodium in Resection of Intracranial Lesions at Routine Neurosurgical Centers.

PubMed

Gollapudi, Prakash Rao; Mohammed, Imran; Pittala, Sandeep R; Kotha, Arjun Reddy; Reddycherla, Naga Raju; Ginjupally, Dhanunjaya Rao

2018-04-01

Fluorescein sodium is one of the fluorophores that is used in the resection of intracranial lesions. It is commonly used along with a customized microscope, which is expensive and not available universally. In this study, we describe a simple, inexpensive method for better visualization of intracranial and spinal cord lesions with fluorescein. After a test dose, 20 mg/kg of fluorescein sodium was administered intravenously at the time of intubation. A blue light source was used before resection for precise localization of the intracranial lesions after durotomy. Most of the resection was done under the white light, while the blue light was used intermittently to delineate the pathologic tissue from the normal tissue and to ensure safe maximal resection. The intensity of fluorescein staining under white light and blue light was noted. The study comprised 40 cases of gliomas, meningiomas, abscesses, spinal cord tumors, and cerebellopontine angle lesions. Thirty-five lesions showed good fluorescence under the blue light, which helped us achieve better resection of the pathologic lesions. Fluorescein sodium is a safe dye; it can be used to aid in precise localization and safe maximal resection of the pathologic tissue with the help of a blue light source at any center with challenged resources. The blue light enhances the fluorescence and visualization of the pathologic tissue, and this technique can be adopted by any surgeon without much difficulty even with a basic neurosurgical setup. Copyright © 2018 Elsevier Inc. All rights reserved.

Fluorescein dye improves microscopic evaluation and counting of demodex in blepharitis with cylindrical dandruff.

PubMed

Kheirkhah, Ahmad; Blanco, Gabriela; Casas, Victoria; Tseng, Scheffer C G

2007-07-01

To show whether fluorescein dye helps detect and count Demodex embedded in cylindrical dandruff (CD) of epilated eyelashes from patients with blepharitis. Two eyelashes with CD were removed from each lid of 10 consecutive patients with blepharitis and subjected to microscopic examination with and without fluorescein solution to detect and count Demodex mites. Of 80 eyelashes examined, 36 (45%) lashes retained their CD after removal. Before addition of the fluorescein solution, the mean total Demodex count per patient was 14.9 +/- 10 and the mean Demodex count per lash was 3.1 +/- 2.5 and 0.8 +/- 0.7 in epilated eyelashes with and without retained CD, respectively (P < 0.0001). After addition of the fluorescein solution, opaque and compact CD instantly expanded to reveal embedded mites in a yellowish and semitransparent background. As a result, the mean total Demodex count per patient was significantly increased to 20.2 +/- 13.8 (P = 0.003), and the mean count per lash was significantly increased to 4.4 +/- 2.8 and 1 +/- 0.8 in eyelashes with and without retained CD (P < 0.0001 and P = 0.007), respectively. This new method yielded more mites in 8 of 10 patients and allowed mites to be detected in 3 lashes with retained CD and 1 lash without retained CD that had an initial count of zero. Addition of fluorescein solution after mounting further increases the proficiency of detecting and counting mites embedded in CD of epilated eyelashes.

Fluorescein-guided intraoperative endoscopy in patients with hereditary hemorrhagic telangiectasia: first impressions.

PubMed

Pagella, Fabio; Pusateri, Alessandro; Zaccari, Dario; Bongetta, Daniele; Zoia, Cesare; Spinozzi, Giuseppe; Olivieri, Carla; Matti, Elina

2017-03-01

Hereditary hemorrhagic telangiectasia (HHT) is a rare genetic disease that results in mucocutaneous telangiectasias and arteriovenous visceral malformations. Nasal telangiectasias lead to recurrent epistaxis, which affects up to 96% of patients. Different morphologic classifications and methods of visualization of nasal lesions have been described in the literature. We developed a new method of intraoperative endoscopy based on the intravenous administration of fluorescein. Preliminary data of this technique are reported. After the intravenous administration of sodium fluorescein, an intraoperative fluorescein-guided endoscopy was carried out using photographic customized yellow filters on top of a 0-degree, 4-mm endoscope. In 2015, 65 HHT patients underwent surgery for their epistaxis in our institution, and in 7 patients (3 males, 4 females; mean age, 54 years) an intraoperative fluorescein-guided intraoperative nasal endoscopy was performed. No adverse events or complications were observed. First impressions regarding the usage of this technique in HHT patients seem to be promising and positive in terms of efficacy and safety. However, further studies with larger cohorts of patients should be performed in order to better investigate the use of this method for diagnostic and surgical purposes in HHT. © 2016 ARS-AAOA, LLC.

Fluoride-induced modulation of ionic transport in asymmetric nanopores functionalized with "caged" fluorescein moieties.

PubMed

Ali, Mubarak; Ahmed, Ishtiaq; Ramirez, Patricio; Nasir, Saima; Cervera, Javier; Niemeyer, Christof M; Ensinger, Wolfgang

2016-04-28

We demonstrate experimentally and theoretically a nanofluidic fluoride sensing device based on a single conical pore functionalized with "caged" fluorescein moieties. The nanopore functionalization is based on an amine-terminated fluorescein whose phenolic hydroxyl groups are protected with tert-butyldiphenylsilyl (TBDPS) moieties. The protected fluorescein (Fcn-TBDPS-NH2) molecules are then immobilized on the nanopore surface via carbodiimide coupling chemistry. Exposure to fluoride ions removes the uncharged TBDPS moieties due to the fluoride-promoted cleavage of the silicon-oxygen bond, leading to the generation of negatively charged groups on the fluorescein moieties immobilized onto the pore surface. The asymmetrical distribution of these groups along the conical nanopore leads to the electrical rectification observed in the current-voltage (I-V) curve. On the contrary, other halides and anions are not able to induce any significant ionic rectification in the asymmetric pore. In each case, the success of the chemical functionalization and deprotection reactions is monitored through the changes observed in the I-V curves before and after the specified reaction step. The theoretical results based on the Nernst-Planck and Poisson equations further demonstrate the validity of an experimental approach to fluoride-induced modulation of nanopore current rectification behaviour.

Human corneal epithelial cell shedding and fluorescein staining in response to silicone hydrogel lenses and contact lens disinfecting solutions.

PubMed

Gorbet, Maud; Peterson, Rachael; McCanna, David; Woods, Craig; Jones, Lyndon; Fonn, Desmond

2014-03-01

A pilot study was conducted to evaluate human corneal epithelial cell shedding in response to wearing a silicone hydrogel contact lens/solution combination inducing corneal staining. The nature of ex vivo collected cells staining with fluorescein was also examined. A contralateral eye study was conducted in which up to eight participants were unilaterally exposed to a multipurpose contact lens solution/silicone hydrogel lens combination previously shown to induce corneal staining (renu® fresh™ and balafilcon A; test eye), with the other eye using a combination of balafilcon A soaked in a hydrogen peroxide care system (Clear Care®; control eye). Lenses were worn for 2, 4 or 6 hours. Corneal staining was graded after lens removal. The Ocular Surface Cell Collection Apparatus was used to collect cells from the cornea and the contact lens. In the test eye, maximum solution-induced corneal staining (SICS) was observed after 2 hours of lens wear (reducing significantly by 4 hours; p < 0.001). There were significantly more cells collected from the test eye after 4 hours of lens wear when compared to the control eye and the collection from the test eye after 2 hours (for both; n = 5; p < 0.001). The total cell yield at 4 hours was 813 ± 333 and 455 ± 218 for the test and control eyes, respectively (N = 5, triplicate, p = 0.003). A number of cells were observed to have taken up the fluorescein dye from the initial fluorescein instillation. Confocal microscopy of fluorescein-stained cells revealed that fluorescein was present throughout the cell cytoplasm and was retained in the cells for many hours after recovery from the corneal surface. This pilot study indicates that increased epithelial cell shedding was associated with a lens-solution combination which induces SICS. Our data provides insight into the transient nature of the SICS reaction and the nature of fluorescein staining observed in SICS.

Staining intraoperative topical solutions with fluorescein: enhancing the safety of sinus surgery.

PubMed

Yao, William C; Regone, Rachel M; Takashima, Masayoshi

2015-09-01

Oxymetazoline HCl 0.05%, 1:100,000 lidocaine with epinephrine (lido+epi), and 1:1000 epinephrine are all colorless solutions employed in sinus surgery. Because lido+epi is injected whereas others are not, care must be taken to label all solutions to avoid inadvertent injection of oxymetazoline or concentrated epinephrine because of life-threatening complications. Dyes have been used to color solutions for visual identification, but efficacy and cost have never been compared. We sought to determine the effectiveness of surgical marking pen (gentian violet) and fluorescein strips as coloring agents in mediums commonly used in sinus surgery. In this specialty techniques study, 4 liquid mediums (normal saline [NS], 0.05% oxymetazoline, 1:1,000 epinephrine, and lido+epi) and 2 coloring agents (fluorescein strip and surgical marking pen) were combined separately. Photos and absorbance of each solution were obtained at 15-minute intervals over 2 hours using a spectrophotometer. Peak absorbance remained stable for all solutions with fluorescein. Absorbance also remained constant for NS (0.37 absorbance units [AU] → 0.37 AU) and oxymetazoline (2.8 AU → 2.8 AU) mixed with marking pen. Absorbance decreased over 2 hours when marking pen was mixed with 1:1000 epinephrine (0.82 AU → 0.32 AU) and lido+epi (1.19 AU → 0.33 AU). Furthermore, the majority of color visibility and absorbance decreased in the first 15 minutes for concentrated epinephrine (0.82 → 0.33) and lido+epi (1.19 → 0.51). Cost of fluorescein strips ($0.14 to $0.20/strip) was less than marking pens ($13 to 46/pen). Compared to marking pen dye, solutions dyed with fluorescein retained their color and did not decay over time. We recommend fluorescein strips to label colorless materials in the operating room because of their lack of color decay and cost advantages. © 2015 ARS-AAOA, LLC.

Is fluorescein-guided technique able to help in resection of high-grade gliomas?

PubMed

Acerbi, Francesco; Broggi, Morgan; Eoli, Marica; Anghileri, Elena; Cavallo, Claudio; Boffano, Carlo; Cordella, Roberto; Cuppini, Lucia; Pollo, Bianca; Schiariti, Marco; Visintini, Sergio; Orsi, Chiara; La Corte, Emanuele; Broggi, Giovanni; Ferroli, Paolo

2014-02-01

Fluorescein, a dye that is widely used as a fluorescent tracer, accumulates in cerebral areas where the blood-brain barrier is damaged. This quality makes it an ideal dye for the intraoperative visualization of high-grade gliomas (HGGs). The authors report their experience with a new fluorescein-guided technique for the resection of HGGs using a dedicated filter on the surgical microscope. The authors initiated a prospective Phase II trial (FLUOGLIO) in September 2011 with the objective of evaluating the safety of fluorescein-guided surgery for HGGs and obtaining preliminary evidence regarding its efficacy for this purpose. To be eligible for participation in the study, a patient had to have suspected HGG amenable to complete resection of the contrast-enhancing area. The present report is based on the analysis of the short- and long-term results in 20 consecutive patients with HGGs (age range 45-74 years), enrolled in the study since September 2011. In all cases fluorescein (5-10 mg/kg) was injected intravenously after intubation. Tumor resection was performed with microsurgical technique and fluorescence visualization by means of BLUE 400 or YELLOW 560 filters on a Pentero microscope. The median preoperative tumor volume was 30.3 cm(3) (range 2.4-87.8 cm(3)). There were no adverse reactions related to fluorescein administration. Complete removal of contrast-enhanced tumor was achieved in 80% of the patients. The median duration of follow-up was 10 months. The 6-months progression-free survival rate was 71.4% and the median survival was 11 months. Analysis of these 20 cases suggested that fluorescein-guided technique with a dedicated filter on the surgical microscope is safe and allows a high rate of complete resection of contrast-enhanced tumor as determined on early postoperative MRI. Clinical trial registration no.: 2011-002527-18 (EudraCT).

Aggressive resection at the infiltrative margins of glioblastoma facilitated by intraoperative fluorescein guidance.

PubMed

Neira, Justin A; Ung, Timothy H; Sims, Jennifer S; Malone, Hani R; Chow, Daniel S; Samanamud, Jorge L; Zanazzi, George J; Guo, Xiaotao; Bowden, Stephen G; Zhao, Binsheng; Sheth, Sameer A; McKhann, Guy M; Sisti, Michael B; Canoll, Peter; D'Amico, Randy S; Bruce, Jeffrey N

2017-07-01

OBJECTIVE Extent of resection is an important prognostic factor in patients undergoing surgery for glioblastoma (GBM). Recent evidence suggests that intravenously administered fluorescein sodium associates with tumor tissue, facilitating safe maximal resection of GBM. In this study, the authors evaluate the safety and utility of intraoperative fluorescein guidance for the prediction of histopathological alteration both in the contrast-enhancing (CE) regions, where this relationship has been established, and into the non-CE (NCE), diffusely infiltrated margins. METHODS Thirty-two patients received fluorescein sodium (3 mg/kg) intravenously prior to resection. Fluorescence was intraoperatively visualized using a Zeiss Pentero surgical microscope equipped with a YELLOW 560 filter. Stereotactically localized biopsy specimens were acquired from CE and NCE regions based on preoperative MRI in conjunction with neuronavigation. The fluorescence intensity of these specimens was subjectively classified in real time with subsequent quantitative image analysis, histopathological evaluation of localized biopsy specimens, and radiological volumetric assessment of the extent of resection. RESULTS Bright fluorescence was observed in all GBMs and localized to the CE regions and portions of the NCE margins of the tumors, thus serving as a visual guide during resection. Gross-total resection (GTR) was achieved in 84% of the patients with an average resected volume of 95%, and this rate was higher among patients for whom GTR was the surgical goal (GTR achieved in 93.1% of patients, average resected volume of 99.7%). Intraoperative fluorescein staining correlated with histopathological alteration in both CE and NCE regions, with positive predictive values by subjective fluorescence evaluation greater than 96% in NCE regions. CONCLUSIONS Intraoperative administration of fluorescein provides an easily visualized marker for glioma pathology in both CE and NCE regions of GBM. These findings support the use of fluorescein as a microsurgical adjunct for guiding GBM resection to facilitate safe maximal removal.

O5.05FLUORESCEIN-GUIDED REMOVAL OF HIGH-GRADE GLIOMAS WITH A DEDICATED FILTER ON THE SURGICAL MICROSCOPE: PRELIMINARY RESULTS OF THE FLUOGLIO STUDY

PubMed Central

Acerbi, F.; Broggi, M.; Cavallo, C.; Anghileri, E.; Eoli, M.; Schiariti, M.; Corte, E. La; Pollo, B.; Boffano, C.; Ferroli, P.

2014-01-01

BACKGROUND: Fluorescein is a fluorescent tracer that can be used for many applications. It is able to accumulate in brain areas with blood-brain barrier disruption, and thus it can be considered an ideal dye for intraoperative visualization of high-grade gliomas (HGG). We report the preliminary results of a phase II trail (FLUOGLIO) on a new fluorescein-guided technique to remove HHG with a dedicated filter on the surgical microscope. METHODS: In September 2011 we started a prospective phase II-trial (FLUOGLIO) to evaluated safety and obtain initial indications about efficacy of fluorescein-guided surgery for HGG. Patients with suspected HGG amenable to complete resection of contrast-enhancing area were eligible to participate in this study. This report is based on the analysis of the short- and long-term results in 28 consecutive patients with HGG (age range 45-74 years), enrolled since September 2011. Fluorescein was intravenous (i.v.) injected after intubation (5-10 mg/Kg). Tumor was removed with microsurgical technique and fluorescence visualization by BLU400 or YELLOW560 filters on Pentero microscope (Carl Zeiss, Germany). The study was approved by our Ethical Committee and registered on the European Regulatory Authorities website (EudraCT No. 2011-002527-18). RESULTS: Median pre-operative tumor volume was 33.1 cm3 (2.4-87.8 cm3). We found no adverse reaction to fluorescein administration. Tumor was completely removed in 80% of the patients. Median follow-up was 10 months. 6 months Progression-free Survival (PFS) and median survival were respectively 71.4 % and 11 months. CONCLUSION: Our analysis suggested that fluorescein-guided technique with a dedicated filter on the surgical microscope is safe and allows high-rate of complete resection of contrast-enhanced tumor at the early post-operative MRI.

Detection of disseminated peritoneal tumors by fluorescein diacrylate in mice

NASA Astrophysics Data System (ADS)

Harada, Yoshinori; Furuta, Hirokazu; Murayama, Yasutoshi; Dai, Ping; Fujikawa, Yuta; Urano, Yasuteru; Nagano, Tetsuo; Morishita, Koki; Hasegawa, Akira; Takamatsu, Tetsuro

2009-02-01

Tumor invasion to the peritoneum is a poor prognostic factor in cancer patients. Accurate diagnosis of disseminated peritoneal tumors is essential to accurate cancer staging. To date, peritoneal washing cytology during laparotomy has been used for diagnosis of peritoneal dissemination of gastrointestinal cancer, but its sensitivity has not been satisfactory. Thus, a more direct approach is indispensable to detect peritoneal dissemination in vivo. Fluorescein diacrylate (FDAcr) is an esterase-sensitive fluorescent probe derived from fluorescein. In cancer cells, fluorescent fluorescein generated by exogenous application of FDAcr selectively deposits owing to its stronger hydrolytic enzyme activity and its lower leakage rate. We examined whether FDAcr can specifically detect disseminated peritoneal tumors in athymic nude mouse models. Intraperitoneally administered FDAcr revealed disseminated peritoneal microscopic tumors not readily recognized on white-light imaging. These results suggest that FDAcr is a useful probe for detecting disseminated peritoneal tumors.

Study of properties and development of sensors based on graphene films grown on SiC (0001) by thermal destruction method

NASA Astrophysics Data System (ADS)

Lebedev, A. A.; Davydov, V. Y.; Usachov, D. Y.; Lebedev, S. P.; Smirnov, A. N.; Levitskii, V. S.; Eliseyev, I. A.; Alekseev, P. A.; Dunaevskiy, M. S.; Rybkin, A. G.; Novikov, S. N.; Makarov, Yu N.

2018-01-01

The structural, chemical, and electronic properties of epitaxial graphene films grown by thermal decomposition of the Si-face of a semi-insulating 6H-SiC substrate in an argon environment are studied by Raman spectroscopy, X-ray photoelectron spectroscopy and angle-resolved photoemission. It was demonstrated the possibility of fabrication of the gas and biosensors that is based on grown graphene films. The gas sensors are sufficiently sensitive to NO2 at low concentrations. The biosensor operation was checked using an immunochemical system comprising fluorescein dye and monoclonal anti fluorescein antibodies. The sensor detects fluorescein concentration on a level of 1-10 ng/mL and bovine serum albumin- fluorescein conjugate on a level of 1-5 ng/mL. The proposed device has good prospects for use for early diagnostics of various diseases.

Evaluation of Intraocular Inflammation with Laser Flare Photometry in Behçet Uveitis.

PubMed

Yalcindag, Fatime Nilufer; Bingol Kiziltunc, Pinar; Savku, Esra

2017-02-01

To evaluate the association between intraocular inflammation and laser flare photometry measurements in Behçet disease. In total, 45 patients were included in the study. The retrospective chart reviews of patients were performed. The flare levels were compared with the grade of anterior chamber cells, the presence of vitreous cells, the complications of uveitis, and fluorescein angiography scores. The attack group had higher flare intensity; the flare levels were higher in both groups compared with the values of healthy controls. The flare levels were related to the grade of the anterior chamber cells, the presence of vitreous cells and the fluorescein angiography scores. Patients with optic atrophy and/or maculopathy also had higher values. Higher flare values were correlated with poor vision. Laser flare photometry may reduce the necessity of fluorescein angiography in monitoring subclinical inflammation and may be an indicator of posterior segment activity when fluorescein angiography is not applicable.

Corneal permeability changes in dry eye disease: an observational study.

PubMed

Fujitani, Kenji; Gadaria, Neha; Lee, Kyu-In; Barry, Brendan; Asbell, Penny

2016-05-13

Diagnostic tests for dry eye disease (DED), including ocular surface disease index (OSDI), tear breakup time (TBUT), corneal fluorescein staining, and lissamine staining, have great deal of variability. We investigated whether fluorophotometry correlated with previously established DED diagnostic tests and whether it could serve as a novel objective metric to evaluate DED. Dry eye patients who have had established signs or symptoms for at least 6 months were included in this observational study. Normal subjects with no symptoms of dry eyes served as controls. Each eye had a baseline fluorescein scan prior to any fluorescein dye. Fluorescein dye was then placed into both eyes, rinsed with saline solution, and scanned at 5, 10, 15, and 30 min. Patients were administered the following diagnostic tests to correlate with fluorophotometry: OSDI, TBUT, fluorescein, and lissamine. Standard protocols were used. P < 0.05 was considered significant. Fifty eyes from 25 patients (DED = 22 eyes, 11 patients; Normal = 28 eyes, 14 patients) were included. Baseline scans of the dry eye and control groups did not show any statistical difference (p = 0.84). Fluorescein concentration of DED and normal patients showed statistical significance at all time intervals (p < 10(-5), 0.001, 0.002, 0.049 for 5, 10, 15, & 30 min respectively). Fluorophotometry values converged towards baseline as time elapsed, but both groups were still statistically different at 30 min (p < 0.01). We used four fluorophotometry scoring methods and correlated them with OSDI, TBUT, fluorescein, and lissamine along with adjusted and aggregate scores. The four scoring schemes did not show any significant correlations with the other tests, except for correlations seen with lissamine and 10 (p = 0.045, 0.034) and 15 min (p = 0.013, 0.012), and with aggregate scores and 15 min (p = 0.042, 0.017). Fluorophotometry generally did not correlate with any other DED tests, even though it showed capability of differentiating between DED and normal eyes up to 30 min after fluorescein dye instillation. There may be an aspect of DED that is missed in the current regimen of DED tests and only captured with fluorophotometry. Adding fluorophotometry may be useful in screening, diagnosing, and monitoring patients with DED.

CAPILLARY ELECTROPHORESIS/LASER-INDUCED FLUORESCENCE DETECTION OF FLUORESCEIN AS A GROUNDWATER MIGRATION TRACER

EPA Science Inventory

Capillary electrophoresis (CE) has been applied to the determination of the groundwater migration tracer dye fluorescein based on laser-induced fluorescence (LIF) detection and compared to determinations obtained with traditional spectrofluorimetry. Detection limits of injected d...

Fluorescein transport properties across artificial lipid membranes, Caco-2 cell monolayers and rat jejunum.

PubMed

Berginc, Katja; Zakelj, Simon; Levstik, Lea; Ursic, Darko; Kristl, Albin

2007-05-01

Membrane transport characteristics of a paracellular permeability marker fluorescein were evaluated using artificial membrane, Caco-2 cell monolayers and rat jejunum, all mounted in side-by-side diffusion cells. Modified Ringer buffers with varied pH values were applied as incubation salines on both sides of artificial membrane, cell culture monolayers or rat jejunum. Passive transport according to pH partition theory was determined using all three permeability models. In addition to that, active transport of fluorescein in the M-S (mucosal-to-serosal) direction through rat jejunum was observed. The highest M-S P(app) values regarding the active transport through the rat jejunum were observed in incubation saline with pH 6.5. Fluorescein transport through the rat jejunum was inhibited by DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid) and alpha-CHC (alpha-cyano-4-hydroxycinnamic acid). Thus, we assume that two pH-dependent influx transporters could be involved in the fluorescein membrane transport through the intestinal (jejunal) epithelium. One is very likely an MCT (monocarboxylic acid cotransporter) isoform, inhibited by specific MCT inhibitor alpha-CHC, while the involvement of the second one with overlapping substrate/inhibitor specificities (most probably a member of the organic anion-transporting polypeptide family, inhibited at least partially by DIDS) could not be excluded.

Caseins from bovine colostrum and milk strongly bind piscidin-1, an antimicrobial peptide from fish.

PubMed

Kütt, Mary-Liis; Stagsted, Jan

2014-09-01

A model system of bovine colostrum and piscidin, a fish-derived antimicrobial peptide, was developed to study potential interactions of antimicrobial peptides in colostrum. We did not detect any antimicrobial activity of colostrum using the radial plate diffusion assay; in fact colostrum completely abrogated activity of added piscidin. This could not be explained by degradation of piscidin by colostrum, which was less than ten percent. We found that colostrum even protected piscidin against degradation by added proteases. We further observed that colostrum and milk rapidly quenched the fluorescence of fluorescein-piscidin but not that of fluorescein. This effect was not seen with BSA and the specific quenching of fluorescein-piscidin by colostrum was saturably inhibited with unlabeled piscidin. Size exclusion chromatography indicated that fluorescein-piscidin bound to casein micelles with no apparent binding to IgG or whey proteins. Further, addition of pure caseins was able to quench fluorescence of fluorescein-piscidin and to inhibit the antimicrobial activity of piscidin. The interaction between caseins and piscidin could be dissociated by guanidine hydrochloride and recovered piscidin had antimicrobial activity against bacteria. Based on our results we propose that caseins could be carriers for antimicrobial peptides in colostrum and milk. Copyright © 2014 Elsevier B.V. All rights reserved.

Determination of torasemide by fluorescence quenching method with some dihalogenated fluorescein dyes as probes

NASA Astrophysics Data System (ADS)

Cui, Zhiping; Liu, Shaopu; Liu, Zhongfang; Li, Yuanfang; Hu, Xiaoli; Tian, Jing

2013-10-01

A novel fluorescence quenching method for the determination of torasemide (TOR) with some dihalogenated fluorescein dyes as fluorescence probes was developed. In acidulous medium, TOR could interact with some dihalogenated fluorescein dyes such as dichlorofluorescein (DCF), dibromofluorescein (DBF) and diiodofluorescein (DIF) to form binary complexes, which could lead to fluorescence quenching of above dihalogenated fluorescein dyes. The maximum fluorescence emission wavelengths were located at 532 nm (TOR-DCF), 535 nm (TOR-DBF) and 554 nm (TOR-DIF). The relative fluorescence intensities (ΔF = F0 - F) were proportional to the concentration of TOR in certain ranges. The detection limits were 4.8 ng mL-1 for TOR-DCF system, 9.8 ng mL-1 for TOR-DBF system and 35.1 ng mL-1 for TOR-DIF system. The optimum reaction conditions, influencing factors were studied; and the effect of coexisting substances was investigated owing to the highest sensitivity of TOR-DCF system. In addition, the reaction mechanism, composition and structure of the complex were discussed by quantum chemical calculation and Job's method. The fluorescence quenching of dihalogenated fluorescein dyes by TOR was a static quenching process judging from the effect of temperature and the Stern-Volmer plots. The method was satisfactorily applied to the determination of TOR in tablets and human urine samples.

DETERMINATION OF ALIPHATIC AMINES IN WATER USING DERIVATIZATION WITH FLUORESCEIN ISOTHIOCYANATE AND CAPILLARY ELECTROPHORESIS/LASER-INDUCED FLUORESCENCE DETECTION.

EPA Science Inventory

Detection-oriented derivatization of aliphatic amines and amine functional groups in coumpounds of environmental interest was studied using fluorescein isothiocyanate (FITC) with separation/determination by capillary electrophoresis/laser-induced fluorescence. Determinative level...

Low-dose intrathecal fluorescein for diagnosis of cerebrospinal fluid rhinorrhea using the scanning fiber endoscope in the human nasal cavities

NASA Astrophysics Data System (ADS)

Hou, Vivian W.; Davis, Calvin G.; Davis, Greg E.; Seibel, Eric J.

2016-03-01

Intrathecal fluorescein (ITF) enhances detection of cerebrospinal fluid rhinorrhea (CSFR). Clinically administered doses fall in the range of 0.1ml to 0.5ml of 5% to 10% fluorescein (1.3×10-3M to 1.3×10-2M). Though uncommon, significant morbidities associated with high doses of fluorescein have been reported. High concentrations are necessary for white light visual assessment; in contrast, fluorescent imaging enhances signal contrast and requires lower ITF concentrations for visualization. The ultrathin and flexible, multimodal scanning fiber endoscope (SFE) can visualize nanomolar concentrations of fluorescein as pseudocolor over reflectance, video-rate imaging. The application of the SFE for CSFR detection was assessed in a cadaver study. Briefly, 10μM (1×10-5M) fluorescein, 100X-1000X less than the standard clinical dose, was injected intra-cranially into the epidural space through an orbital roof puncture. The resulting rhinorrhea was assessed with a conventional, rigid ENT scope and second with the SFE in both video reflectance and multimodal fluorescent imaging modes. Neither system could visualize the 10μM ITF during white light imaging however the nanomolar sensitive SFE visualized the rhinorrhea during fluorescent imaging. Despite the low concentration used, a target-to-background ratio of 5.6 +/- 2.7 was achieved. To demonstrate SFE guidance of CSFR detection and repair, de-identified patient computed tomography (CT) scans were used to generate 3D printed phantoms. Cases were selected for unique anatomical features and overall clinical difficulty as determined by an experienced ENT clinician (GED). The sensitivity and minimally invasive nature of the SFE provide a unique platform for enhancing diagnosis and monitoring interventions in surgical endoscopic approaches into the sinuses.

Temporal Characteristics of Sodium Fluorescein in the Tear Meniscus.

PubMed

Markoulli, Maria; Isa, Nur Amalina M D; Papas, Eric B

2017-02-01

To observe the emission intensity profile of sodium fluorescein in the human tear film as a function of time and concentration. Twenty-two participants with no dry eye signs or symptoms were randomly allocated to receive 1 μL of either a 2 or 10% concentration of fluorescein to one eye. Images of the inferior tear meniscus were captured at regular intervals over 30 minutes and the process repeated for the other eye with the alternate concentration. Fluorescence intensity was quantified on the basis of the grayscale pixel values in the tear meniscus images. The fluorescein-decay profile over time and between concentrations was determined. Peak fluorescence intensity was reached in 3.9 ± 3.0 and 8.7 ± 4.4 minutes after instillation for the 2 and 10% concentrations, respectively. The 10% concentration of fluorescein maintained its peak fluorescence intensity longer than the 2% concentration (about 9 and 2 minutes, respectively). The peak fluorescence intensity was not significantly different between the higher and lower concentrations (44 ± 37 vs. 38 ± 32 units, P = .22). For both concentrations, the observed intensity did not return to baseline levels by the end of the 30-minute observation time. The fluorescence intensity of fluorescein in a clinical setting varies with time such that both the onset and duration of maximum brightness are concentration dependent. At low concentration (2%), maximum brightness occurs almost immediately after instillation and lasts about 2 minutes. With a higher concentration (10%), the effective working window is delayed for about 7 to 8 minutes. Irrespective of initial concentration, observable fluorescence remains in the tear film beyond 30 minutes post-instillation.

Quantitative Spatial and Temporal Analysis of Fluorescein Angiography Dynamics in the Eye

PubMed Central

Hui, Flora; Nguyen, Christine T. O.; Bedggood, Phillip A.; He, Zheng; Fish, Rebecca L.; Gurrell, Rachel; Vingrys, Algis J.; Bui, Bang V.

2014-01-01

Purpose We describe a novel approach to analyze fluorescein angiography to investigate fluorescein flow dynamics in the rat posterior retina as well as identify abnormal areas following laser photocoagulation. Methods Experiments were undertaken in adult Long Evans rats. Using a rodent retinal camera, videos were acquired at 30 frames per second for 30 seconds following intravenous introduction of sodium fluorescein in a group of control animals (n = 14). Videos were image registered and analyzed using principle components analysis across all pixels in the field. This returns fluorescence intensity profiles from which, the half-rise (time to 50% brightness), half-fall (time for 50% decay) back to an offset (plateau level of fluorescence). We applied this analysis to video fluorescein angiography data collected 30 minutes following laser photocoagulation in a separate group of rats (n = 7). Results Pixel-by-pixel analysis of video angiography clearly delineates differences in the temporal profiles of arteries, veins and capillaries in the posterior retina. We find no difference in half-rise, half-fall or offset amongst the four quadrants (inferior, nasal, superior, temporal). We also found little difference with eccentricity. By expressing the parameters at each pixel as a function of the number of standard deviation from the average of the entire field, we could clearly identify the spatial extent of the laser injury. Conclusions This simple registration and analysis provides a way to monitor the size of vascular injury, to highlight areas of subtle vascular leakage and to quantify vascular dynamics not possible using current fluorescein angiography approaches. This can be applied in both laboratory and clinical settings for in vivo dynamic fluorescent imaging of vasculature. PMID:25365578

Micro-instillation of fluorescein with an inoculation loop for ocular surface staining in dry eye syndrome.

PubMed

Courrier, Emilie; Renault, Didier; Kaspi, Mathilde; Marcon, Agathe; Lambert, Victor; Garcin, Thibaud; Chiambaretta, Frederic; Garhofer, Gerhard; Thuret, Gilles; Gain, Philippe

2018-03-01

To describe and validate the micro-instillation of fluorescein on the ocular surface by a disposable calibrated inoculation loop to improve corneal and conjunctival staining quality. Accuracy and precision of the volume of 0.5% sodium fluorescein collected by a single use 1 μl-calibrated inoculation loop were measured using a precision balance. Twenty patients (40 eyes) suffering from dry eye syndrome were enrolled in a prospective interventional nonrandomized study. Fluorescein was instilled with the loop, and slit-lamp images were taken within 30 seconds using cobalt blue light with and without a yellow barrier filter. For comparison, after a washout period, the same images were retaken after instillation of one drop of fluorescein from a single-dose unit. The main outcome measure was the staining quality assessed by three experts, blind to the instillation method. Patient discomfort (tolerance, by a questionnaire) was also compared. The mean volume collected by the loop was 1.18 ± 0.12 μl, compared with 33.70 ± 6.10 μl using the single-dose unit. The loop avoided excess dye responsible for unpleasant tearing, masking of lesions and rapid diffusion into the stroma. Micro-instillation greatly improved image quality without losing information. The yellow filter further improved image contrast. Tolerance was excellent. The 1 μl-calibrated inoculation loop is a safe, convenient, inexpensive, disposable, sterile, well-tolerated tool for reproducible micro-instillation of commercial fluorescein. By greatly improving staining quality, it will help standardize assessment of dry eye severity. © 2017 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

Fluorescein-Guided Resection of Intramedullary Spinal Cord Tumors: Results from a Preliminary, Multicentric, Retrospective Study.

PubMed

Acerbi, Francesco; Cavallo, Claudio; Schebesch, Karl-Michael; Akçakaya, Mehmet Osman; de Laurentis, Camilla; Hamamcioglu, Mustafa Kemal; Broggi, Morgan; Brawanski, Alexander; Falco, Jacopo; Cordella, Roberto; Ferroli, Paolo; Kiris, Talat; Höhne, Julius

2017-12-01

Intramedullary spinal cord tumors (IMSCTs) are rare, heterogenous lesions that are usually enhanced on preoperative magnetic resonance imaging (MRI) because of a damaged blood-brain barrier. Sodium fluorescein is a dye that accumulates in areas of the central nervous system with a damaged BBB. Given the pattern of MRI contrast enhancement of the majority of IMSCTs, the use of this fluorescent tracer could improve tumor visualization and quality of resection. In this article, we present the first experience with the application of fluorescein-guided technique for surgical removal of IMSCTs. Eleven patients (6 men, 5 women; mean age, 50.1 years), harboring 5 ependymomas, 3 hemangioblastomas, 1 astrocytoma, 1 pilocytic astrocytoma, and 1 glioneuronal tumor forming rosettes were included. Sodium fluorescein (5 mg/kg) was injected immediately after patient intubation. Tumors were removed with microsurgical technique and standard neurophysiological monitoring, under YELLOW 560 filter (Pentero 900) visualization. Surgical reports were reviewed regarding usefulness and grade of fluorescein staining. Postoperative MRI was performed within 72 hours after surgery, and postoperative clinical outcome was registered. No adverse events were registered. Fluorescent staining was reported in 9 of 11 cases (82%), all of them enhancing on preoperative MRI (100% of ependymomas, 100% of pilocytic astrocytomas, 100% of hemangioblastomas). No fluorescence was reported in 1 astrocytoma and 1 glioneuronal tumor-forming rosette. Intraoperative fluorescence was considered helpful for tumor resection in 9 of 11 cases (82%). Gross total resection was obtained in 8 of 11 cases (72.7%). Our results suggest that fluorescein-guided surgery is a safe and effective technique that can be used during the surgical resection of IMSCTs presenting with contrast-enhancement on preoperative MRI. Copyright © 2017 Elsevier Inc. All rights reserved.

Validity of fluorexon disodium versus sodium fluorescein for use in Goldmann tonometry.

PubMed

Ng, Loretta T; Tong, Judy W; De Land, Paul N

2006-07-01

To evaluate the safety, validity, and comfort of 0.35% fluorexon disodium and 0.4% benoxinate (Flura-Safe) compared with the gold standard of 0.25% sodium fluorescein and 0.4% benoxinate for Goldmann applanation tonometry (GAT). This was a double-masked, randomized, crossover clinical trial. Subjects received either the standard or study formulation for GAT on visit 1 and the other formulation 1 week later. At each visit, tonometer mire quality, adequacy of fluorescence, ease of intraocular pressure (IOP) measurements, the IOP value, and anesthetizing efficacy of the formulation were assessed. Subjects graded general comfort, soreness and irritation, and burning and stinging of each formulation at 1 and 5 minutes after drop instillation. Sixty-seven subjects completed the study. The mean IOP was 13.9 +/- 2.7 with fluorexon and 13.9 +/- 2.8 mm Hg with fluorescein OD and 14.0 +/- 2.8 with fluorexon and 13.9 +/- 2.5 mm Hg with fluorescein OS. The measurements with the 2 formulations were highly correlated for OD and OS, and the differences between the 2 measurements were not clinically significant. There was also no significant difference between the 2 drops in mire clarity, adequacy of fluorescence, or corneal anesthesia. However, fluorexon was statistically more comfortable (P = 0.039) and caused less stinging and burning (P = 0.014) at 1 minute versus the fluorescein formulation. Not only was the new fluorexon product accurate and effective in GAT, it was also statistically more comfortable and had a less stinging and burning effect at 1 minute after drop instillation than the traditional fluorescein formulation. Because fluorexon is less likely to stain soft contact lenses, this may be the dye-anesthetic formulation of choice for practices that routinely perform GAT.

Resonance energy transfer between sites in rat liver glutathione S-transferase, 1-1, selectively modified at cysteine-17 and cysteine-111.

PubMed

Hu, L; Colman, R F

1997-02-18

Monobromobimane (mBBr) can label both Cys111 and Cys17 of rat liver glutathione S-transferase, 1-1 (GST 1-1). However, selective modification of Cys111 was achieved by the maleimide-based sulfhydryl reagents N-ethylmaleimide (NEM) and fluorescein 5-maleimide (NFM). Incubation of GST 1-1 with 5 mM NEM for 30 min at pH 7.5 and 25 degrees C leads to the formation of modified enzyme with 92% residual activity toward 1-chloro-2,4-dinitrobenzene and completely blocks Cys111 from subsequent reaction with either NFM or mBBr. Reaction of GST 1-1 with 0.2 mM NFM under the same conditions affords a modified enzyme with only 14% residual activity even though NFM and NEM target the same Cys111. The results indicate that when the bulky fluorescein is covalently bound to Cys111, the ligand projects into both the xenobiotic binding site and the glutathione site. After NEM or NFM modification of GST 1-1, the enzyme was further modified by monobromobimane at Cys17 with loss of activity. Together with the only tryptophan (Trp20), fluorescein linked to Cys111 and bimane to Cys17 provide three fluorescent probes to study the solution structure of GST 1-1. Fluorescence spectral analysis suggests that Trp20 and bimane linked to Cys17 are located in a relatively hydrophobic environment, while fluorescein linked to Cys111 is located in a charged environment. These fluorescent probes constitute three sets of donor-acceptor pairs for the measurement of fluorescence energy transfer, and distances calculated from such measurements are 20 A between Trp20 and bimane at Cys17, 19 A between Trp20 and fluorescein at Cys111, and < 22 A between bimane at Cys17 and fluorescein at Cys111. Molecular modeling studies indicate that fluorescein lies between the two subunits, is surrounded by charged residues, and is extended into the xenobiotic binding site. They also suggest that mBBr must approach from the dimer interface in order to reach the reaction site at Cys17.

Azinphos-methyl residues in apples and spatial distribution of fluorescein in vase-shaped apple trees.

PubMed

Bélanger, A; Bostanian, N J; Boivin, G; Boudreau, F

1991-06-01

Vase-shaped standard apple trees cv. McIntosh were sprayed with azinphos-methyl at pink, pink and 1st cover and 1st cover only. Residue analyses by gas chromatography revealed detectable residues on foliage until mid summer. At harvest, negligible residue levels were found on the peel and the whole apple. On four trees, fluorescein was sprayed in the same manner as the insecticide and maximum levels of the dye were detected on the outside lower canopy along the row. Minimal concentration of fluorescein was detected on the inner upper canopy away from the direction of the row.

Fluorescence Imaging/Agents in Tumor Resection.

PubMed

Stummer, Walter; Suero Molina, Eric

2017-10-01

Intraoperative fluorescence imaging allows real-time identification of diseased tissue during surgery without being influenced by brain shift and surgery interruption. 5-Aminolevulinic acid, useful for malignant gliomas and other tumors, is the most broadly explored compound approved for fluorescence-guided resection. Intravenous fluorescein sodium has recently received attention, highlighting tumor tissue based on extravasation at the blood-brain barrier (defective in many brain tumors). Fluorescein in perfused brain, unselective extravasation in brain perturbed by surgery, and propagation with edema are concerns. Fluorescein is not approved but targeted fluorochromes with affinity to brain tumor cells, in development, may offer future advantages. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Fluorescein sodium-guided surgery of a brain abscess: A case report

PubMed Central

Höhne, Julius; Brawanski, Alexander; Schebesch, Karl-Michael

2016-01-01

Background: Up to now, the feasibility and benefit of using fluorescein sodium under a dedicated surgical microscope filter (YE560, YELLOW 560 nm filter, Carl Zeiss Meditec, Germany) has never been clinically evaluated in infectious disorders of the brain. Case Description: Here, we report the case of a male patient with a brain abscess in the right parietal lobe that was removed under fluorescence-guidance (intravenous administration of fluorescein sodium 10%, 5 mg/kg bodyweight). The abscess capsule showed intensive yellow fluorescent staining, while − under white light − the cortex appeared normal. Conclusion: This technique may improve the identification and surgical removal of brain abscesses. PMID:28031990

The use of lysozyme modified with fluorescein for the detection of Gram-positive bacteria.

PubMed

Arabski, Michał; Konieczna, Iwona; Tusińska, Ewa; Wąsik, Sławomir; Relich, Inga; Zając, Krzysztof; Kamiński, Zbigniew J; Kaca, Wiesław

2015-01-01

Lysozyme (1,4-β-N-acetylmuramidase) is commonly applied in the food, medical, and pharmaceutical industries. In this study, we tested a novel application of fluorescein-modified lysozyme (using carboxyfluorescein with a triazine-based coupling reagent) as a new tool for the detection of Gram-positive soil bacteria. The results, obtained by cultivation methods, fluorescence analysis, and laser interferometry, showed that, after optimization, fluorescein-modified lysozyme could be used to evaluate the prevalence of Gram-positive bacteria essential in bioremediation of soils with low pH, such as those degraded by sulfur. Copyright © 2014 Elsevier GmbH. All rights reserved.

The significance of fluorescein angiography in the early diagnosis of lesions on ocular fundus at pseudoxanthoma elasticum patients.

PubMed

Bogdanowski, T; Gluza, J; Rasiewicz, D

1977-05-27

The role of fluorescein angiography in early diagnosis of lesions on fundus of the eye at pseudoxanthoma elasticum patients has been shown. The authors show angiographic changes on the basis of three cases of pseudoxanthoma elasticum patients.

Italian multicentre study on intrathecal fluorescein for craniosinusal fistulae.

PubMed

Felisati, G; Bianchi, A; Lozza, P; Portaleone, S

2008-08-01

Cerebrospinal fluid leak (CSF), clinical sign of a dural lesion of the skull base, is a relatively rare event that can present with a variety of symptoms. Every craniosinus fistula should be considered a serious, potentially life-threatening situation (even those cases with hidden CSF leak). Reports of experience concerning diagnosis and treatment of craniosinus fistulae have appeared in the Literature. In the last few years, the endoscopic nasal approach is proving effective as it makes diagnosis much easier and is the least invasive surgical approach, with the greatest percentage of success. Various classifications are being proposed to improve clinical evaluation of CSF leaks and to simplify the diagnostic and therapeutic approach. The most common parameters of classification are: aetiology (traumatic, iatrogenic, non-traumatic, etc.) site, type of flow (high or low pressure) and, as far as concerns treatment, the type of graft used, all of which have contributed to various diagnostic and therapeutic algorithms being proposed. Therefore, the subject seems to be widely schematized and the therapeutic attitude widely agreed. However, one of the diagnostic and therapeutic approaches is now being questioned. For some, it is the heart of the clinical approach, while for others, it is a useful tool yet too dangerous to be used on account of potential side effects: namely, the fluorescein test. This procedure, consisting of intrathecal injection of a colorant (fluorescein), is well known by the Food and Drug Administration (FDA) which neither explicitly prohibits it, nor allows it, intrathecal administration is, therefore, an off label use. As far as the Authors know, authorization of this procedure has not been forthcoming anywhere in the world although the procedure itself is widely employed. As far as concerns the use of intrathecal fluorescein, many scientific papers have been written, clearly supporting its clinical usefulness. One limit to the use of fluorescein derives from frequent reports of complications, often related to the intrathecal administration; such complications are, however, always due to an incorrect dosage. In order to perform correct monitoring of any complication related to the use of intrathecal fluorescein and to investigate in a strictly scientific fashion, the legal problem related to the off label use (intrathecal administration) of an authorised substance, the Authors coordinated an Italian multicentre study aimed at establishig the tolerability of the lumbar intrathecal administration of fluorescein. Aim of the study was to review the literature focusing on CSF leaks, to set up to date diagnostic and therapeutic indications of fluorescein and to report the preliminary results of the Italian multicentre study.

General Synthetic Method for Si-Fluoresceins and Si-Rhodamines

PubMed Central

2017-01-01

The century-old fluoresceins and rhodamines persist as flexible scaffolds for fluorescent and fluorogenic compounds. Extensive exploration of these xanthene dyes has yielded general structure–activity relationships where the development of new probes is limited only by imagination and organic chemistry. In particular, replacement of the xanthene oxygen with silicon has resulted in new red-shifted Si-fluoresceins and Si-rhodamines, whose high brightness and photostability enable advanced imaging experiments. Nevertheless, efforts to tune the chemical and spectral properties of these dyes have been hindered by difficult synthetic routes. Here, we report a general strategy for the efficient preparation of Si-fluoresceins and Si-rhodamines from readily synthesized bis(2-bromophenyl)silane intermediates. These dibromides undergo metal/bromide exchange to give bis-aryllithium or bis(aryl Grignard) intermediates, which can then add to anhydride or ester electrophiles to afford a variety of Si-xanthenes. This strategy enabled efficient (3–5 step) syntheses of known and novel Si-fluoresceins, Si-rhodamines, and related dye structures. In particular, we discovered that previously inaccessible tetrafluorination of the bottom aryl ring of the Si-rhodamines resulted in dyes with improved visible absorbance in solution, and a convenient derivatization through fluoride-thiol substitution. This modular, divergent synthetic method will expand the palette of accessible xanthenoid dyes across the visible spectrum, thereby pushing further the frontiers of biological imaging. PMID:28979939

Spectral optical coherence tomography vs. fluorescein pattern for rigid gas-permeable lens fit.

PubMed

Piotrowiak, Ilona; Kaluzny, Bartłomiej Jan; Danek, Beata; Chwiędacz, Adam; Sikorski, Bartosz Lukasz; Malukiewicz, Grażyna

2014-07-04

This study aimed to evaluate anterior segment spectral optical coherence tomography (AS SOCT) for assessing the lens-to-cornea fit of rigid gas-permeable (RGP) lenses. The results were verified with the fluorescein pattern method, considered the criterion standard for RGP lens alignment evaluations. Twenty-six eyes of 14 patients were enrolled in the study. Initial base curve radius (BCR) of each RGP lens was determined on the basis of keratometry readings. The fluorescein pattern and AS SOCT tomograms were evaluated, starting with an alignment fit, and subsequently, with BCR reductions in increments of 0.1 mm, up to 3 consecutive changes. AS SOCT examination was performed with the use of RTVue (Optovue, California, USA). The average BCR for alignment fits, defined according to the fluorescein pattern, was 7.8 mm (SD=0.26). Repeatability of the measurements was 18.2%. BCR reductions of 0.1, 0.2, and 0.3 mm resulted in average apical clearances detected with AS SOCT of 12.38 (SD=9.91, p<0.05), 28.79 (SD=15.39, p<0.05), and 33.25 (SD=10.60, p>0.05), respectively. BCR steepening of 0.1 mm or more led to measurable changes in lens-to-cornea fits. Although AS SOCT represents a new method of assessing lens-to-cornea fit, apical clearance detection with current commercial technology showed lower sensitivity than the fluorescein pattern assessment.

Influence of ionization states of antigen on anti-fluorescein antibodies

NASA Astrophysics Data System (ADS)

Fukunishi, Hiroaki

2012-10-01

Ratios of anion and di-anion states of fluorescein (FLU(-1) and FLU(-2)) are 21.2% and 78.8%, respectively, in the neutral pH. We investigated the influence of ionization states of antigen on anti-fluorescein antibodies. For this purpose, steered molecular dynamics (SMD) simulations were performed. Potential of mean forces (PMF) based on Jarzynski equality showed that wild-type (4-4-20) more strongly binds to FLU(-1) than FLU(-2), whereas its femtomolar-affinity mutant (4M5.3) more strongly binds to FLU(-2) than FLU(-1). It was speculated that the environment or the process of in vivo antibody production had been different from those of the protein engineering.

21 CFR 74.1321 - D&C Red No. 21.

Code of Federal Regulations, 2010 CFR

2010-04-01

... ADDITIVES SUBJECT TO CERTIFICATION Drugs § 74.1321 D&C Red No. 21. (a) Identity. (1) The color additive D&C... (CAS Reg. No. 25709-84-6). The color additive is manufactured by brominating fluorescein with elemental... anhydride. The fluorescein is isolated and partially purified prior to bromination. (2) Color additive...

Immunochemical Parameters of Some Commercial Conjugates for the Fluorescent Treponemal Antibody-Absorption Test

PubMed Central

Hunter, E. F.; Smith, J. F.; Lewis, J. S.; McGrew, B. E.; Schmale, J. D.

1972-01-01

Fluorescein-labeled anti-human globulins were examined to determine the need for standardization of conjugates used in the fluorescent treponemal antibody-absorption (FTA-ABS) test. Twenty-one of 33 conjugates submitted by commercial manufacturers to the Reagents Control Activity, Venereal Disease Research Laboratory, for evaluation in the FTA-ABS test were available for study. Conjugates, after evaluation in FTA-ABS performance tests, were examined by immunoelectrophoresis, by titration against immunoglobulins G and M (IgG, IgM) with FTA-ABS techniques, and by the biuret protein and fluorescein diacetate methods for determining fluorescein to protein (F/P) ratios. The conjugates were predominately anti-IgG globulin with anti-light-chain activity. Differences were noted in the ability of some conjugates to detect IgM antibody. The F/P ratios of those conjugates that could be determined varied from 2.6 to 17.8 μg of fluorescein per mg of protein. The need to identify and standardize both the immunologic capabilities and the optimum F/P ratio for FTA-ABS test conjugates is presented. PMID:4564403

Fluorescein angiography and retinal vascular development in premature infants.

PubMed

Purcaro, Velia; Velia, Purcaro; Baldascino, Antonio; Antonio, Baldascino; Papacci, Patrizia; Patrizia, Papacci; Giannantonio, Carmen; Carmen, Giannantonio; Molisso, Anna; Anna, Molisso; Molle, Fernando; Fernando, Molle; Lepore, Domenico; Domenico, Lepore; Romagnoli, Costantino; Costantino, Romagnoli

2012-10-01

To investigate the role of fluorescein angiography (FA) in the management of retinopathy of prematurity (ROP) in preterm newborns. An observational case series of 13 extremely low birth weight infants. From September 2009 to March 2010, 13 newborn infants with a gestational age <29 weeks end/or birth weight <1000 g underwent serial fluorescein angiography with RetCam (Clarity, Pleasanton, CA) every 2 weeks. The fluorescein angiograms were examined to optimize the timing of diagnosis of ROP and to investigate development of retinal and choroidal vascularization. There were no side effects related to FA. Variable features of retinal and choroidal circulation in preterm infants with a high risk of developing ROP were noted. FA allows vessels branching at the junction between vascular and avascular retina (V-Av junction) to be viewed easily and shows the ROP findings that sometimes cannot be seen by indirect ophthalmoscopy. Dye leakage is the most significant sign of progression to severe ROP or the need for surgery in newborn babies with ROP. RetCam-assisted intravenous FA is safe and allows a more objective assessment of the ROP stage and zone.

[Evaluation of transfection effectiveness using fluorescein-labelled oligonucleotides and entraster-R siRNA transfection into Plasmodium falciparum].

PubMed

Zhou, Hong-Chang; Gao, Yu-Hui; Shao, Sheng-Wen; Zhang, Hui; Zhang, Ting

2013-12-01

The cultured Plasmodium falciparum parasites were synchronized twice by 5% sorbitol treatment twice (8-hour window), and then incubated at 37 degrees C for 16 h. Parasites were transfected with fluorescein-labelled oligonucleotides (group A) or fluorescein-labelled oligonucleotides+Entranster-R siRNA transfection reagent (group B). After 5 h a part of parasites was evaluated by fluorescence microscopy and flow cytometry. The rest of parasites were washed with RPMI 1640 medium, and then incubated with 500 microl new medium containing 2% fresh erythrocytes for another 12 h, and detected by flow cytometry. The fluorescein-labelled oligonucleotides were localized in erythrocytes in group B, but nearly no fluorescence was observed for group A. Flow cytometry analysis indicated that the transfection efficiency of group B [(47.40 +/- 3.39)%] was higher than that of group A [(0.60 +/- 0.27)%]. In the second cell cycle, the transfection efficiency in group B was (26.85 +/- 2.90)%, while that of group A was nearly zero. The results indicated that Entranster-R siRNA transfection reagent may increase the oligonucleotides transfection efficiency.

Potential bacterial contamination of eyedrops used for tonometry.

PubMed

Palmberg, R; Gutierrez, Y S; Miller, D; Feuer, W J; Anderson, D R

1994-05-15

We compared the potential for bacterial contamination of a proparacaine hydrochloride solution preserved with benzalkonium chloride, which is used with fluorescein paper for tonometry, to that of a fluorescein-benoxinate hydrochloride combination solution preserved with chlorobutanol. We contaminated bottles of each solution with Pseudomonas aeruginosa or Staphylococcus aureus (10(7) organisms per milliliter of eyedrop solution). From the fluorescein-benoxinate hydrochloride solution, Staphylococcus organisms were cultured in declining numbers over time, with a half-life of nine seconds, and no Staphylococcus organisms were recovered after five minutes. Pseudomonas organisms disappeared from this solution within 15 seconds. Neither species was viable after 15 seconds in the proparacaine hydrochloride solution. Additionally, we cultured 12 bottles of fluorescein-benoxinate hydrochloride and 15 bottles of proparacaine hydrochloride that had been in use in the clinic for one month. No organisms were recovered from a drop dispensed from any bottle or from the tips, caps, rims, or solution from within any bottle, except that five colonies of Staphylococcus grew from the external rim of one bottle. We conclude that both solutions sterilize themselves rapidly and effectively. Either may be used safely for tonometry.

STUDIES ON FLUORESCENT ANTIBODY STAINING

PubMed Central

Goldstein, Gerald; Slizys, Irene S.; Chase, Merrill W.

1961-01-01

1. A study has been made of the non-specific fluorescent staining of splenic imprints treated with fluorescent sheep antibody globulins. 2. In tissue imprints made with the spleens of antigen-stimulated animals, no morphological distinction was evident between areas showing non-specific fluorescence and specific fluorescence. 3. Elimination of non-specific fluorescence was not achieved by any one, or any combination of the following: (a) conjugating only gamma globulins with fluorescein isothiocyanate; (b) removal of dialyzable fluorescent products on sephadex, followed by concentration through the use of pressure dialysis; (c) use of crystalline preparations of fluorescein isothiocyanate. 4. Individual preparations of fluorescent antibodies were separated by gradient elution chromatography on diethylaminoethyl (DEAE) cellulose into fractions possessing different numbers of fluorescein radicals per molecule of globulin. 5. The coupling ratio of 50 mg fluorescein isothiocyanate (FITC) per gm of protein, as commonly advocated, can not be recommended for the precise localization of antibody globulin in tissues owing to the capacity of the coupled products to give non-specific fluorescent staining. When crystalline preparations of FITC are used instead of the amorphous product at 50 mg/gm protein, far too high non-specific fluorescence results. 6. A fraction with bright specific fluorescence and no or negligible nonspecific fluorescence was obtained from each fluorescent antibody that was prepared by using 6 to 8 mg of crystalline fluorescein isothiocyanate per gm of globulin and was then subjected to DEAE-cellulose chromatography and gradient elution to eliminate the most highly coupled molecules. PMID:13706641

Fellow Eye Changes in Patients with Nonischemic Central Retinal Vein Occlusion: Assessment of Perfused Foveal Microvascular Density and Identification of Nonperfused Capillaries

PubMed Central

Pinhas, Alexander; Dubow, Michael; Shah, Nishit; Cheang, Eric; Liu, Chun L.; Razeen, Moataz; Gan, Alexander; Weitz, Rishard; Sulai, Yusufu N.; Chui, Toco Y.; Dubra, Alfredo; Rosen, Richard B.

2016-01-01

Purpose Eyes fellow to nonischemic central retinal vein occlusion (CRVO) were examined for abnormalities, which might explain their increased risk for future occlusion, using adaptive optics scanning light ophthalmoscope fluorescein angiography. Methods Adaptive optics scanning light ophthalmoscope fluorescein angiography foveal microvascular densities were calculated. Nonperfused capillaries adjacent to the foveal avascular zone were identified. Spectral domain optical coherence tomography, ultrawide field fluorescein angiographies, and microperimetry were also performed. Results Ten fellow eyes of nine nonischemic CRVO and 1 nonischemic hemi-CRVO subjects and four affected eyes of three nonischemic CRVO and one nonischemic hemi-CRVO subjects were imaged. Ninety percent of fellow eyes and 100% of affected eyes demonstrated at least 1 nonperfused capillary compared with 31% of healthy eyes. Fellow eye microvascular density (35 ± 3.6 mm−1) was significantly higher than that of affected eyes (25 ± 5.2 mm−1) and significantly lower than that of healthy eyes (42 ± 4.2 mm−1). Compared with healthy controls, spectral domain optical coherence tomography thicknesses showed no significant difference, whereas microperimetry and 2/9 ultrawide field fluorescein angiography revealed abnormalities in fellow eyes. Conclusion Fellow eye changes detectable on adaptive optics scanning light ophthalmoscope fluorescein angiography reflect subclinical pathology difficult to detect using conventional imaging technologies. These changes may help elucidate the pathogenesis of nonischemic CRVO and help identify eyes at increased risk of future occlusion. PMID:25932560

Mucoadhesive ocular insert based on thiolated poly(acrylic acid): development and in vivo evaluation in humans.

PubMed

Hornof, Margit; Weyenberg, Wim; Ludwig, Annick; Bernkop-Schnürch, Andreas

2003-05-20

The aim of the study was to develop a mucoadhesive ocular insert for the controlled delivery of ophthalmic drugs and to evaluate its efficacy in vivo. The inserts tested were based either on unmodified or thiolated poly(acrylic acid). Water uptake and swelling behavior of the inserts as well as the drug release rates of the model drugs fluorescein and two diclofenac salts with different solubility properties were evaluated in vitro. Fluorescein was used as fluorescent tracer to study the drug release from the insert in humans. The mean fluorescein concentration in the cornea/tearfilm compartment as a function of time was determined after application of aqueous eye drops and inserts composed of unmodified and of thiolated poly(acrylic acid). The acceptability of the inserts by the volunteers was also evaluated. Inserts based on thiolated poly(acrylic acid) were not soluble and had good cohesive properties. A controlled release was achieved for the incorporated model drugs. The in vivo study showed that inserts based on thiolated poly(acrylic acid) provide a fluorescein concentration on the eye surface for more than 8 h, whereas the fluorescein concentration rapidly decreased after application of aqueous eye drops or inserts based on unmodified poly(acrylic acid). Moreover, these inserts were well accepted by the volunteers. The present study indicates that ocular inserts based on thiolated poly(acrylic acid) are promising new solid devices for ocular drug delivery.

Spectroscopic properties and amplified spontaneous emission of fluorescein laser dye in ionic liquids as green media

NASA Astrophysics Data System (ADS)

AL-Aqmar, Dalal M.; Abdelkader, H. I.; Abou Kana, Maram T. H.

2015-09-01

The use of ionic liquids (ILs) as milieu materials for laser dyes is a promising field and quite competitive with volatile organic solvents and solid state-dye laser systems. This paper investigates some photo-physical parameters of fluorescein dye incorporated into ionic liquids; 1-Butyl-3-methylimidazolium chloride (BMIM Cl), 1-Butyl-3-methylimidazolium tetrachloroaluminate (BMIM AlCl4) and 1-Butyl-3-methylimidazolium tetrafluoroborate (BMIM BF4) as promising host matrix in addition to ethanol as reference. These parameters are: absorption and emission cross-sections, fluorescence lifetime and quantum yield, in addition to the transition dipole moment, the attenuation length and oscillator strength were also investigated. Lasing characteristics such as amplified spontaneous emission (ASE), the gain, and the photostability of fluorescein laser dye dissolved in different host materials were assessed. The composition and properties of the matrix of ILs were found that it has great interest in optimizing the laser performance and photostability of the investigated laser dye. Under transverse pumping of fluorescein dye by blue laser diode (450 nm) of (400 mW), the initial ASE for dye dissolved in BMIM AlCl4 and ethanol were decreased to 39% and 36% respectively as time progressed 132 min. Relatively high efficiency and high fluorescence quantum yield (11.8% and 0.82% respectively) were obtained with good photostability in case of fluorescein in BMIM BF4 that was decreased to ∼56% of the initial ASE after continuously pumping with 400 mW for 132 min.

Molecular engineering of fluorescein dyes as complementary absorbers in dye co-sensitized solar cells

DOE PAGES

Pepe, Giulio; Cole, Jacqueline M.; Waddell, Paul G.; ...

2016-09-22

Fluorescein dye derivatives exhibit extended optical absorption up to 500 nm, rendering these compounds suitable as co-absorbers in dye-sensitized solar cells (DSCs). A molecular engineering approach is presented, which embraces this intrinsic optical attribute of fluoresceins, while modifying the dye chemistry to enhance their light harvesting efficiency, in order to effectively tailor them for DSC applications. This approach first realizes relationships between the molecular structure and the optoelectronic properties for a series of five a priori known (parent) fluorescein dyes: 5-carboxyfluorescein (1), a mixture of m-carboxyfluorescein where m = 5 or 6 (2), 5-carboxyfluorescein diacetate (3), 6-carboxyfluorescein diacetate (4), amore » mixture of n-carboxy-2',7'-dichlorofluorescein diacetate where n = 5 or 6 (5). The first step in this approach combines, where available, experimental and computational methods so that electronic structure calculations can also be validated for representative fluorescein dyes. Such calculations can then be used reliably to predict the structure and properties of fluorescein dyes for cases where experimental data are lacking. Structure-function relationships established from this initial step inform the selection of parent dye 1 that is taken forward to the second step in molecular engineering: in silico chemical derivation to re-functionalize 1 for DSC applications. For this purpose, computational calculations are used to extend the charge conjugation in 1 between its donor and acceptor moieties. These structural modifications result in a bathochromic shift of the lowest excitation by ~1.3-1.9 eV (100-170 nm), making the dye optically absorb in the visible region. Further calculations on dye molecules adsorbed onto the surface of a TiO 2 cluster are used to investigate the dye sensitization behavior via dye adsorption energies and anchoring modes. The results of this theoretical investigation lead to two molecularly engineered fluoresceins being proposed to act as co-sensitizers together with a rhodamine dye. This combination of three dyes ensures chemical compatibility, panchromatic absorption, and restores optical absorption dipping otherwise observed in a DSC device at ~350-400 nm owing to the I-/I- 3 electrolyte. Altogether, the results of this study demonstrate that molecular engineering can be used to identify suitable chemical modifications for organic dyes with improved light harvesting properties for photovoltaic applications.« less

Molecular engineering of fluorescein dyes as complementary absorbers in dye co-sensitized solar cells

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

Pepe, Giulio; Cole, Jacqueline M.; Waddell, Paul G.

Fluorescein dye derivatives exhibit extended optical absorption up to 500 nm, rendering these compounds suitable as co-absorbers in dye-sensitized solar cells (DSCs). A molecular engineering approach is presented, which embraces this intrinsic optical attribute of fluoresceins, while modifying the dye chemistry to enhance their light harvesting efficiency, in order to effectively tailor them for DSC applications. This approach first realizes relationships between the molecular structure and the optoelectronic properties for a series of five a priori known (parent) fluorescein dyes: 5-carboxyfluorescein (1), a mixture of m-carboxyfluorescein where m = 5 or 6 (2), 5-carboxyfluorescein diacetate (3), 6-carboxyfluorescein diacetate (4), amore » mixture of n-carboxy-2',7'-dichlorofluorescein diacetate where n = 5 or 6 (5). The first step in this approach combines, where available, experimental and computational methods so that electronic structure calculations can also be validated for representative fluorescein dyes. Such calculations can then be used reliably to predict the structure and properties of fluorescein dyes for cases where experimental data are lacking. Structure-function relationships established from this initial step inform the selection of parent dye 1 that is taken forward to the second step in molecular engineering: in silico chemical derivation to re-functionalize 1 for DSC applications. For this purpose, computational calculations are used to extend the charge conjugation in 1 between its donor and acceptor moieties. These structural modifications result in a bathochromic shift of the lowest excitation by ~1.3-1.9 eV (100-170 nm), making the dye optically absorb in the visible region. Further calculations on dye molecules adsorbed onto the surface of a TiO 2 cluster are used to investigate the dye sensitization behavior via dye adsorption energies and anchoring modes. The results of this theoretical investigation lead to two molecularly engineered fluoresceins being proposed to act as co-sensitizers together with a rhodamine dye. This combination of three dyes ensures chemical compatibility, panchromatic absorption, and restores optical absorption dipping otherwise observed in a DSC device at ~350-400 nm owing to the I-/I- 3 electrolyte. Altogether, the results of this study demonstrate that molecular engineering can be used to identify suitable chemical modifications for organic dyes with improved light harvesting properties for photovoltaic applications.« less

pH dependence and unsuitability of fluorescein dye as a tracer for pesticide mobility studies in acid soil

Treesearch

Chris Peterson

2009-01-01

The mobility of fluorescein and bromide used as tracers in packed soil columns was investigated.Five different soils were used in two application methods: soil surface application and soil incorporation, both of which simulate accepted methods of soil application of termiticides to prevent structural infestation. The...

Teaching Old Dyes New Tricks: Biological Probes Built from Fluoresceins and Rhodamines.

PubMed

Lavis, Luke D

2017-06-20

Small-molecule fluorophores, such as fluorescein and rhodamine derivatives, are critical tools in modern biochemical and biological research. The field of chemical dyes is old; colored molecules were first discovered in the 1800s, and the fluorescein and rhodamine scaffolds have been known for over a century. Nevertheless, there has been a renaissance in using these dyes to create tools for biochemistry and biology. The application of modern chemistry, biochemistry, molecular genetics, and optical physics to these old structures enables and drives the development of novel, sophisticated fluorescent dyes. This critical review focuses on an important example of chemical biology-the melding of old and new chemical knowledge-leading to useful molecules for advanced biochemical and biological experiments.

Investigation of the usefulness of fluorescein sodium fluorescence in stereotactic brain biopsy.

PubMed

Thien, Ady; Han, Julian Xinguang; Kumar, Krishan; Ng, Yew Poh; Rao, Jai Prashanth; Ng, Wai Hoe; King, Nicolas Kon Kam

2018-02-01

Intraoperative frozen section assessment, to confirm acquisition of pathological tissues, is used in stereotactic brain biopsy to minimise sampling errors. Limitations include the dependence on dedicated neuro-oncology pathologists and an increase in operative duration. We investigated the use of intraoperative fluorescein sodium, and compared it to frozen section assessment, for confirming pathological tissue samples in the stereotactic biopsy of gadolinium-contrast-enhancing brain lesions. This prospective observational study consisted of 18 consecutive patients (12 men; median age, 63 years) who underwent stereotactic biopsy of gadolinium-contrast-enhancing brain lesions with intravenous fluorescein sodium administration. Twenty-three specimens were obtained and examined for the presence of fluorescence using a microscope with fluorescence visualisation capability. Positive and negative predictive values were calculated based on the fluorescence status of the biopsy samples with its corresponding intraoperative frozen section and definitive histopathological diagnosis. Nineteen specimens (83%) were fluorescent and four (17%) were non-fluorescent. All 19 fluorescent specimens were confirmed to be lesional on intraoperative frozen section assessment and were suitable for histopathological diagnosis. Three of the non-fluorescent specimens were confirmed to be lesional on intraoperative frozen section assessment. One non-fluorescent specimen was non-diagnostic on frozen section and histological assessments. The positive predictive value was 100% and the negative predictive value was 25%. Fluorescein sodium fluorescence is as accurate as frozen section assessment in confirming sampling of pathological tissue in the stereotactic biopsy of gadolinium-contrast-enhancing brain lesions. Fluorescein sodium fluorescence-guided stereotactic biopsy is a useful addition to the neurosurgical armamentarium.

A smart membrane based on an antigen-responsive hydrogel.

PubMed

Zhang, Rongsheng; Bowyer, Adrian; Eisenthal, Robert; Hubble, John

2007-07-01

Hydrogel membranes have been fabricated that incorporate antibody/antigen moieties. The permeability of large solutes through these membranes is dependent on the presence of soluble antigen that can compete with the internal interactions between antibody and antigen leading to an increase in gel mesh size. Specifically, the membrane's structure is based on a dextran backbone grafted with a fluorescein isothiocyanate (FITC) antigen and a sheep anti-FITC IgG antibody. The backbone is covalently cross-linked by conjugated divinyl sulfone (DVS) groups. The gel structure is additionally stabilized by affinity crosslinks formed by biospecific interactions between the bound IgG and FITC. FTIR spectra of the gel are consistent with formation of covalent bonds between cysteine groups in the IgG and DVS groups in the dextran. Results obtained using isothermal titration calorimetry (ITC) confirmed the competitive interaction binding between IgG-FITC-dextran and free sodium fluorescein at pH 5.0. Scanning electron microscopy (SEM) of samples prepared using cryofixation and cryofracturing techniques showed that observed changes in permeability correlate with free fluorescein-dependent structural changes in the gel. Three-dimensional images obtained from confocal laser scanning microscopy show that these changes occur throughout the gel and indicate that SEM results are not artifacts of sample preparation. The permeability of these gels, as shown by blue-dextran (12 kDa) diffusion, increases in response to the presence of free fluorescein of the external medium, which causes competitive displacement of the affinity cross-links. Sequential addition and removal of sodium fluorescein showed that these permeability changes are reversible. (c) 2006 Wiley Periodicals, Inc.

Use of indocyanine green and sodium fluorescein for anterior segment angiography in ophthalmologically normal cats.

PubMed

Pirie, Chris G; Alario, Anthony

2015-10-01

To assess and compare results of anterior segment angiography of ophthalmologically normal cats following IV injection with indocyanine green and sodium fluorescein dyes. 10 client-owned cats. Anterior segment angiography was performed in anesthetized cats following administration of 0.25% indocyanine green (1.0 mg/kg, IV) or 10% sodium fluorescein (20 mg/kg, IV) solution. All cats received both treatments. Imaging (1 eye/cat) was performed with a full-spectrum digital single-lens reflex camera equipped with an adaptor (1 image/s for 30 seconds) immediately following IV dye injection and 1, 2, 3, 4, and 5 minutes after injection. Onset and duration of arterial, capillary, and venous phases of iris vasculature were identified and compared statistically between treatments. Degree of iridal pigmentation, leakage of dye from iris vasculature, and image quality were subjectively assessed. No differences were found in onset or duration of vascular phases between treatments. Visibility of the iris vasculature was not impaired by poor or moderate iridal pigmentation with either method. Indocyanine green provided subjectively better vascular detail and image contrast than sodium fluorescein. No vascular dye leakage was observed following indocyanine green administration. Leakage of dye from blood vessels in the stroma (in 10 cats) and presence of dye in the anterior chamber (in 5 cats) were detected after sodium fluorescein administration. Images obtained with either fluorescent dye were considered to be of diagnostic quality. Lack of leakage following indocyanine green administration suggested this treatment may have better diagnostic utility for anterior segment angiography. The photographic equipment used provided a cost-effective alternative to existing imaging systems.

Fluorescein for resection of high-grade gliomas: A safety study control in a single center and review of the literature

PubMed Central

Francaviglia, Natale; Iacopino, Domenico Gerardo; Costantino, Gabriele; Villa, Alessandro; Impallaria, Pietro; Meli, Francesco; Maugeri, Rosario

2017-01-01

Background: The importance of a complete resection of high-grade gliomas (HGGs) has been highlighted in scientific literature, in order to limit tumor recurrence and above all to improve disease-free survival rates. Several fluorescent biomarkers have been tested to improve intraoperative identification of residual tumor; 5-aminolevulinic acid (5-ALA) and fluorescein sodium (FS) are now starting to play a central role in glioma surgery. We performed a retrospective analysis on 47 patients operated for HGGs. Here we report our preliminary data. Methods: Data of 47 consecutive patients with HGG have been collected in our study (25 males, 22 females; mean age: 60.3 years, range: 27–86 years). Fluorescein (5 mg/kg of body weight) was injected intravenously right after the induction of general anesthesia. A YELLOW 560 filter was used on an OPMI Pentero 900 microscope (Carl Zeiss Meditec, Oberkochen, Germany) to complete a microsurgical tumor removal. Glioma resection and quality of life were evaluated preoperative and postoperatively. Results: Gross total resection (GTR) was achieved in 53.2% (n = 25) of patients. A subtotal resection (STR) (>95%) was achieved in 29.8% (n = 14), while a partial resection (PR) (<95%) was obtained in 17% (n = 8) of patients. Overall, in 83% (n = 39) of patients who underwent fluorescence-guided surgery the resection rate achieved was >95%. No adverse effects correlated to fluorescein have been recorded. Conclusion: Fluorescein seems to be safe and effective in the resection of HGGs, allowing a high rate of gross total removal of contrast enhanced areas. PMID:28781922

Fluorescein for resection of high-grade gliomas: A safety study control in a single center and review of the literature.

PubMed

Francaviglia, Natale; Iacopino, Domenico Gerardo; Costantino, Gabriele; Villa, Alessandro; Impallaria, Pietro; Meli, Francesco; Maugeri, Rosario

2017-01-01

The importance of a complete resection of high-grade gliomas (HGGs) has been highlighted in scientific literature, in order to limit tumor recurrence and above all to improve disease-free survival rates. Several fluorescent biomarkers have been tested to improve intraoperative identification of residual tumor; 5-aminolevulinic acid (5-ALA) and fluorescein sodium (FS) are now starting to play a central role in glioma surgery. We performed a retrospective analysis on 47 patients operated for HGGs. Here we report our preliminary data. Data of 47 consecutive patients with HGG have been collected in our study (25 males, 22 females; mean age: 60.3 years, range: 27-86 years). Fluorescein (5 mg/kg of body weight) was injected intravenously right after the induction of general anesthesia. A YELLOW 560 filter was used on an OPMI Pentero 900 microscope (Carl Zeiss Meditec, Oberkochen, Germany) to complete a microsurgical tumor removal. Glioma resection and quality of life were evaluated preoperative and postoperatively. Gross total resection (GTR) was achieved in 53.2% ( n = 25) of patients. A subtotal resection (STR) (>95%) was achieved in 29.8% ( n = 14), while a partial resection (PR) (<95%) was obtained in 17% ( n = 8) of patients. Overall, in 83% ( n = 39) of patients who underwent fluorescence-guided surgery the resection rate achieved was >95%. No adverse effects correlated to fluorescein have been recorded. Fluorescein seems to be safe and effective in the resection of HGGs, allowing a high rate of gross total removal of contrast enhanced areas.

Sensitivity and specificity of intrathecal fluorescein and white light excitation for detecting intraoperative cerebrospinal fluid leak in endoscopic skull base surgery: a prospective study.

PubMed

Raza, Shaan M; Banu, Matei A; Donaldson, Angela; Patel, Kunal S; Anand, Vijay K; Schwartz, Theodore H

2016-03-01

The intraoperative detection of CSF leaks during endonasal endoscopic skull base surgery is critical to preventing postoperative CSF leaks. Intrathecal fluorescein (ITF) has been used at varying doses to aid in the detection of intraoperative CSF leaks. However, the sensitivity and specificity of ITF at certain dosages is unknown. A prospective database of all endoscopic endonasal procedures was reviewed. All patients received 25 mg ITF diluted in 10 ml CSF and were pretreated with dexamethasone and Benadryl. Immediately after surgery, the operating surgeon prospectively noted if there was an intraoperative CSF leak and fluorescein was identified. The sensitivity, specificity, and positive and negative predictive power of ITF for detecting intraoperative CSF leak were calculated. Factors correlating with postoperative CSF leak were determined. Of 419 patients, 35.8% of patients did not show a CSF leak. Fluorescein-tinted CSF (true positive) was noted in 59.7% of patients and 0 false positives were encountered. CSF without fluorescein staining (false negative) was noted in 4.5% of patients. The sensitivity and specificity of ITF were 92.9% and 100%, respectively. The negative and positive predictive values were 88.8% and 100%, respectively. Postoperative CSF leaks only occurred in true positives at a rate of 2.8%. ITF is extremely specific and very sensitive for detecting intraoperative CSF leaks. Although false negatives can occur, these patients do not appear to be at risk for postoperative CSF leak. The use of ITF may help surgeons prevent postoperative CSF leaks by intraoperatively detecting and confirming a watertight repair.

[Channelography and mechanism of action in canaloplasty].

PubMed

Grieshaber, M C

2015-04-01

Canaloplasty lowers the intraocular pressure (IOP) by restoring the natural outflow system. The success of canaloplasty depends on the function of this system. To evaluate the natural outflow system regarding canaloplasty by two clinical tests, provocative gonioscopy and channelography and to describe the mechanism of action of canaloplasty. Provocative gonioscopy evaluates the pattern of blood reflux which is induced by ocular hypotension as the result of a reversed pressure gradient between the episcleral venous pressure and IOP following paracentesis. In channelography the transtrabecular diffusion and the filling properties of the episcleral venous system are assessed by a microcatheter and a fluorescein tracer. Blood reflux varied greatly in glaucomatous eyes and showed an inverse correlation with the preoperative IOP. The higher the IOP, the poorer the blood reflux. The filling qualities of the episcleral venous system and diffusion through the trabecular meshwork were different. Poor trabecular passage and good episcleral fluorescein outflow indicates patent distal outflow pathways, poor trabecular passage and poor episcleral fluorescein outflow indicates obstructed trabecular meshwork and closed collector channels and good trabecular passage together with poor episcleral fluorescein outflow suggests that the site of impairment is mainly in the distal outflow system. The quality of blood reflux and the characteristics of the episcleral filling and the transtrabecular diffusion by fluorescein represent the clinical state of the outflow pathway and help in the prediction of the surgical outcome in canaloplasty. The mechanism for canaloplasty is not yet completely clarified; currently under discussion are circumferential viscodilation, permanent distension of the inner wall of Schlemm's canal using a suture and a Stegmann canal expander.

Synthesis of Rhodamines from Fluoresceins Using Pd-Catalyzed C–N Cross-Coupling

PubMed Central

2011-01-01

A unified, convenient, and efficient strategy for the preparation of rhodamines and N,N′-diacylated rhodamines has been developed. Fluorescein ditriflates were found to undergo palladium-catalyzed C–N cross-coupling with amines, amides, carbamates, and other nitrogen nucleophiles to provide direct access to known and novel rhodamine derivatives, including fluorescent dyes, quenchers, and latent fluorophores. PMID:22091952

Lighting up Protons with MorphFl, a Fluorescein-Morpholine Dyad: An Experiment for the Organic Laboratory

ERIC Educational Resources Information Center

Miller, Tyson A.; Spangler, Michael; Burdette, Shawn C.

2011-01-01

A two-period organic laboratory experiment that includes fluorescence sensing is presented. The pH-sensitive sensor MorphFl is prepared using a Mannich reaction between a fluorescein derivative and the iminium ion of morpholine. During the first laboratory, students prepare MorphFl. The second session begins with characterizing the sensor using…

The effect of rigor mortis on the passage of erythrocytes and fluid through the myocardium of isolated dog hearts.

PubMed

Nevalainen, T J; Gavin, J B; Seelye, R N; Whitehouse, S; Donnell, M

1978-07-01

The effect of normal and artificially induced rigor mortis on the vascular passage of erythrocytes and fluid through isolated dog hearts was studied. Increased rigidity of 6-mm thick transmural sections through the centre of the posterior papillary muscle was used as an indication of rigor. The perfusibility of the myocardium was tested by injecting 10 ml of 1% sodium fluorescein in Hanks solution into the circumflex branch of the left coronary artery. In prerigor hearts (20 minute incubation) fluorescein perfused the myocardium evenly whether or not it was preceded by an injection of 10 ml of heparinized dog blood. Rigor mortis developed in all hearts after 90 minutes incubation or within 20 minutes of perfusing the heart with 50 ml of 5 mM iodoacetate in Hanks solution. Fluorescein injected into hearts in rigor did not enter the posterior papillary muscle and adjacent subendocardium whether or not it was preceded by heparinized blood. Thus the vascular occlusion caused by rigor in the dog heart appears to be so effective that it prevents flow into the subendocardium of small soluble ions such as fluorescein.

Fluorescein thiocarbamyl amino acids as internal standards for migration time correction in capillary sieving electrophoresis

PubMed Central

Pugsley, Haley R.; Swearingen, Kristian E.; Dovichi, Norman J.

2009-01-01

A number of algorithms have been developed to correct for migration time drift in capillary electrophoresis. Those algorithms require identification of common components in each run. However, not all components may be present or resolved in separations of complex samples, which can confound attempts for alignment. This paper reports the use of fluorescein thiocarbamyl derivatives of amino acids as internal standards for alignment of 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ)-labeled proteins in capillary sieving electrophoresis. The fluorescein thiocarbamyl derivative of aspartic acid migrates before FQ-labeled proteins and the fluorescein thiocarbamyl derivative of arginine migrates after the FQ-labeled proteins. These compounds were used as internal standards to correct for variations in migration time over a two-week period in the separation of a cellular homogenate. The experimental conditions were deliberately manipulated by varying electric field and sample preparation conditions. Three components of the homogenate were used to evaluate the alignment efficiency. Before alignment, the average relative standard deviation in migration time for these components was 13.3%. After alignment, the average relative standard deviation in migration time for these components was reduced to 0.5%. PMID:19249052

Retinal Image Quality Assessment for Spaceflight-Induced Vision Impairment Study

NASA Technical Reports Server (NTRS)

Vu, Amanda Cadao; Raghunandan, Sneha; Vyas, Ruchi; Radhakrishnan, Krishnan; Taibbi, Giovanni; Vizzeri, Gianmarco; Grant, Maria; Chalam, Kakarla; Parsons-Wingerter, Patricia

2015-01-01

Long-term exposure to space microgravity poses significant risks for visual impairment. Evidence suggests such vision changes are linked to cephalad fluid shifts, prompting a need to directly quantify microgravity-induced retinal vascular changes. The quality of retinal images used for such vascular remodeling analysis, however, is dependent on imaging methodology. For our exploratory study, we hypothesized that retinal images captured using fluorescein imaging methodologies would be of higher quality in comparison to images captured without fluorescein. A semi-automated image quality assessment was developed using Vessel Generation Analysis (VESGEN) software and MATLAB® image analysis toolboxes. An analysis of ten images found that the fluorescein imaging modality provided a 36% increase in overall image quality (two-tailed p=0.089) in comparison to nonfluorescein imaging techniques.

Synthesis of novel fluorescently labeled water-soluble fullerenes and their application to its cellar uptake and distribution properties

NASA Astrophysics Data System (ADS)

Hashimoto, Akiko; Yamanaka, Takehiro; Takamura-Enya, Takeji

2017-12-01

Fullerene is a well-known carbon nanomaterial, which can be potentially used for drug manufacture or delivery. Despite several successful examples of utilizing fullerene derivatives as drug candidate materials, their low water solubility under physiological conditions negatively affects the cell penetration efficiency after treatment. In this work, we successfully synthesized two fullerene derivatives with covalently attached fluorescein and boron dipyrromethene (BODIPY) fluorophore moieties, which exhibited cellular uptake and intracellular localization. While both fluorophores decreased their fluorescence intensity in the vicinity of fullerene, the cellar uptake of the fluorescein-modified fullerene was detected via fluorescence microscopy observations. Moreover, decreases in the fluorescence intensities of the intact fluorescein and BODIPY species were observed when both fluorophores and fullerene coexisted in aqueous media.

A hot-spot-active magnetic graphene oxide substrate for microRNA detection based on cascaded chemiluminescence resonance energy transfer

NASA Astrophysics Data System (ADS)

Bi, Sai; Chen, Min; Jia, Xiaoqiang; Dong, Ying

2015-02-01

Herein, a cascaded chemiluminescence resonance energy transfer (C-CRET) process was demonstrated from horseradish peroxidase (HRP)-mimicking DNAzyme-catalyzed luminol-H2O2 to fluorescein and further to graphene oxide (GO) when HRP-mimicking DNAzyme/fluorescein was in close proximity to the GO surface. The proposed C-CRET system was successfully implemented to construct three modes of C-CRET hot-spot-active substrates (modes I, II and III) by covalently immobilizing HRP-mimicking DNAzyme/fluorescein-labeled hairpin DNAs (hot-spot-generation probes) on magnetic GO (MGO), resulting in a signal ``off'' state due to the quenching of the luminol/H2O2/HRP-mimicking DNAzyme/fluorescein CRET system by GO. Upon the introduction of microRNA-122 (miRNA-122), the targets (mode I) or the new triggers that were generated through a strand displacement reaction (SDR) initiated by miRNA-122 (modes II and III) hybridized with the loop domains of hairpin probes on MGO to form double-stranded (modes I and II) or triplex-stem structures (mode III), causing an ``open'' configuration of the hairpin probe and a CRET signal ``on'' state, thus achieving sensitive and selective detection of miRNA-122. More importantly, the substrate exhibited excellent controllability, reversibility and reproducibility through SDR and magnetic separation (modes II and III), especially sequence-independence for hairpin probes in mode III, holding great potential for the development of a versatile platform for optical biosensing.Herein, a cascaded chemiluminescence resonance energy transfer (C-CRET) process was demonstrated from horseradish peroxidase (HRP)-mimicking DNAzyme-catalyzed luminol-H2O2 to fluorescein and further to graphene oxide (GO) when HRP-mimicking DNAzyme/fluorescein was in close proximity to the GO surface. The proposed C-CRET system was successfully implemented to construct three modes of C-CRET hot-spot-active substrates (modes I, II and III) by covalently immobilizing HRP-mimicking DNAzyme/fluorescein-labeled hairpin DNAs (hot-spot-generation probes) on magnetic GO (MGO), resulting in a signal ``off'' state due to the quenching of the luminol/H2O2/HRP-mimicking DNAzyme/fluorescein CRET system by GO. Upon the introduction of microRNA-122 (miRNA-122), the targets (mode I) or the new triggers that were generated through a strand displacement reaction (SDR) initiated by miRNA-122 (modes II and III) hybridized with the loop domains of hairpin probes on MGO to form double-stranded (modes I and II) or triplex-stem structures (mode III), causing an ``open'' configuration of the hairpin probe and a CRET signal ``on'' state, thus achieving sensitive and selective detection of miRNA-122. More importantly, the substrate exhibited excellent controllability, reversibility and reproducibility through SDR and magnetic separation (modes II and III), especially sequence-independence for hairpin probes in mode III, holding great potential for the development of a versatile platform for optical biosensing. Electronic supplementary information (ESI) available: Sequences of RNA and DNA used in this study, relationship of the proposed three modes, CRET mechanism of the luminol/H2O2/HRP-mimicking DNAzyme/fluorescein system, calculation of the surface coverage of hairpin probe I-1 on MGO, control experiment, comparison between different modes for microRNA detection, and advantages of the proposed strategy. See DOI: 10.1039/c4nr06603k

Distribution of Particles, Small Molecules and Polymeric Formulation Excipients in the Suprachoroidal Space after Microneedle Injection

PubMed Central

Chiang, Bryce; Venugopal, Nitin; Edelhauser, Henry F.; Prausnitz, Mark R.

2016-01-01

The purpose of this work was to determine the effect of injection volume, formulation composition, and time on circumferential spread of particles, small molecules and polymeric formulation excipients in the suprachoroidal space (SCS) after microneedle injection into New Zealand White rabbit eyes ex vivo and in vivo. Microneedle injections of 25–150 μL Hank’s Balanced Salt Solution (HBSS) containing 0.2 μm red-fluorescent particles and a model small molecule (fluorescein) were performed in rabbit eyes ex vivo, and visualized via flat mount. Particles with diameters of 0.02 – 2 μm were co-injected into SCS in vivo with fluorescein or a polymeric formulation excipient: fluorescein isothiocyanate (FITC)-labeled Discovisc or FITC-labeled carboxymethyl cellulose (CMC). Fluorescent fundus images were acquired over time to determine area of particle, fluorescein and polymeric formulation excipient spread, as well as their co-localization. We found that fluorescein covered a significantly larger area than co-injected particles when suspended in HBSS, and that this difference was present from 3 min post-injection onwards. We further showed that there was no difference in initial area covered by FITC-Discovisc and particles; the transport time (i.e., the time until the FITC-Discovisc and particle area began dissociating) was 2 d. There was also no difference in initial area covered by FITC-CMC and particles; the transport time in FITC-CMC was 4 d. We also found that particle size (20 nm – 2 μm) had no effect on spreading area when delivered in HBSS or Discovisc. We conclude that (i) the area of particle spread in SCS during injection generally increased with increasing injection volume, was unaffected by particle size and was significantly less than the area of fluorescein spread, (ii) particles suspended in low-viscosity HBSS formulation were entrapped in the SCS after injection, whereas fluorescein was not and (iii) particles co-injected with viscous polymeric formulation excipients co-localized near the site of injection in the SCS, continued to co-localize while spreading over larger areas for 2 – 4 days, and then no longer co-localized as the polymeric formulation excipients were cleared within 1 – 3 weeks and the particles remained largely in place. These data suggest that particles encounter greater barriers to flow in SCS compared to molecules and that co-localization of particles and polymeric formulation excipients allow spreading over larger areas of the SCS until the particles and excipients dissociate. PMID:27742547

Distribution of particles, small molecules and polymeric formulation excipients in the suprachoroidal space after microneedle injection.

PubMed

Chiang, Bryce; Venugopal, Nitin; Edelhauser, Henry F; Prausnitz, Mark R

2016-12-01

The purpose of this work was to determine the effect of injection volume, formulation composition, and time on circumferential spread of particles, small molecules, and polymeric formulation excipients in the suprachoroidal space (SCS) after microneedle injection into New Zealand White rabbit eyes ex vivo and in vivo. Microneedle injections of 25-150 μL Hank's Balanced Salt Solution (HBSS) containing 0.2 μm red-fluorescent particles and a model small molecule (fluorescein) were performed in rabbit eyes ex vivo, and visualized via flat mount. Particles with diameters of 0.02-2 μm were co-injected into SCS in vivo with fluorescein or a polymeric formulation excipient: fluorescein isothiocyanate (FITC)-labeled Discovisc or FITC-labeled carboxymethyl cellulose (CMC). Fluorescent fundus images were acquired over time to determine area of particle, fluorescein, and polymeric formulation excipient spread, as well as their co-localization. We found that fluorescein covered a significantly larger area than co-injected particles when suspended in HBSS, and that this difference was present from 3 min post-injection onwards. We further showed that there was no difference in initial area covered by FITC-Discovisc and particles; the transport time (i.e., the time until the FITC-Discovisc and particle area began dissociating) was 2 d. There was also no difference in initial area covered by FITC-CMC and particles; the transport time in FITC-CMC was 4 d. We also found that particle size (20 nm-2 μm) had no effect on spreading area when delivered in HBSS or Discovisc. We conclude that (i) the area of particle spread in SCS during injection generally increased with increasing injection volume, was unaffected by particle size, and was significantly less than the area of fluorescein spread, (ii) particles suspended in low-viscosity HBSS formulation were entrapped in the SCS after injection, whereas fluorescein was not and (iii) particles co-injected with viscous polymeric formulation excipients co-localized near the site of injection in the SCS, continued to co-localize while spreading over larger areas for 2-4 days, and then no longer co-localized as the polymeric formulation excipients were cleared within 1-3 weeks and the particles remained largely in place. These data suggest that particles encounter greater barriers to flow in SCS compared to molecules and that co-localization of particles and polymeric formulation excipients allows spreading over larger areas of the SCS until the particles and excipients dissociate. Copyright © 2016 Elsevier Ltd. All rights reserved.

Targeting PCNA Phosphorylation in Breast Cancer

DTIC Science & Technology

2012-04-01

inhibitor and dyes. These basic peptides and polycationic peptoids were labeled with fluorescein and used to assess the effect of structure on cellular... substitution of a peptoid conjugate that was synthesized during the project period. Additional types of analogs were prepared as well each providing...Broceta, A., Diezmann, F., Ou-Yang, C. Y., Fara, M. A. & Bradley, M. Synthesis, penetrability and intracellular targeting of fluorescein -tagged

The Spectrum of Optic Disc Ischemia in Patients Younger than 50 Years (An Amercian Ophthalmological Society Thesis)

PubMed Central

Arnold, Anthony C.; Costa, Roberta M. S.; Dumitrascu, Oana M.

2013-01-01

Purpose: To identify the spectrum of clinical and fluorescein angiographic features of optic disc ischemia in patients younger than 50 years. Methods: This retrospective comparative case series from a university consultative neuro-ophthalmology practice consisted of two phases. The first compared 108 cases of nonarteritic anterior ischemic optic neuropathy in patients younger than 50 years (NAIONy) to a cohort of 108 cases in patients 50 years or older (NAIONo). Predisposing risk factors, fluorescein angiographic features, and clinical course were compared. In the second phase, 12 cases of diabetic papillopathy under age 50 were assessed by fluorescein angiographic criteria for evidence of optic disc ischemia and compared to patients with NAIONy. Results: NAIONy comprised 108 (12.7%) of 848 NAION cases reviewed. Chronic renal failure with dialysis and migraine were more common in NAIONy. Fellow eye involvement rate was significantly higher for NAIONy patients (46/108, 42.6%) than for NAIONo patients (32/108, 29.6%). Fluorescein angiographic features of ischemia were documented in 44 (81.5%) of 54 eyes studied. In one case, these features were documented in pre-NAION edema. Diabetic papillopathy demonstrated delayed filling consistent with ischemia in 7 of 10 (70.0%), without significant visual field loss. Conclusions: Ischemic optic neuropathy in patients younger than 50 years is not rare. Fellow eye involvement is more frequent in younger patients. Fluorescein angiography confirmation of impaired perfusion in multiple syndromes of optic neuropathy corroborates a spectrum of optic disc ischemia ranging from perfusion delay without visual loss to severely impaired perfusion and visual loss and incorporates optic neuropathies previously considered nonischemic. PMID:24167327

Dodecyl and octyl esters of fluorescein as protonophores and uncouplers of oxidative phosphorylation in mitochondria at submicromolar concentrations.

PubMed

Shchepinova, Maria M; Denisov, Stepan S; Kotova, Elena A; Khailova, Ljudmila S; Knorre, Dmitry A; Korshunova, Galina A; Tashlitsky, Vadim N; Severin, Fedor F; Antonenko, Yuri N

2014-01-01

In our search for fluorescent uncouplers of oxidative phosphorylation, three esters of fluorescein, n-butyl-, n-octyl-, and n-dodecyl-oxycarbonyl-fluorescein (C4-FL, C8-FL, C12-FL) were synthesized and characterized. With increasing liposomal lipid content, the long-chain alkyl derivatives of fluorescein (C8-FL, C12-FL and commercially available C18-FL), but not C4-FL and unsubstituted fluorescein, exhibited an increase in fluorescence polarization reflecting the dye binding to liposomes. C12-FL induced proton permeability in lipid membranes, while C4-FL was inactive. In contrast to C4-FL and C18-FL, C12-FL and C8-FL increased the respiration rate and decreased the membrane potential of isolated rat liver mitochondria with half-maximal effective concentrations of 700nM and 300nM, respectively. The effect of Cn-FL on the respiration correlated with that on proton permeability of the inner mitochondrial membrane, as measured by induction of mitochondria swelling in the potassium acetate medium. Binding of C8-FL to mitochondria depended on their energization, which was apparently associated with pH gradient generation across the inner mitochondrial membrane in the presence of a respiratory substrate. In wild-type yeast cells, C12-FL localized predominantly in plasma membrane, whereas in AD1-8 mutants lacking MDR pumps, it stained cytoplasmic organelles with some preference for mitochondria. Fluorescent uncouplers can be useful as a tool for determining their localization in a cell or distribution between different tissues in a living animal by fluorescent microscopy. © 2013.

Fluorophilia: Fluorophore-containing compounds adhere non-specifically to injured neurons

PubMed Central

Hawkins, Bridget E.; Frederickson, Christopher J.; DeWitt, Douglas S.; Prough, Donald S.

2012-01-01

Ionic (free) zinc (Zn2+) is implicated in apoptotic neuronal degeneration and death. In our attempt to examine the effects of Zn2+ in neurodegeneration following brain injury, we serendipitously discovered that injured neurons bind fluorescein moieties, either alone or as part of an indicator dye, in histologic sections. This phenomenon, that we have termed “fluorophilia”, is analogous to the ability of degenerating neuronal somata and axons to bind silver ions (argyrophilia — the basis of silver degeneration stains). To provide evidence that fluorophilia occurs in sections of brain tissue, we used a wide variety of indicators such as Fluoro-Jade (FJ), a slightly modified fluorescein sold as a marker for degenerating neurons; Newport Green, a fluorescein-containing Zn2+ probe; Rhod-5N, a rhodamine-containing Ca2+ probe; and plain fluorescein. All yielded remarkably similar staining of degenerating neurons in the traumatic brain-injured tissue with the absence of staining in our sham-injured brains. Staining of presumptive injured neurons by these agents was not modified when Zn2+ in the brain section was removed by prior chelation with EDTA or TPEN, whereas staining by a non-fluorescein containing Zn2+ probe, N-(6-methoxy-8-quinolyl)-p-toluenesulfonamide (TSQ), was suppressed by prior chelation. Thus, certain fluorophore-containing compounds nonspecifically stain degenerating neuronal tissue in histologic sections and may not reflect the presence of Zn2+. This may be of concern to researchers using indicator dyes to detect metals in brain tissue sections. Further experiments may be advised to clarify whether Zn2+-binding dyes bind more specifically in intact neurons in culture or organotypic slices. PMID:22137653

The fluorescence resonance energy transfer (FRET) gate: a time-resolved study.

PubMed

Xu, Qing-Hua; Wang, Shu; Korystov, Dmitry; Mikhailovsky, Alexander; Bazan, Guillermo C; Moses, Daniel; Heeger, Alan J

2005-01-18

The two-step energy-transfer process in a self-assembled complex comprising a cationic conjugated polymer (CCP) and a dsDNA is investigated by using pump-dump-emission spectroscopy and time-correlated single-photon counting; energy is transferred from the CCP to an ethidium bromide (EB) molecule intercalated into the dsDNA through a fluorescein molecule linked to one terminus of the DNA. Time-dependent anisotropy measurements indicate that the inefficient direct energy transfer from the CCP to the intercalated EB results from the near orthogonality of their transition moments. These measurements also show that the transition moment of the fluorescein spans a range of angular distributions and lies between that of the CCP and EB. Consequently, the fluorescein acts as a fluorescence resonance energy-transfer gate to relay the excitation energy from the CCP to the EB.

The fluorescence resonance energy transfer (FRET) gate: A time-resolved study

PubMed Central

Xu, Qing-Hua; Wang, Shu; Korystov, Dmitry; Mikhailovsky, Alexander; Bazan, Guillermo C.; Moses, Daniel; Heeger, Alan J.

2005-01-01

The two-step energy-transfer process in a self-assembled complex comprising a cationic conjugated polymer (CCP) and a dsDNA is investigated by using pump-dump-emission spectroscopy and time-correlated single-photon counting; energy is transferred from the CCP to an ethidium bromide (EB) molecule intercalated into the dsDNA through a fluorescein molecule linked to one terminus of the DNA. Time-dependent anisotropy measurements indicate that the inefficient direct energy transfer from the CCP to the intercalated EB results from the near orthogonality of their transition moments. These measurements also show that the transition moment of the fluorescein spans a range of angular distributions and lies between that of the CCP and EB. Consequently, the fluorescein acts as a fluorescence resonance energy-transfer gate to relay the excitation energy from the CCP to the EB. PMID:15642946

[Nursing care in fluorescein angiography].

PubMed

Santos-Blanco, Feliciano

2008-01-01

Fluoresceinic angiography of the ocular fundus is a diagnostic technique to study retinal and choroidal circulation. This technique consists of parenteral administration of 500 mg of sodium fluorescein 10% and photographing the fluorescence in the eye vessels. Although this substance is fairly safe, it may also produce mild, moderate or severe local and/or general adverse reactions. The nursing process is routinely used in hospital units but not always in outpatient clinics, even through the use of invasive procedures with intravenous medication administration is common. Therefore, nurses, as those reponsible for intravenous administration, should use the nursing process to guarantee the quality of care required by the patient. To do this, we describe an individualized care plan based on evaluation by Marjorie Gordon's functional health patterns, NANDA's nursing diagnoses Taxonomy II, Nursing Outcomes Classification (NOC), Nursing Interventions Classifications (NIC) and potential complications of the procedure.

Metal Ion Sensor with Catalytic DNA in a Nanofluidic Intelligent Processor

DTIC Science & Technology

2011-12-01

attributed to decreased diffusion and less active DNAzyme complex because of pore constraints. Uncleavable Alexa546 intensity is shown in gray ...is shown in gray , cleavable fluorescein in green, and the ratio of Fl/Alexa in red. Error bars represent one standard deviation of four independent...higher concentrations inhibiting cleaved fragment release. Uncleavable Alexa 546 intensity is shown in gray , cleavable fluorescein in green, and the

A nanodiamond-fluorescein conjugate for cell studies

NASA Astrophysics Data System (ADS)

Pedroso-Santana, Seidy; Fleitas-Salazar, Noralvis; Sarabia-Sainz, Andrei; Silva-Campa, Erika; Burgara-Estrella, Alexel; Angulo-Molina, Aracely; Melendrez, Rodrigo; Pedroza-Montero, Martin; Riera, Raul

2018-03-01

The use of nanodiamonds in studies with living systems generally involves the modification of their surfaces with functional groups. Fluorescent molecules can be attached to these groups, so that one can know the exact position of the particles in each moment of the interaction with the cells. Here we modify the surface of detonation nanodiamonds and nitrogen-vacancy center nanodiamonds using carboxylation and hydroxylation procedures. Subsequent reactions with silicates and cysteine, before addition of fluorescein allow to obtain fluorescent nano-conjugates. We used confocal microscopy to observe the position of nanodiamonds interacting with HeLa cells. At 3 h post-incubation the green fluorescence is localized in extracellular rounded like-vesicles assemblies while at 24 h the conjugates can be observed inside the cells. The measurement of the fluorescence emitted by both conjugates allowed to find an enhanced emission of fluorescein isothiocyanate (FITC) when the nitrogen-vacancy center is present. We propose the existence of a fluorescence enhancement by electron transference process. The procedure described in this work allows the functionalization of nanodiamonds with FITC and other molecules using functional surface groups and small size mediators. Also, as was proved in our work, the nanodiamond-fluorescein conjugates can be used to track nanoparticles position within the cell. Localization studies are particularly important for drug delivery applications of nanodiamonds.

Method for accurate quantitation of background tissue optical properties in the presence of emission from a strong fluorescence marker

NASA Astrophysics Data System (ADS)

Bravo, Jaime; Davis, Scott C.; Roberts, David W.; Paulsen, Keith D.; Kanick, Stephen C.

2015-03-01

Quantification of targeted fluorescence markers during neurosurgery has the potential to improve and standardize surgical distinction between normal and cancerous tissues. However, quantitative analysis of marker fluorescence is complicated by tissue background absorption and scattering properties. Correction algorithms that transform raw fluorescence intensity into quantitative units, independent of absorption and scattering, require a paired measurement of localized white light reflectance to provide estimates of the optical properties. This study focuses on the unique problem of developing a spectral analysis algorithm to extract tissue absorption and scattering properties from white light spectra that contain contributions from both elastically scattered photons and fluorescence emission from a strong fluorophore (i.e. fluorescein). A fiber-optic reflectance device was used to perform measurements in a small set of optical phantoms, constructed with Intralipid (1% lipid), whole blood (1% volume fraction) and fluorescein (0.16-10 μg/mL). Results show that the novel spectral analysis algorithm yields accurate estimates of tissue parameters independent of fluorescein concentration, with relative errors of blood volume fraction, blood oxygenation fraction (BOF), and the reduced scattering coefficient (at 521 nm) of <7%, <1%, and <22%, respectively. These data represent a first step towards quantification of fluorescein in tissue in vivo.

Fluorescein-labeled stable neurotensin derivatives.

PubMed

Maes, Veronique; Hultsch, Christina; Kohl, Suzann; Bergmann, Ralf; Hanke, Thomas; Tourwé, Dirk

2006-08-01

Neurotensin(8-13) analogs containing a glycine or 5-aminovaleroyl spacer were labeled with fluorescein through formation of an N-terminal thiourea function. The receptor binding was measured in HT-29 cell cultures and showed a substantial decrease in affinity, especially for the metabolically stabilized [MeArg(9), Tle(11)] analog. Using fluorescence microscopy, the internalization of the fluorescent neurotensin analogs into HT-29 cells was observed. Copyright (c) 2006 European Peptide Society and John Wiley & Sons, Ltd.

[Simultaneous staining with fluorescein diacetate-propidium iodide to determine isolated cochlear outer hair cell viability of guinea pig].

PubMed

Yu, Q; Shi, H; Wang, J

1995-01-01

A simultaneous double-staining procedure using fluorescein diacetate (FDA) and propidium iodide (PI) is discribed for use in the determination of isolated cochlear outer hair cell viability. With exciter light, viable cells fluoresce bright green, while nonviable cells are bright red. In cell culture and cytotoxicity studies, double-staining with FDA-PI is a accurate method to discriminate between live and nonviable cells.

Cryogenic Collection of Complete Subsurface Samples for Molecular Biological Analysis

DTIC Science & Technology

2012-05-01

Nitrate was analyzed by ion chromatography ( Dionex IC25) and had a detection limit of 0.01 mg/L. Fluorescein was measured using a flow-through...dissolved oxygen (DO) with a flow through electrode, Nitrate by ion chromatography , and fluorescein with a flow through fluorometer. 1.9 LARGE...measured by headspace gas chromatography (HP 7694 Headspace Sampler attached to an HP 5890 GC with an FID detector). The GC method had a detection

Development of poly-l-lysine-coated calcium-alginate microspheres encapsulating fluorescein-labeled dextrans

NASA Astrophysics Data System (ADS)

Charron, Luc; Harmer, Andrea; Lilge, Lothar

2005-09-01

A technique to produce fluorescent cell phantom standards based on calcium alginate microspheres with encapsulated fluorescein-labeled dextrans is presented. An electrostatic ionotropic gelation method is used to create the microspheres which are then exposed to an encapsulation method using poly-l-lysine to trap the dextrans inside. Both procedures were examined in detail to find the optimal parameters producing cell phantoms meeting our requirements. Size distributions favoring 10-20 microns microspheres were obtained by varying the high voltage and needle size parameters. Typical size distributions of the samples were centered at 150 μm diameter. Neither the molecular weight nor the charge of the dextrans had a significant effect on their retention in the microspheres, though anionic dextrans were chosen to help in future capillary electrophoresis work. Increasing the exposure time of the microspheres to the poly-l-lysine solution decreased the leakage rates of fluorescein-labeled dextrans.

Isolation and Characterization of Precise Dye/Dendrimer Ratios

PubMed Central

Dougherty, Casey A.; Furgal, Joseph C.; van Dongen, Mallory A.; Goodson, Theodore; Banaszak Holl, Mark M.; Manono, Janet; DiMaggio, Stassi

2014-01-01

Fluorescent dyes are commonly conjugated to nanomaterials for imaging applications using stochastic synthesis conditions that result in a Poisson distribution of dye/particle ratios and therefore a broad range of photophysical and biodistribution properties. We report the isolation and characterization of generation 5 poly(amidoamine) (G5 PAMAM) dendrimer samples containing 1, 2, 3, and 4 fluorescein (FC) or 6-carboxytetramethylrhodamine succinimidyl ester (TAMRA) dyes per polymer particle. For the fluorescein case, this was achieved by stochastically functionalizing dendrimer with a cyclooctyne `click' ligand, separation into sample containing precisely defined `click' ligand/particle ratios using reverse-phase high performance liquid chromatography (rp-HPLC), followed by reaction with excess azide-functionalized fluorescein dye. For the TAMRA samples, stochastically functionalized dendrimer was directly separated into precise dye/particle ratios using rp-HPLC. These materials were characterized using 1H and 19F NMR, rp-HPLC, UV-Vis and fluorescence spectroscopy, lifetime measurements, and MALDI. PMID:24604830

Bilateral ophthalmic artery occlusion in rhino-orbito-cerebral mucormycosis.

PubMed

Song, Yoo Mi; Shin, Sun Young

2008-03-01

To report a case of bilateral ophthalmic artery occlusion in rhino-orbito-cerebral mucormycosis. Reviewed clinical charts, photographs, and fluorescein angiography An 89-year-old man with poorly controlled diabetes developed sudden bilateral ptosis, complete ophthalmoplegia of the right eye, and superior rectus palsy of the left eye. Brain and orbit magnetic resonance imaging showed midbrain infarction and mild diffuse sinusitis. On the 2nd day of hospitalization, sudden visual loss and light reflex loss developed. There were retinal whitening, absence of retinal arterial filling, and a total lack of choroidal perfusion on fluorescein angiography of the right eye. The left eye showed a cherry red spot in the retina and the absence of retinal arterial filling and partial choroidal perfusion on fluorescein angiography. On rhinologic examination, mucormyosis was noticed. Despite treatment, visual acuity and light reflex did not recover and he died 4 days after admission. Bilateral ophthalmic artery occlusion can occur in rhino-orbital-cerebral mucormycosis.

Functionality screen of streptavidin mutants by non-denaturing SDS-PAGE using biotin-4-fluorescein.

PubMed

Humbert, Nicolas; Ward, Thomas R

2008-01-01

Site-directed mutagenesis or directed evolution of proteins often leads to the production of inactive mutants. For streptavidin and related proteins, mutations may lead to the loss of their biotin-binding properties. With high-throughput screening methodologies in mind, it is imperative to detect, prior to the high-density protein production, the bacteria that produce non-functional streptavidin isoforms. Based on the incorporation of biotin-4-fluorescein in streptavidin mutants present in Escherichia coli bacterial extracts, we detail a functional screen that allows the identification of biotin-binding streptavidin variants. Bacteria are cultivated in a small volume, followed by a rapid treatment of the cells; biotin-4-fluorescein is added to the bacterial extract and loaded on an Sodium Dodecyl Sulfate Poly-Acrylamide Gel Electrophoresis (SDS-PAGE) under non-denaturing conditions. Revealing is performed using a UV transilluminator. This screen is thus easy to implement, cheap and requires only readily available equipment.

Acute toxicity of heavy metals for benthic epiphytic foraminifera Pararotalia spinigera (Le Calvez) and influence of seaweed-derived DOC

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

Bresler, V.; Yanko, V.

1995-10-01

The acute toxicity of cadmium, copper, and mercury to the benthic epiphytic foraminiferan Pararotalia spinigera (Le Calvez) was investigated using seven different vital cytophysiological and cytochemical methods. The ability to enzymatically hydrolyze the fluorogenic substrates fluorescein diacetate or fluorescein dibutyrate was the most sensitive method of LC50 value determination. The LC50 (24-h) values for cadmium, copper, and mercury determined by this assay with fluorescein diacetate was 0.56, 1.4, and 0.07 {micro}M, respectively. The content of seaweed-derived dissolved organic carbon (DOC), measured by absorbance at 436 nm, produced a dramatic increase of LC50 values for the heavy metals in a dose-dependentmore » manner. ``Intact`` epiphytic foraminifera attached to seaweeds are less sensitive to acute toxicity of cadmium, copper, and mercury than are ``detached`` foraminifera.« less

Fundus autofluorescence patterns in primary intraocular lymphoma.

PubMed

Casady, Megan; Faia, Lisa; Nazemzadeh, Maryam; Nussenblatt, Robert; Chan, Chi-Chao; Sen, H Nida

2014-02-01

To evaluate fundus autofluorescence (FAF) patterns in patients with primary intraocular (vitreoretinal) lymphoma. Records of all patients with primary intraocular lymphoma who underwent FAF imaging at the National Eye Institute were reviewed. Fundus autofluorescence patterns were evaluated with respect to clinical disease status and the findings on fluorescein angiography and spectral-domain optical coherence tomography. There were 18 eyes (10 patients) with primary intraocular lymphoma that underwent FAF imaging. Abnormal autofluorescence in the form of granular hyperautofluorescence and hypoautofluorescence was seen in 11 eyes (61%), and blockage by mass lesion was seen in 2 eyes (11%). All eyes with granular pattern on FAF had active primary intraocular lymphoma at the time of imaging, but there were 5 eyes with unremarkable FAF, which were found to have active lymphoma. The most common pattern on fluorescein angiography was hypofluorescent round spots with a "leopard spot" appearance (43%). These hypofluorescent spots on fluorescein angiography correlated with hyperautofluorescent spots on FAF in 5 eyes (36%) (inversion of FAF). Nodular hyperreflective spots at the level of retinal pigment epithelium on optical coherence tomography were noted in 43% of eyes. The hyperautofluorescent spots on FAF correlated with nodular hyperreflective spots on optical coherence tomography in 6 eyes (43%). Granularity on FAF was associated with active lymphoma in majority of the cases. An inversion of FAF (hyperautofluorescent spots on FAF corresponding to hypofluorescent spots on fluorescein angiography) was observed in less than half of the eyes.

Microscope-Integrated Optical Coherence Tomography Angiography in the Operating Room in Young Children With Retinal Vascular Disease.

PubMed

Chen, Xi; Viehland, Christian; Carrasco-Zevallos, Oscar M; Keller, Brenton; Vajzovic, Lejla; Izatt, Joseph A; Toth, Cynthia A

2017-05-01

Intraoperative optical coherence tomography (OCT) has gained traction as an important adjunct for clinical decision making during vitreoretinal surgery, and OCT angiography (OCTA) has provided novel insights in clinical evaluation of retinal diseases. To date, these two technologies have not been applied in combination to evaluate retinal vascular disease in the operating suite. To conduct microscope-integrated, swept-source OCTA (MIOCTA) in children with retinal vascular disease. In this case report analysis, OCT imaging in pediatric patients, MIOCTA images were obtained during examination under anesthesia from a young boy with a history of idiopathic vitreous hemorrhage and a female infant with familial exudative vitreoretinopathy. Side-by-side comparison of research MIOCT angiograms and clinically indicated fluorescein angiograms. In 2 young children with retinal vascular disease, the MIOCTA images showed more detailed vascular patterns than were visible on the fluorescein angiograms although within a more posterior field of view. The MIOCTA system allowed visualization of small pathological retinal vessels in the retinal periphery that were obscured in the fluorescein angiograms by fluorescein staining from underlying, preexisting laser scars. This is the first report to date of the use of MIOCTA in the operating room for young children with retinal vascular disease. Further optimization of this system may allow noninvasive detailed evaluation of retinal vasculature during surgical procedures and in patients who could not cooperate with in-office examinations.

Angiographic lesion size associated with LOC387715 A69S genotype in subfoveal polypoidal choroidal vasculopathy.

PubMed

Sakurada, Yoichi; Kubota, Takeo; Imasawa, Mitsuhiro; Tsumura, Toyoaki; Mabuchi, Fumihiko; Tanabe, Naohiko; Iijima, Hiroyuki

2009-01-01

To investigate whether the LOC387715/ARMS2 variants are associated with an angiographic phenotype, including lesion size and composition, in subfoveal polypoidal choroidal vasculopathy. Ninety-two subjects with symptomatic subfoveal polypoidal choroidal vasculopathy, whose visual acuity was from 0.1 to 0.5 on the Landolt chart, were genotyped for the LOC387715 polymorphism (rs10490924) using denaturing high-performance chromatography. The angiographic phenotype, including lesion composition and size, was evaluated by evaluators who were masked for the genotype. Lesion size was assessed by the greatest linear dimension based on fluorescein or indocyanine green angiography. Although there was no statistically significant difference in lesion size on indocyanine green angiography (P = 0.36, Kruskal-Wallis test) and in lesion composition (P = 0.59, chi-square test) among the 3 genotypes, there was a statistically significant difference in lesion size on fluorescein angiography (P = 0.0022, Kruskal-Wallis test). The LOC387715 A69S genotype is not associated with lesion composition or size on indocyanine green angiography but with lesion size on fluorescein angiography in patients with subfoveal polypoidal choroidal vasculopathy. Because fluorescein angiography findings represent secondary exudative changes, including subretinal hemorrhages and retinal pigment epithelial detachment, the results in the present study likely indicate that the T allele at the LOC387715 gene is associated with the exudative activity of polypoidal lesions.

Finite element modeling of drug distribution in the vitreous humor of the rabbit eye.

PubMed

Friedrich, S; Cheng, Y L; Saville, B

1997-01-01

Direct intravitreal injection of drug is a common method for treating diseases of the retina or vitreous. The stagnant nature of the vitreous humor and surrounding tissue barriers creates concentration gradients within the vitreous that must be accounted for when developing drug therapy. The objective of this research was to study drug distribution in the vitreous humor of the rabbit eye after an intravitreal injection, using a finite element model. Fluorescein and fluorescein glucuronide were selected as model compounds due to available experimental data. All required model parameters were known except for the permeability of these compounds through the retina, which was determined by fitting model predictions to experimental data. The location of the intravitreal injection in the experimental studies was not precisely known; therefore, several injection locations were considered, and best-fit retinal permeability was determined for each case. Retinal permeability of fluorescein and fluorescein glucuronide estimated by the model ranged from 1.94 x 10(-5) to 3.5 x 10(-5) cm s(-1) and from 0 to 7.62 x 10(-7) cm s(-1), respectively, depending on the assumed site of the injection. These permeability values were compared with values previously calculated from other models, and the limitations of the models are discussed. Intravitreal injection position was found to be an important variable that must be controlled in both experimental and clinical settings.

A fluorometric lateral flow assay for visual detection of nucleic acids using a digital camera readout.

PubMed

Magiati, Maria; Sevastou, Areti; Kalogianni, Despina P

2018-06-04

A fluorometric lateral flow assay has been developed for the detection of nucleic acids. The fluorophores phycoerythrin (PE) and fluorescein isothiocyanate (FITC) were used as labels, while a common digital camera and a colored vinyl-sheet, acting as a cut-off optical filter, are used for fluorescence imaging. After DNA amplification by polymerase chain reaction (PCR), the biotinylated PCR product is hybridized to its complementary probe that carries a poly(dA) tail at 3΄ edge and then applied to the lateral flow strip. The hybrids are captured to the test zone of the strip by immobilized poly(dT) sequences and detected by streptavidin-fluorescein and streptavidin-phycoerythrin conjugates, through streptavidin-biotin interaction. The assay is widely applicable, simple, cost-effective, and offers a large multiplexing potential. Its performance is comparable to assays based on the use of streptavidin-gold nanoparticles conjugates. As low as 7.8 fmol of a ssDNA and 12.5 fmol of an amplified dsDNA target were detectable. Graphical abstract Schematic presentation of a fluorometric lateral flow assay based on fluorescein and phycoerythrin fluorescent labels for the detection of single-stranded (ssDNA) and double-stranded DNA (dsDNA) sequences and using a digital camera readout. SA: streptavidin, BSA: Bovine Serum Albumin, B: biotin, FITC: fluorescein isothiocyanate, PE: phycoerythrin, TZ: test zone, CZ: control zone.

NMR structure and dynamics of the engineered fluorescein-binding lipocalin FluA reveal rigidification of beta-barrel and variable loops upon enthalpy-driven ligand binding.

PubMed

Mills, Jeffrey L; Liu, Gaohua; Skerra, Arne; Szyperski, Thomas

2009-08-11

The NMR structure of the 21 kDa lipocalin FluA, which was previously obtained by combinatorial design, elucidates a reshaped binding site specific for the dye fluorescein resulting from 21 side chain replacements with respect to the parental lipocalin, the naturally occurring bilin-binding protein (BBP). As expected, FluA exhibits the lipocalin fold of BBP, comprising eight antiparallel beta-strands forming a beta-barrel with an alpha-helix attached to its side. Comparison of the NMR structure of free FluA with the X-ray structures of BBP.biliverdin IX(gamma) and FluA.fluorescein complexes revealed significant conformational changes in the binding pocket, which is formed by four loops at the open end of the beta-barrel as well as adjoining beta-strand segments. An "induced fit" became apparent for the side chain conformations of Arg 88 and Phe 99, which contact the bound fluorescein in the complex and undergo concerted rearrangement upon ligand binding. Moreover, slower internal motional modes of the polypeptide backbone were identified by measuring transverse (15)N backbone spin relaxation times in the rotating frame for free FluA and also for the FluA.fluorescein complex. A reduction in the level of such motions was detected upon complex formation, indicating rigidification of the protein structure and loss of conformational entropy. This hypothesis was confirmed by isothermal titration calorimetry, showing that ligand binding is enthalpy-driven, thus overcompensating for the negative entropy associated with both ligand binding per se and rigidification of the protein. Our investigation of the solution structure and dynamics as well as thermodynamics of lipocalin-ligand interaction not only provides insight into the general mechanism of small molecule accommodation in the deep and narrow cavity of this abundant class of proteins but also supports the future design of corresponding binding proteins with novel specificities, so-called "anticalins".

Evaluation of the Safety and Tolerability of Conjunctival Ring for Posterior Segment of the Eye.

PubMed

Kinoshita, Satoshi; Ohguchi, Takeshi; Noda, Kousuke; Murata, Miyuki; Yasueda, Shin-Ichi; Obata, Haruka; Matsunaga, Toru; Fukushima, Tsutomu; Kanda, Atsuhiro; Ishida, Susumu

2017-08-01

To evaluate the safety and tolerability of conjunctival rings (CRs), a novel device for drug delivery to the posterior segment of the eye. In animal studies, CRs containing 5% dexamethasone sodium phosphate (DSP) or vehicle solution were placed on the right and left eyes of C57BL/6J mice, respectively. Contact lenses (CLs) containing vehicle solution were used as a control. Twenty-four hours after placement of the CRs, corneal fluorescein staining was graded based on the McDonald-Shadduck scoring system, ranging from 0 to 4. In humans, CRs containing vehicle solution were placed on the right eye of healthy volunteers for 9 hours. The corneal curvature, corneal thickness, intraocular pressure, visual acuity, tear production (Schirmer I test), tear film break-up time and fluorescein staining scores of the cornea (scores ranging from 0 to 3) and conjunctiva (scores ranging from 0 to 6) were assessed before and after wearing the CRs. The release characteristics of DSP from CRs were also evaluated. In animal experiments, corneal fluorescein staining scores were 1 or less in all the groups, and there was no significant difference between the CR group and the CL group. In the preclinical safety evaluation of CR for humans, ophthalmic examination revealed that CR caused no significant changes in all the parameters investigated including corneal curvature (p = 0.77), corneal thickness (p = 0.96), intraocular pressure (p = 0.59), visual acuity (p = 0.14), Schirmer I test results (p = 0.76), tear film break-up time (p = 0.68), corneal fluorescein staining scores (p = 0.64), and conjunctival fluorescein staining scores (p = 0.52). The DSP release from CRs occurs within a few hours, which is similar to the drug-release property of medicated CL, as reported previously. The current data showed the safety and tolerability of CR as a drug delivery device for the treatment of posterior segment diseases.

Net Fluorescein Flux Across Corneal Endothelium Strongly Suggests Fluid Transport is due to Electro-osmosis.

PubMed

Sanchez, J M; Cacace, V; Kusnier, C F; Nelson, R; Rubashkin, A A; Iserovich, P; Fischbarg, J

2016-08-01

We have presented prior evidence suggesting that fluid transport results from electro-osmosis at the intercellular junctions of the corneal endothelium. Such phenomenon ought to drag other extracellular solutes. We have investigated this using fluorescein-Na2 as an extracellular marker. We measured unidirectional fluxes across layers of cultured human corneal endothelial (HCE) cells. SV-40-transformed HCE layers were grown to confluence on permeable membrane inserts. The medium was DMEM with high glucose and no phenol red. Fluorescein-labeled medium was placed either on the basolateral or the apical side of the inserts; the other side carried unlabeled medium. The inserts were held in a CO2 incubator for 1 h (at 37 °C), after which the entire volume of the unlabeled side was collected. After that, label was placed on the opposite side, and the corresponding paired sample was collected after another hour. Fluorescein counts were determined with a (Photon Technology) DeltaScan fluorometer (excitation 380 nm; emission 550 nm; 2 nm bwth). Samples were read for 60 s. The cells utilized are known to transport fluid from the basolateral to the apical side, just as they do in vivo in several species. We used 4 inserts for influx and efflux (total: 20 1-h periods). We found a net flux of fluorescein from the basolateral to the apical side. The flux ratio was 1.104 ± 0.056. That difference was statistically significant (p = 0.00006, t test, paired samples). The endothelium has a definite restriction at the junctions. Hence, an asymmetry in unidirectional fluxes cannot arise from osmosis, and can only point instead to paracellular solvent drag. We suggest, once more, that such drag is due to electro-osmotic coupling at the paracellular junctions.

Effects of loading concentration, blood and synovial fluid on antibiotic release and anti-biofilm activity of bone cement beads.

PubMed

Dusane, Devendra H; Diamond, Scott M; Knecht, Cory S; Farrar, Nicholas R; Peters, Casey W; Howlin, Robert P; Swearingen, Matthew C; Calhoun, Jason H; Plaut, Roger D; Nocera, Tanya M; Granger, Jeffrey F; Stoodley, Paul

2017-02-28

Antibiotic loaded cement beads are commonly used for the treatment of biofilm related orthopaedic periprosthetic infections; however the effects of antibiotic loading and exposure of beads to body fluids on release kinetics are unclear. The purpose of this study was to determine the effects of (i) antibiotic loading density (ii) loading amount (iii) material type and (iv) exposure to body fluids (blood or synovial fluid) on release kinetics and efficacy of antibiotics against planktonic and lawn biofilm bacteria. Short-term release into an agar gel was evaluated using a fluorescent tracer (fluorescein) incorporated in the carrier materials calcium sulfate (CaSO 4 ) and poly methyl methacrylate (PMMA). Different fluorescein concentrations in CaSO 4 beads were evaluated. Mechanical properties of fluorescein-incorporated beads were analyzed. Efficacy of the antibiotics vancomycin (VAN) or tobramycin (TOB) alone and in combination was evaluated against lawn biofilms of bioluminescent strains of Staphylococcus aureus and Pseudomonas aeruginosa. Zones of inhibition of cultures (ZOI) were measured visually and using an in-vivo imaging system (IVIS). The influence of body fluids on release was assessed using CaSO 4 beads that contained fluorescein or antibiotics and were pre-coated with human blood or synovial fluid. The spread from the beads followed a square root of time relationship in all cases. The loading concentration had no influence on short-term fluorescein release and pre-coating of beads with body fluids did not affect short-term release or antibacterial activity. Compared to PMMA, CaSO 4 had a more rapid short term rate of elution and activity against planktonic and lawn biofilms. This study highlights the importance of considering antibiotic loading and packing density when investigating the clinical application of bone cements for infection management. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Sarcoid granuloma of the choroid.

PubMed

Marcus, D F; Bovino, J A; Burton, T C

1982-12-01

Two patients were found to have macular choroidal granulomas associated with systemic sarcoidosis. This unusual fundus picture was documented by serial fundus photography and fluorescein angiography. Both patients had a similar clinical picture of decreased vision, chorioretinal granulomas, and overlying neurosensory detachments. Staining of the inflammatory mass with fluorescein and leakage of dye into the neurosensory space was typical. Lymph node biopsies were performed to substantiate the diagnosis. Both patients responded promptly to systemic corticosteroid therapy with dramatic improvement in visual acuity and resolution of the choroidal lesions.

pH sensor based on boron nitride nanotubes.

PubMed

Huang, Q; Bando, Y; Zhao, L; Zhi, C Y; Golberg, D

2009-10-14

A submicrometer-sized pH sensor based on biotin-fluorescein-functionalized multiwalled BN nanotubes with anchored Ag nanoparticles is designed. Intrinsic pH-dependent photoluminescence and Raman signals in attached fluorescein molecules enhanced by Ag nanoparticles allow this novel nanohybrid to perform as a practical pH sensor. It is able to work in a submicrometer-sized space. For example, the sensor may determine the environmental pH of sub-units in living cells where a traditional optical fiber sensor fails because of spatial limitations.

pH sensor based on boron nitride nanotubes

NASA Astrophysics Data System (ADS)

Huang, Q.; Bando, Y.; Zhao, L.; Zhi, C. Y.; Golberg, D.

2009-10-01

A submicrometer-sized pH sensor based on biotin-fluorescein-functionalized multiwalled BN nanotubes with anchored Ag nanoparticles is designed. Intrinsic pH-dependent photoluminescence and Raman signals in attached fluorescein molecules enhanced by Ag nanoparticles allow this novel nanohybrid to perform as a practical pH sensor. It is able to work in a submicrometer-sized space. For example, the sensor may determine the environmental pH of sub-units in living cells where a traditional optical fiber sensor fails because of spatial limitations.

Development of LEDs-based microplate reader for bioanalytical assay measurements

NASA Astrophysics Data System (ADS)

Alaruri, Sami D.; Katzlinger, Michael; Schinwald, Bernhard; Kronberger, Georg; Atzler, Joseph

2013-10-01

The optical design for an LEDs-based microplate reader that can perform fluorescence intensity (top and bottom), absorbance, luminescence and time-resolved fluorescence measurements is described. The microplate reader is the first microplate reader in the marketplace that incorporates LEDs as excitation light sources. Absorbance measurements over the 0-3.5 optical density range for caffeine solution are presented. Additionally, fluorescence intensity readings collected at 535 and 625 nm from a green and a red RediPlateTM are reported. Furthermore, fluorescence decay lifetime measurements obtained for Eu (europium) and Sm (samarium) standard solutions using 370 nm excitation are presented. The microplate reader detection limits for the fluorescence intensity top, fluorescence intensity bottom, fluorescence polarization and time-resolved fluorescence modes are 1.5 fmol 100 µL-1 fluorescein (384-well plate), 25 fmol 100 µL-1 fluorescein (384-well plate), 5 mP at 10 nM fluorescein (black 384-well plate) and 30 amol 100 µL-1 europium solution (white 384-well plate), respectively.

Detection of new-onset choroidal neovascularization.

PubMed

Do, Diana V

2013-05-01

To highlight the most common methods that are used to detect new-onset choroidal neovascularization (CNV) as a result of age-related macular degeneration (AMD). Numerous modalities are available to try to detect CNV. Amsler grid testing, preferential hyperacuity perimetry (PHP), optical coherence tomography (OCT), and fluorescein angiography are tools that may be used to detect CNV. The Age-Related Macular Degeneration: Detection of Onset of new Choroidal neovascularization Study (AMD DOC Study) evaluated the sensitivity of time domain OCT, relative to fluorescein angiography, in detecting new-onset neovascular AMD within a 2-year period. The sensitivity of each modality for detecting CNV was OCT 0.40 [(95% confidence interval (95% CI) (0.16-0.68), supervised Amsler grid 0.42 (95% CI 0.15-0.72), and PHP 0.50 (95% CI 0.23-0.77)]. Numerous modalities are available to try to detect CNV. The prospective AMD DOC Study demonstrated that fluorescein angiography still remains the best method to detect new-onset CNV.

Photodynamic therapy for treatment subretinal neovascularization

NASA Astrophysics Data System (ADS)

Avetisov, Sergey E.; Budzinskaja, Maria V.; Kiseleva, Tatyana N.; Balatskaya, Natalia V.; Gurova, Irina V.; Loschenov, Viktor B.; Shevchik, Sergey A.; Kuzmin, Sergey G.; Vorozhtsov, Georgy N.

2007-07-01

This work are devoted our experience with photodynamic therapy (PDT) with <> for patients with choroidal neovascularization (CNV). 18 patients with subfoveal CNV in age-related macular degeneration (AMD), 24 patients with subfoveal CNV in pathological myopia (PM) and 4 patients with subfoveal CNV associated with toxoplasmic retinochoroiditis were observed. CNV was 100% classic in all study patients. Standardized protocol refraction, visual acuity testing, ophthalmologic examinations, biomicroscopy, fluorescein angiography, and ultrasonography were performed before treatment and 1 month, 3 months, 6 months, and 1 year after treatment; were used to evaluate the results of photodynamic therapy with <> (0.02% solution of mixture sulfonated aluminium phtalocyanine 0.05 mg/kg, intravenously). A diode laser (<>, Inc, Moscow) was used operating in the range of 675 nm. Need for retreatment was based on fluorescein angiographic evidence of leakage at 3-month follow-up intervals. At 3, 6, 9 month 26 (56.5%) patients had significant improvement in the mean visual acuity. At the end of the 12-month minimal fluorescein leakage from choroidal neovascularization was seen in 12 (26.1%) patients and the mean visual acuity was slightly worse than 0.2 which was not statistically significant as compared with the baseline visual acuity. Patients with fluorescein leakage from CNV underwent repeated PDT with <>. 3D-mode ultrasound shown the decreasing thickness of chorioretinal complex in CNV area. Photodynamic therapy with <> can safely reduce the risk of severe vision loss in patients with predominantly classic subfoveal choroidal neovascularization secondary to AMD, PM and toxoplasmic retinochoroiditis.

A hot-spot-active magnetic graphene oxide substrate for microRNA detection based on cascaded chemiluminescence resonance energy transfer.

PubMed

Bi, Sai; Chen, Min; Jia, Xiaoqiang; Dong, Ying

2015-02-28

Herein, a cascaded chemiluminescence resonance energy transfer (C-CRET) process was demonstrated from horseradish peroxidase (HRP)-mimicking DNAzyme-catalyzed luminol-H2O2 to fluorescein and further to graphene oxide (GO) when HRP-mimicking DNAzyme/fluorescein was in close proximity to the GO surface. The proposed C-CRET system was successfully implemented to construct three modes of C-CRET hot-spot-active substrates (modes I, II and III) by covalently immobilizing HRP-mimicking DNAzyme/fluorescein-labeled hairpin DNAs (hot-spot-generation probes) on magnetic GO (MGO), resulting in a signal "off" state due to the quenching of the luminol/H2O2/HRP-mimicking DNAzyme/fluorescein CRET system by GO. Upon the introduction of microRNA-122 (miRNA-122), the targets (mode I) or the new triggers that were generated through a strand displacement reaction (SDR) initiated by miRNA-122 (modes II and III) hybridized with the loop domains of hairpin probes on MGO to form double-stranded (modes I and II) or triplex-stem structures (mode III), causing an "open" configuration of the hairpin probe and a CRET signal "on" state, thus achieving sensitive and selective detection of miRNA-122. More importantly, the substrate exhibited excellent controllability, reversibility and reproducibility through SDR and magnetic separation (modes II and III), especially sequence-independence for hairpin probes in mode III, holding great potential for the development of a versatile platform for optical biosensing.

Murine fundus fluorescein angiography: An alternative approach using a handheld camera.

PubMed

Ehrenberg, Moshe; Ehrenberg, Scott; Schwob, Ouri; Benny, Ofra

2016-07-01

In today's modern pharmacologic approach to treating sight-threatening retinal vascular disorders, there is an increasing demand for a compact, mobile, lightweight and cost-effective fluorescein fundus camera to document the effects of antiangiogenic drugs on laser-induced choroidal neovascularization (CNV) in mice and other experimental animals. We have adapted the use of the Kowa Genesis Df Camera to perform Fundus Fluorescein Angiography (FFA) in mice. The 1 kg, 28 cm high camera has built-in barrier and exciter filters to allow digital FFA recording to a Compact Flash memory card. Furthermore, this handheld unit has a steady Indirect Lens Holder that firmly attaches to the main unit, that securely holds a 90 diopter lens in position, in order to facilitate appropriate focus and stability, for photographing the delicate central murine fundus. This easily portable fundus fluorescein camera can effectively record exceptional central retinal vascular detail in murine laser-induced CNV, while readily allowing the investigator to adjust the camera's position according to the variable head and eye movements that can randomly occur while the mouse is optimally anesthetized. This movable image recording device, with efficiencies of space, time, cost, energy and personnel, has enabled us to accurately document the alterations in the central choroidal and retinal vasculature following induction of CNV, implemented by argon-green laser photocoagulation and disruption of Bruch's Membrane, in the experimental murine model of exudative macular degeneration. Copyright © 2016 Elsevier Ltd. All rights reserved.

Empty Turnip yellow mosaic virus capsids as delivery vehicles to mammalian cells.

PubMed

Kim, Doyeong; Lee, Younghee; Dreher, Theo W; Cho, Tae-Ju

2018-05-03

Turnip yellow mosaic virus (TYMV) was able to enter animal cells when the spherical plant virus was conjugated with Tat, a cell penetrating peptide (CPP). Tat was chemically attached to the surface lysine residues of TYMV using hydrazone chemistry. Baby hamster kidney (BHK) cells were incubated with either unmodified or Tat-conjugated TYMV and examined by flow cytometry and confocal microscopic analyses. Tat conjugation was shown to be more efficient than Lipofectamine in allowing TYMV to enter the mammalian cells. Tat-assisted-transfection was also associated with less loss of cell viability than lipofection. Among the CPPs tested (Tat, R8, Pep-1 and Pen), it was observed that R8 and Pen were also effective while Pep-1 was not. We also examined if the internal space of TYMV can be used to load fluorescein dye as a model cargo. When TYMV is treated by freezing and thawing, the virus is known to convert into a structure with a 6-8 nm hole and release viral RNA. When the resultant pot-like particles were reacted with fluorescein-5-maleimide using interior sulfhydryl groups as conjugation sites, about 145 fluorescein molecules were added per particle. The fluorescein-loaded TYMV particles were conjugated with Tat and introduced into BHK cells, again with higher transfection efficiency compared to lipofection. Our studies demonstrate the potential of modified TYMV as an efficient system for therapeutic cargo delivery to mammalian cells. Copyright © 2018 Elsevier B.V. All rights reserved.

EVALUATION OF MULTISPECTRAL IMAGING IN DIAGNOSING DIABETIC RETINOPATHY.

PubMed

Li, Liang; Zhang, Pu; Liu, Hong; Liu, Yu-Hua; Gao, Ling

2018-06-12

To investigate multispectral imaging (MSI) as a novel diagnostic approach for diabetic retinopathy (DR) in clinic. A total of 50 Type-2 diabetic patients (99 eyes) were enrolled in this cross-sectional study. All subjects underwent digital fundus photography (DFP), MSI, and fundus fluorescein angiography. A total exact agreement, sensitivity, specificity, positive predictive value, and negative predictive value of no DR/mild nonproliferative diabetic retinopathy (NPDR) and severe NPDR/proliferative diabetic retinopathy (PDR) grading were calculated based on DFP and MSI and were compared with fundus fluorescein angiography. Compared with fundus fluorescein angiography, the exact agreement for MSI was 0.835; for DFP, it was 0.614; the sensitivity for no DR/mild NPDR in both MSI and DFP was 100%, and for severe NPDR/PDR, it was 97.4% and 88.3%. The specificity for no DR/mild NPDR in MSI and DFP was 96.3% and 95%, and for severe NPDR/PDR, it was 100% in both. The positive predictive value for no DR/mild NPDR in MSI and DFP was 86.4% and 82.6%, and for severe NPDR/PDR, it was 100% in both; the negative predictive value for no DR/mild NPDR in MSI and DFP was 100%, and for severe NPDR/PDR, it was 91.7% and 71.0% in both. Multispectral imaging displayed an excellent agreement with fundus fluorescein angiography in DR grading, which suggested that it might serve as a new diagnostic technique and an informative tool for evaluating DR.

The photostability of the commonly used biotin-4-fluorescein probe.

PubMed

Haack, Richard A; Swift, Kerry M; Ruan, Qiaoqiao; Himmelsbach, Richard J; Tetin, Sergey Y

2017-08-15

Biotin-4-fluorescein (B4F) is a commonly used fluorescent probe for studying biotin-(strept)avidin interactions. During a characterization study of an anti-biotin antibody, using B4F as the probe, we noticed a discrepancy in the expected and experimentally determined number of biotin binding sites. Analytical testing showed that the biotin moiety in the probe undergoes a photosensitized oxidation to produce a mixture of biotin sulfoxides which has the potential to impact the quantitation of binding sites using this fluorescent probe. Copyright © 2017 Elsevier Inc. All rights reserved.

Fluorogenic Substrate Detection of Viable Intracellular and Extracellular Pathogenic Protozoa

NASA Astrophysics Data System (ADS)

Jackson, Peter R.; Pappas, Michael G.; Hansen, Brian D.

1985-01-01

Viable Leishmania promastigotes and amastigotes were detected by epifluorescence microscopy with fluorescein diacetate being used to mark living parasites and the nucleic acid-binding compound ethidium bromide to stain dead cells. This procedure is superior to other assays because it is faster and detects viable intracellular as well as extracellular Leishmania. Furthermore, destruction of intracellular pathogens by macrophages is more accurately determined with fluorescein diacetate than with other stains. The procedure may have applications in programs to develop drugs and vaccines against protozoa responsible for human and animal disease.

Fluorescence imaging of the nanoparticles modified with indocyanine green

NASA Astrophysics Data System (ADS)

Gareev, K. G.; Babikova, K. Y.; Postnov, V. N.; Naumisheva, E. B.; Korolev, D. V.

2017-11-01

The comparative research of silica, the magnetite and magnetite-silica nanoparticles modified with fluorescent dyes using gas-phase and liquid-phase methods was conducted. At the content of fluorescent dye comparable in size a particular spectrophotometric method, nanoparticles with fluorescein have up to 1000 times larger overall luminous efficiency. It is revealed that magnetic nanoparticles are characterized by a smaller light efficiency in comparison with silica particles, at the same time particles of a magnetite are most effective at modification with fluorescein, and magnetite-silica particles - at modification with indocyanine green.

Multispectral Fluorescence Imaging During Robot-assisted Laparoscopic Sentinel Node Biopsy: A First Step Towards a Fluorescence-based Anatomic Roadmap.

PubMed

van den Berg, Nynke S; Buckle, Tessa; KleinJan, Gijs H; van der Poel, Henk G; van Leeuwen, Fijs W B

2017-07-01

During (robot-assisted) sentinel node (SN) biopsy procedures, intraoperative fluorescence imaging can be used to enhance radioguided SN excision. For this combined pre- and intraoperative SN identification was realized using the hybrid SN tracer, indocyanine green- 99m Tc-nanocolloid. Combining this dedicated SN tracer with a lymphangiographic tracer such as fluorescein may further enhance the accuracy of SN biopsy. Clinical evaluation of a multispectral fluorescence guided surgery approach using the dedicated SN tracer ICG- 99m Tc-nanocolloid, the lymphangiographic tracer fluorescein, and a commercially available fluorescence laparoscope. Pilot study in ten patients with prostate cancer. Following ICG- 99m Tc-nanocolloid administration and preoperative lymphoscintigraphy and single-photon emission computed tomograpy imaging, the number and location of SNs were determined. Fluorescein was injected intraprostatically immediately after the patient was anesthetized. A multispectral fluorescence laparoscope was used intraoperatively to identify both fluorescent signatures. Multispectral fluorescence imaging during robot-assisted radical prostatectomy with extended pelvic lymph node dissection and SN biopsy. (1) Number and location of preoperatively identified SNs. (2) Number and location of SNs intraoperatively identified via ICG- 99m Tc-nanocolloid imaging. (3) Rate of intraoperative lymphatic duct identification via fluorescein imaging. (4) Tumor status of excised (sentinel) lymph node(s). (5) Postoperative complications and follow-up. Near-infrared fluorescence imaging of ICG- 99m Tc-nanocolloid visualized 85.3% of the SNs. In 8/10 patients, fluorescein imaging allowed bright and accurate identification of lymphatic ducts, although higher background staining and tracer washout were observed. The main limitation is the small patient population. Our findings indicate that a lymphangiographic tracer can provide additional information during SN biopsy based on ICG- 99m Tc-nanocolloid. The study suggests that multispectral fluorescence image-guided surgery is clinically feasible. We evaluated the concept of surgical fluorescence guidance using differently colored dyes that visualize complementary features. In the future this concept may provide better guidance towards diseased tissue while sparing healthy tissue, and could thus improve functional and oncologic outcomes. Copyright © 2016 European Association of Urology. Published by Elsevier B.V. All rights reserved.

Comparison of in vivo efficacy of different ocular lubricants in dry eye animal models.

PubMed

Zheng, Xiaodong; Goto, Tomoko; Ohashi, Yuichi

2014-04-29

To compare the efficacy of three types of ocular lubricants in protecting corneal epithelial cells in dry eye animal models. Ocular lubricants containing 0.1% or 0.3% sodium hyaluronate (SH), carboxymethylcellulose (CMC), or hydroxypropyl methylcellulose (HPMC) were tested. First, ocular lubricant containing 0.002% fluorescein was dropped onto the rabbit corneas. The fluorescein intensity as an index of retention was measured. Second, a rabbit dry eye model was made by holding the eye open with a speculum, and 50 μL of each ocular lubricant was dropped onto the cornea. After 3 hours, the corneas were stained with 1% methylene blue (MB), and the absorbance of MB was measured. Third, a rat dry eye model was treated with the ocular lubricants for 4 weeks, and the corneal fluorescein staining was scored. Eyes treated with physiological saline were used as controls. Finally, immunohistochemistry was used to analyze occludin, an epithelial barrier protein, in cultured human corneal epithelial cells pretreated with ocular lubricants and desiccated for 20 or 60 minutes. Our results showed that 0.3% SH had a significantly longer retention time than the other lubricants (all P < 0.01). The absorbance of MB was significantly lower in the 0.3% SH group. The corneas of rats exposed to 0.3% SH had significantly lower fluorescein staining scores. A significantly higher number of occludin-positive cells were found after exposure to 0.3% SH than other lubricants. Ocular lubricant containing 0.3% SH would be preferable to treat patients with dry eye syndrome. Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.

A microfluidic device for studying cell signaling with multiple inputs and adjustable amplitudes and frequencies

NASA Astrophysics Data System (ADS)

Ningsih, Zubaidah; Chon, James W. M.; Clayton, Andrew H. A.

2013-12-01

Cell function is largely controlled by an intricate web of macromolecular interactions called signaling networks. It is known that the type and the intensity (concentration) of stimulus affect cell behavior. However, the temporal aspect of the stimulus is not yet fully understood. Moreover, the process of distinguishing between two stimuli by a cell is still not clear. A microfluidic device enables the delivery of a precise and exact stimulus to the cell due to the laminar flow established inside its micro-channel. The slow stream delivers a constant stimulus which is adjustable according to the experiment set up. Moreover, with controllable inputs, microfluidic facilitates the stimuli delivery according to a certain pattern with adjustable amplitude, frequency and phase. Several designs of PDMS microfluidic device has been produced in this project via photolithography and soft lithography processes. To characterize the microfluidic performance, two experiments has been conducted. First, by comparing the fluorescence intensity and the lifetime of fluorescein in the present of KI, mixing extent between two inputs was observed using Frequency Lifetime Imaging Microscopy (FLIM). Furthermore, the input-output relationship of fluorescein concentration delivered was also drawn to characterize the amplitude, frequency and phase of the inputs. Second experiment involved the cell culturing inside microfluidic. Using NG108-15 cells, proliferation and differentiation were observed based on the cell number and cell physiological changes. Our results demonstrate that hurdle design gives 86% mixing of fluorescein and buffer. Relationship between inputoutput fluorescein concentrations delivered has also been demonstrated and cells were successfully cultured inside the microfluidic.

Development and characterization of glutathione-conjugated albumin nanoparticles for improved brain delivery of hydrophilic fluorescent marker.

PubMed

Patel, Prerak J; Acharya, Niyati S; Acharya, Sanjeev R

2013-01-01

The glutathione-conjugated bovine serum albumin (BSA) nanoparticles were constructed in the present exploration as a novel biodegradable carrier for brain-specific drug delivery with evaluation of its in vitro and in vivo delivery properties. BSA nanocarriers were activated and conjugated to the distal amine functions of the glutathione via carbodiimide chemistry using EDAC as a mediator. These nanoparticles were characterized for particle shape, average size, SPAN value, drug entrapment and in vitro drug release. Further, presence of glutathione on the surface of BSA nanoparticles was confirmed by Ellman's assay, which has suggested that approximately 750 units of glutathione were conjugated per BSA nanoparticle. To evaluate the brain delivery properties of the glutathione-conjugated BSA nanoparticles fluorescein sodium was used as a model hydrophilic compound. Permeability and neuronal uptake properties of developed formulations were evaluated against the MDCK-MDR1 endothelial and neuro-glial cells, respectively. The permeability of glutathione-conjugated BSA nanoparticles across the monolayer of MDCK-MDR1 endothelial tight junction was shown significantly higher than that of unconjugated nanoparticles and fluorescein sodium solution. Similarly, glutathione-conjugated nanoparticles exhibited considerably higher uptake by neuro-glial cells which was inferred by high fluorescence intensity under microscope in comparison to unconjugated nanoparticles and fluorescein sodium solution. Following an intravenous administration, nearly three folds higher fluorescein sodium was carried to the rat brain by glutathione-conjugated nanoparticles as compared to unconjugated nanoparticles. The significant in vitro and in vivo results suggest that glutathione-conjugated BSA nanoparticles is a promising brain drug delivery system with low toxicity.

In vivo Delivery of Fluoresceinated Dextrans to the Murine Growth Plate: Imaging of Three Vascular Routes by Multiphoton Microscopy

PubMed Central

Farnum, Cornelia; Lenox, Michelle; Zipfel, Warren; Horton, William; Williams, Rebecca

2008-01-01

Bone elongation by endochondral ossification occurs through the differentiation cascade of chondrocytes of cartilaginous growth plates. Molecules from the systemic vasculature reach the growth plate from three different directions: epiphyseal, metaphyseal, and via a ring vessel and plexus associated with the perichondrium. This study is an analysis of the real-time dynamics of entrance of fluoresceinated tracers of different molecular weights into the growth plate from the systemic vasculature, and tests the hypothesis that molecular weight is a key variable in the determination of both the directionality and the extent of tracer movement into the growth plate. Multiphoton microscopy was used for direct in vivo imaging of the murine proximal tibial growth plate in anesthetized 4-5-week-old transgenic mice with green fluorescent protein linked to the collagen II promoter. Mice were given an intracardiac injection of either fluorescein (332.3 Da), or fluoresceinated dextrans of 3, 10, 40, 70 kDa, singly or sequentially. For each tracer, directionality and rate of arrival, together with extent of movement within the growth plate, were imaged in real time. For small molecules (up to 10 kDa) vascular access from all three directions was observed and entrance was equally permissive from the metaphyseal and the epiphyseal sides. Within our detection limit (a few per cent of vascular concentration) 40 kDa and larger dextrans did not enter. These results have implications both for understanding systemic and paracrine regulation of growth plate chondrocytic differentiation, as well as variables associated with effective drug delivery to growth plate chondrocytes. PMID:16342207

An investigation of paper based microfluidic devices for size based separation and extraction applications.

PubMed

Zhong, Z W; Wu, R G; Wang, Z P; Tan, H L

2015-09-01

Conventional microfluidic devices are typically complex and expensive. The devices require the use of pneumatic control systems or highly precise pumps to control the flow in the devices. This work investigates an alternative method using paper based microfluidic devices to replace conventional microfluidic devices. Size based separation and extraction experiments conducted were able to separate free dye from a mixed protein and dye solution. Experimental results showed that pure fluorescein isothiocyanate could be separated from a solution of mixed fluorescein isothiocyanate and fluorescein isothiocyanate labeled bovine serum albumin. The analysis readings obtained from a spectrophotometer clearly show that the extracted tartrazine sample did not contain any amount of Blue-BSA, because its absorbance value was 0.000 measured at a wavelength of 590nm, which correlated to Blue-BSA. These demonstrate that paper based microfluidic devices, which are inexpensive and easy to implement, can potentially replace their conventional counterparts by the use of simple geometry designs and the capillary action. These findings will potentially help in future developments of paper based microfluidic devices. Copyright © 2015 Elsevier B.V. All rights reserved.

Fluorescence Ratiometric Assay Strategy for Chemical Transmitter of Living Cells Using H2O2-Sensitive Conjugated Polymers.

PubMed

Wang, Yunxia; Li, Shengliang; Feng, Liheng; Nie, Chenyao; Liu, Libing; Lv, Fengting; Wang, Shu

2015-11-04

A new water-soluble conjugated poly(fluorene-co-phenylene) derivative (PFP-FB) modified with boronate-protected fluorescein (peroxyfluor-1) via PEG linker has been designed and synthesized. In the presence of H2O2, the peroxyfluor-1 group can transform into green fluorescent fluorescein by deprotecting the boronate protecting groups. In this case, upon selective excitation of PFP-FB backbone at 380 nm, efficient fluorescence resonance energy transfer (FRET) from PFP-FB backbone to fluorescein occurs, and accordingly, the fluorescence color of PFP-FB changes from blue to green. Furthermore, the emission color of PFP-FB and the FRET ratio change in a concentration-dependent manner. By taking advantage of PFP-FB, ratiometric detection of choline and acetylcholine (ACh) through cascade enzymatic reactions and further dynamic monitoring of the choline consumption process of cancer cells have been successfully realized. Thus, this new polymer probe promotes the development of enzymatic biosensors and provides a simpler and more effective way for detecting the chemical transmitter of living cells.

Direct comparison of single- and multi-walled carbon nanotubes in fluorescence quenching phenomenon

NASA Astrophysics Data System (ADS)

Oura, Shusuke; Umemura, Kazuo

2018-03-01

Here, we report the fluorescence quenching ability of single-stranded DNA (ssDNA)-wrapped single- and multi-walled carbon nanotubes (ssDNA-SWNTs and ssDNA-MWNTs, respectively) using fluorescein dye-labeled ssDNA (Fluor-ssDNA). To compare the quenching abilities of SWNTs and MWNTs, we measured the quenching ratios of fluorescence emission from fluorescein when Fluor-ssDNA reacted with the hybrids of 30-mers of thymine (T30) and SWNTs or MWNTs (T30-SWNTs and T30-MWNTs, respectively). The fluorescence quenching ratios of Fluor-T30 in SWNT and MWNT samples were 28 ± 3.1 and 36 ± 2.0% relative to free fluorescein at the same concentration, respectively. On the other hand, those of Fluor-A30 with SWNT and MWNT hybrids were 11 ± 1.9 and 32 ± 1.9%, respectively. Our results suggest that although the fluorescence quenching ability of MWNT was greater than that of SWNT, SWNT quenching ratios were more sensitive to the base sequences of Fluor-ssDNA.

Tailoring of optical properties of fluorescein using green synthesized gold nanoparticles.

PubMed

John, Jisha; Thomas, Lincy; George, Nibu A; Kurian, Achamma; George, Sajan D

2015-06-28

Dye-nanoparticle mixtures hold great promise in biological as well as photonics applications due to their capability to tailor the emission behavior of dye by tuning the nanoparticles parameters. However, as compared to the well-defined dye-nanoparticle distance, studies lack the understanding of homogenous mixtures of dye and nanoparticles. In this work, we investigate the influence of shape and concentration of gold nanoparticles prepared via green synthesis on the optical properties of fluorescein dye in a dye-nanoparticle mixture. We have investigated the radiative path of deexcitation using steady state fluorescence and the non-radiative path is probed using a laser based dual-beam thermal lens technique. The energy transfer efficiency as well as dye-nanoparticle distance is studied using both techniques. Furthermore, we have explored the influence of nanoparticles parameters on the fluorescence quantum yield of fluorescein using the thermal lens technique. The studies indicate that spherical nanoparticles are efficient quenchers while star shaped nanoparticles can probe larger dye-NP distances. The tailoring of dye properties by tuning nanoparticle parameters can be utilized in diverse areas including bioimaging, solar cells, and sensors.

Individual marking of soft-bodied subtidal invertebrates in situ – A novel staining technique applied to the giant plumose anemone Metridium farcimen (Tilesius, 1809)

PubMed Central

Sebens, Kenneth P.

2017-01-01

The ability to recognize individuals and track growth over time is crucial to population dynamics research as well as studies of animal behavior. Invertebrates are particularly difficult to track as they often molt, have regenerative capabilities, or lack hard parts to attach markers. We tested, in laboratory and field studies, a new way of marking sea anemones (order Actiniaria) by injection of three vital stains (i.e., neutral red, methylene blue, and fluorescein). Neutral red and methylene blue did not affect growth or survival, but fluorescein was lethal at high concentrations. Marked individuals could be identified up to seven months after injection with neutral red, six weeks with methylene blue, and three days with low concentrations of fluorescein. Neutral red could be used for long-term monitoring of growth and survival in the field, and in combination with methylene blue could be used to mark individuals in distinguishable patterns for short-term studies such as examining predator-prey interactions, movement of individuals, and recruitment survival. PMID:29161292

Classification of Human Retinal Microaneurysms Using Adaptive Optics Scanning Light Ophthalmoscope Fluorescein Angiography

PubMed Central

Dubow, Michael; Pinhas, Alexander; Shah, Nishit; Cooper, Robert F.; Gan, Alexander; Gentile, Ronald C.; Hendrix, Vernon; Sulai, Yusufu N.; Carroll, Joseph; Chui, Toco Y. P.; Walsh, Joseph B.; Weitz, Rishard; Dubra, Alfredo; Rosen, Richard B.

2014-01-01

Purpose. Microaneurysms (MAs) are considered a hallmark of retinal vascular disease, yet what little is known about them is mostly based upon histology, not clinical observation. Here, we use the recently developed adaptive optics scanning light ophthalmoscope (AOSLO) fluorescein angiography (FA) to image human MAs in vivo and to expand on previously described MA morphologic classification schemes. Methods. Patients with vascular retinopathies (diabetic, hypertensive, and branch and central retinal vein occlusion) were imaged with reflectance AOSLO and AOSLO FA. Ninety-three MAs, from 14 eyes, were imaged and classified according to appearance into six morphologic groups: focal bulge, saccular, fusiform, mixed, pedunculated, and irregular. The MA perimeter, area, and feret maximum and minimum were correlated to morphology and retinal pathology. Select MAs were imaged longitudinally in two eyes. Results. Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging revealed microscopic features of MAs not appreciated on conventional images. Saccular MAs were most prevalent (47%). No association was found between the type of retinal pathology and MA morphology (P = 0.44). Pedunculated and irregular MAs were among the largest MAs with average areas of 4188 and 4116 μm2, respectively. Focal hypofluorescent regions were noted in 30% of MAs and were more likely to be associated with larger MAs (3086 vs. 1448 μm2, P = 0.0001). Conclusions. Retinal MAs can be classified in vivo into six different morphologic types, according to the geometry of their two-dimensional (2D) en face view. Adaptive optics scanning light ophthalmoscope fluorescein angiography imaging of MAs offers the possibility of studying microvascular change on a histologic scale, which may help our understanding of disease progression and treatment response. PMID:24425852

pH-Dependent silica nanoparticle dissolution and cargo release.

PubMed

Giovaninni, Giorgia; Moore, Colin J; Hall, Andrew J; Byrne, Hugh J; Gubala, Vladimir

2018-05-16

The dissolution of microporous silica nanoparticles (NP) in aqueous environments of different biologically relevant pH was studied in order to assess their potential as drug delivery vehicles. Silica NPs, loaded with fluorescein, were prepared using different organosilane precursors (tetraethoxysilane, ethyl triethoxysilane or a 1:1 molar ratio of both) and NP dissolution was evaluated in aqueous conditions at pH 4, pH 6 and pH 7.4. These conditions correspond to the acidity of the intracellular environment (late endosome, early endosome, cytosol respectively) and gastrointestinal tract ('fed' stomach, duodenum and jejunum respectively). All NPs degraded at pH 6 and pH 7.4, while no dissolution was observed at pH 4. NP dissolution could be clearly visualised as mesoporous hollows and surface defects using electron microscopy, and was supported by UV-vis, fluorimetry and DLS data. The dissolution profiles of the NPs are particularly suited to the requirements of oral drug delivery, whereby NPs must resist degradation in the harsh acidic conditions of the stomach (pH 4), but dissolve and release their cargo in the small intestine (pH 6-7.4). Particle cores made solely of ethyl triethoxysilane exhibited a 'burst release' of encapsulated fluorescein at pH 6 and pH 7.4, whereas NPs synthesised with tetraethoxysilane released fluorescein in a more sustained fashion. Thus, by varying the organosilane precursor used in NP formation, it is possible to modify particle dissolution rates and tune the release profile of encapsulated fluorescein. The flexible synthesis afforded by silica NPs to achieve pH-responsive dissolution therefore makes this class of nanomaterial an adaptable platform that may be well suited to oral delivery applications. Copyright © 2018 Elsevier B.V. All rights reserved.

Effect of ranibizumab on high-speed indocyanine green angiography and minimum intensity projection optical coherence tomography findings in neovascular age-related macular degeneration.

PubMed

Nicholson, Benjamin P; Nigam, Divya; Toy, Brian; Stetson, Paul F; Agrón, Elvira; Jacobs-El, Naima; Cunningham, Denise; Cukras, Catherine; Wong, Wai; Wiley, Henry; Chew, Emily; Ferris, Frederick; Meyerle, Catherine B

2015-01-01

The purpose of this 1-year prospective study was to investigate how induction/pro re nata ranibizumab intravitreal treatment of eyes with neovascular age-related macular degeneration affects the anatomy of choroidal neovascularization (CNV) and the overlying outer retinal tissue. High-speed indocyanine green (HS-ICG) angiography measurements provided quantification of the CNV size in 60 patients followed for 1 year. Minimum intensity projection optical coherence tomography (MinIP OCT), a novel algorithm assessing minimum optical intensity between the internal limiting membrane and retinal pigment epithelium, measured the area of outer retinal disruption overlying the CNV. Fluorescein angiography was also assessed to evaluate late retinal leakage. After 1 year, the mean area of CNV measured with indocyanine green angiography decreased by 5.8%. The mean area of MinIP OCT of outer retinal disruption overlying the CNV decreased by 4.2%. Mean area of fluorescein angiography leakage decreased by 6.3%. Both the area of outer retinal disruption measured with MinIP OCT and the area of leakage on fluorescein angiography typically exceeded the area of CNV on indocyanine green angiography at baseline and 1 year. Choroidal neovascularization treated with induction/pro re nata intravitreal ranibizumab for 1 year essentially remained static. Minimum intensity projection optical coherence tomography suggests that the area of outer retinal disruption overlying the CNV may be greater than the CNV itself and often correlates with the leakage area on fluorescein angiography. Additionally, there was minimal change in the area of outer retinal disruption on MinIP OCT even when fluid resolved. Measurements of the extent of CNV lesions based on indocyanine green angiography and MinIP OCT may provide useful outcome variables to help assess the CNV complex longitudinally and warrant further validation.

Degree of corneal anesthesia after topical application of 0.4% oxybuprocaine ophthalmic solution in normal equids.

PubMed

Little, Erika; Yvorchuk-St Jean, Kathy; Little, William; Sithole, Fortune; St Jean, Guy

2016-10-01

Oxybuprocaine hydrochloride ophthalmic solution has been widely used off-label in horses and donkeys, despite lack of data demonstrating efficacy and safety in these species. The objective of this study was to assess anesthetic efficacy of 0.4% oxybuprocaine hydrochloride ophthalmic solution in horses ( n = 5) and donkeys ( n = 24) and compare the effects with 0.5% proparacaine hydrochloride ophthalmic solution. The baseline corneal touch threshold (CTT) was measured with a Cochet-Bonnet esthesiometer. Donkeys ( n = 12) and horses ( n = 5) in group A received sterile ophthalmic solutions 0.4% oxybuprocaine with fluorescein (also termed benoxinate with fluorescein, abbreviated as ben + flu) instilled in one eye and 0.9% sterile sodium chloride solution (NaCl) with fluorescein (Na + flu) in the contralateral eye. Donkeys ( n = 12) and horses ( n = 5) in group B received sterile ophthalmic solutions (ben + flu) in one eye and 0.5% proparacaine with fluorescein (prop + flu) in the contralateral eye. The CTT was measured at 1 and 5 min post-application and at 5-minute intervals until 75 min after treatment. The CTT changes over time differed significantly between oxybuprocaine-treated and control eyes ( P < 0.001). The CTT continued to decrease throughout the duration of the study when compared with baseline values. No statistically significant difference in onset, depth, or duration of corneal anesthesia was found between oxybuprocaine and proparacaine treated eyes during the time of the study. Interestingly, horses were shown to have a significantly more sensitive cornea than donkeys ( P = 0.002). Oxybuprocaine and proparacaine reduced corneal sensitivity in donkeys and horses. No local irritation was observed with 0.4% oxybuprocaine.

Degree of corneal anesthesia after topical application of 0.4% oxybuprocaine ophthalmic solution in normal equids

PubMed Central

Little, Erika; Yvorchuk-St. Jean, Kathy; Little, William; Sithole, Fortune; St. Jean, Guy

2016-01-01

Oxybuprocaine hydrochloride ophthalmic solution has been widely used off-label in horses and donkeys, despite lack of data demonstrating efficacy and safety in these species. The objective of this study was to assess anesthetic efficacy of 0.4% oxybuprocaine hydrochloride ophthalmic solution in horses (n = 5) and donkeys (n = 24) and compare the effects with 0.5% proparacaine hydrochloride ophthalmic solution. The baseline corneal touch threshold (CTT) was measured with a Cochet-Bonnet esthesiometer. Donkeys (n = 12) and horses (n = 5) in group A received sterile ophthalmic solutions 0.4% oxybuprocaine with fluorescein (also termed benoxinate with fluorescein, abbreviated as ben + flu) instilled in one eye and 0.9% sterile sodium chloride solution (NaCl) with fluorescein (Na + flu) in the contralateral eye. Donkeys (n = 12) and horses (n = 5) in group B received sterile ophthalmic solutions (ben + flu) in one eye and 0.5% proparacaine with fluorescein (prop + flu) in the contralateral eye. The CTT was measured at 1 and 5 min post-application and at 5-minute intervals until 75 min after treatment. The CTT changes over time differed significantly between oxybuprocaine-treated and control eyes (P < 0.001). The CTT continued to decrease throughout the duration of the study when compared with baseline values. No statistically significant difference in onset, depth, or duration of corneal anesthesia was found between oxybuprocaine and proparacaine treated eyes during the time of the study. Interestingly, horses were shown to have a significantly more sensitive cornea than donkeys (P = 0.002). Oxybuprocaine and proparacaine reduced corneal sensitivity in donkeys and horses. No local irritation was observed with 0.4% oxybuprocaine. PMID:27733789

Degree of corneal anaesthesia after topical application of 0.4% oxybuprocaine hydrochloride and 0.5% proparacaine hydrochloride ophthalmic solution in clinically normal cattle.

PubMed

Little, W B; Jean, G St; Sithole, F; Little, E; Jean, K Yvorchuk-St

2016-06-01

The use of corneal anaesthesia is necessary for a range of clinical purposes. Therefore, we assessed and compared the efficacy of corneal anaesthesia after application of 0.4% oxybuprocaine hydrochloride and 0.5% proparacaine hydrochloride ophthalmic solution in clinically normal cattle. The 24 clinically normal cows were allocated into two groups. Cows in group 1 (n = 12) received 0.2 mL of 0.4% oxybuprocaine hydrochloride with fluorescein ophthalmic solution in one eye and 0.2 mL of sterile saline (0.9% NaCl) with fluorescein in the contralateral eye (control). Group 2 (n = 12) received 0.2 mL of 0.4% oxybuprocaine hydrochloride with fluorescein ophthalmic solution in one eye and 0.2 mL of 0.5% proparacaine hydrochloride with fluorescein in the contralateral eye (control). In each group, corneal touch threshold was determined by Cochet-Bonnet aesthesiometer for both eyes immediately prior to topical administration of solutions, at 1 min and 5 min after administration of topical solutions and every 5 min thereafter for a total of 75 min. Significant corneal anaesthesia was noted immediately following topical application of both oxybuprocaine and proparacaine as compared with controls, with maximal corneal anaesthesia noted 1 min after administration. Both oxybuprocaine and proparacaine produced significant corneal anaesthesia for the duration of the 75-min study. Neither oxybuprocaine hydrochloride nor proparacaine hydrochloride treatment resulted in visible adverse effects. There are limited data available demonstrating the efficacy and duration of corneal anaesthetic agents in cattle. Both oxybuprocaine hydrochloride and proparacaine hydrochloride should be considered practical options for providing corneal anaesthesia in cattle in a clinical setting. © 2016 Australian Veterinary Association.

Comparison of Wide-Field Fluorescein Angiography and Nine-Field Montage Angiography in Uveitis

PubMed Central

Nicholson, Benjamin P.; Nigam, Divya; Miller, Darby; Agrón, Elvira; Dalal, Monica; Jacobs-El, Naima; Lima, Breno da Rocha; Cunningham, Denise; Nussenblatt, Robert; Sen, H. Nida

2014-01-01

Purpose To qualitatively and quantitatively compare Optos© fundus camera fluorescein angiographic images of retinal vascular leakage with 9-field montage Topcon© fluorescein angiography (FA) images in patients with uveitis. We hypothesized that Optos images reveal more leakage in uveitis patients. Design Retrospective, observational case series. Methods Images of all uveitis patients imaged with same-sitting Optos FA and 9-field montage FA during a 9 month period at a single institution (52 eyes of 31 patients) were graded for the total area of retinal vascular leakage. The main outcome measure was area of fluorescein leakage. Results The area of apparent FA leakage was greater in Optos images than in 9-field montage images (median 22.5 mm2 vs. 4.8 mm2, P<0.0001). Twenty-two of 49 (45%) eyes with gradable photos had at least 25% more leakage on the Optos image than on the montage image. Two (4.1%) had at least 25% less leakage on Optos, and 25 (51%) were similar between the two modalities. Two eyes had no apparent retinal vascular leakage on 9-field montage but were found to have apparent leakage on Optos images. Twenty-three of the 49 eyes had posterior pole leakage, and of these 17 (73.9%) showed more posterior pole leakage on the Optos image. A single 200 degree Optos FA image captured a mean 1.50x the area captured by montage photography. Conclusion More retinal vascular pathology, both in the periphery and the posterior pole, is seen with Optos FA in uveitis patients when compared with 9-field montage. The clinical implications of Optos FA findings have yet to be determined. PMID:24321475

Intravitreal Stanniocalcin-1 Enhances New Blood Vessel Growth in a Rat Model of Laser-Induced Choroidal Neovascularization

PubMed Central

Zhao, Min; Xie, Wankun; Tsai, Shu-Huai; Hein, Travis W.; Rocke, Brent A.; Kuo, Lih; Rosa, Robert H.

2018-01-01

Purpose The purpose of this study was to investigate the impact of stanniocalcin-1 (STC-1), a photoreceptor-protective glycoprotein, on the development of choroidal neovascularization (CNV) in relation to VEGF and its main receptor (VEGFR2) expression after laser injury. Methods In rats, CNV was induced by laser photocoagulation in both eyes, followed by intravitreal injection of STC-1 in the right eye and vehicle or denatured STC-1 injection in the left eye as control. Two weeks after laser injury, fundus autofluorescence (FAF) imaging and fundus fluorescein angiography (FFA) were performed. Fluorescein leakage from CNV was graded using a defined scale system. The size of CNV was quantified with spectral domain optical coherence tomography (SD-OCT), fluorescein-labeled choroid-sclera flat mounts, and hematoxylin-eosin staining. Protein expressions were evaluated by Western blot. Results Photocoagulation produced a well-circumscribed area of CNV. With STC-1 treatment, CNV lesions assessed by FAF were increased by 50% in both intensity and area. The CNV lesions were also increased with SD-OCT, flat-mount, and histologic analyses. FFA disclosed enhanced fluorescein leakage in CNV lesions in STC-1 treated eyes. The STC-1 protein was detected in the choroidal tissue and its level was increased with CNV lesions in correlation with VEGF and VEGFR2 expressions. Intravitreal administration of STC-1 significantly increased choroidal expression of both VEGF and VEGFR2 proteins. Conclusions Chorodial tissue expresses STC-1, which seemingly acts as a stress response protein by enhancing pathological new blood vessel growth in laser-induced CNV. It is likely that STC-1 promotes CNV development via VEGF signaling. PMID:29490350

Assessment of murine bladder permeability with fluorescein: validation with cyclophosphamide and protamine.

PubMed

Eichel, L; Scheidweiler, K; Kost, J; Shojaie, J; Schwarz, E; Messing, E; Wood, R

2001-07-01

Bladder hyperpermeability should result in elevated blood levels of intravesically administered agents. Reabsorption from a hyperpermeable bladder should result in prolonged urinary excretion of an agent after parenteral administration. To test these hypotheses, urinary clearance and plasma levels of sodium fluorescein (NaF) were measured in mice before and during cyclophosphamide (CYP) and protamine-induced hemorrhagic cystitis. To measure the plasma uptake of NaF from the bladder, 10 mg/mL NaF was instilled, either by catheter or retrograde urethral infusion, 15 minutes before retro-orbital or ventricular sampling. The plasma levels were measured 24 hours and 14 days after exposure to CYP 300 mg/kg or 15 minutes after instillation of protamine 10 mg/mL. Hourly urine concentrations were measured immediately after intraperitoneal administration of 10 mg/kg NaF. Pretreatment samples were compared with those obtained 24 hours after intraperitoneal administration of 300 mg/kg CYP. Urinary NaF excretion was delayed in CYP-exposed mice. A bi-exponential model provided an appropriate fit of the data, both before and after CYP administration. The plasma levels of NaF were significantly elevated at 24 hours and 14 days after CYP exposure when sampled by ventricular nick or retro-orbitally. The median concentration of fluorescein in the protamine-treated mice was significantly higher than in the control mice. Fluorescein can be used to measure alterations in bladder permeability after bladder mucosal injury in mice. Urinary excretion of NaF is a bi-exponential process that is delayed after bladder mucosal injury, presumably because of increased mucosal permeability and resorption from the urine into the bloodstream.

Efficacy and safety of retinol palmitate ophthalmic solution in the treatment of dry eye: a Japanese Phase II clinical trial.

PubMed

Toshida, Hiroshi; Funaki, Toshinari; Ono, Koichi; Tabuchi, Nobuhito; Watanabe, Sota; Seki, Tamotsu; Otake, Hiroshi; Kato, Takuji; Ebihara, Nobuyuki; Murakami, Akira

2017-01-01

The purpose of this study was to investigate the efficacy and safety of the administration of retinol palmitate (VApal) ophthalmic solution (500 IU/mL) for the treatment of patients with dry eye. This study included 66 patients with dry eye. After a 2-week washout period, patients were randomized (1:1) into either a VApal ophthalmic solution or a placebo group, and a single drop of either solution was administered six times daily for 4 weeks. Efficacy measures were 12 subjective symptoms, rose bengal (RB) and fluorescein staining scores, tear film breakup time, and tear secretion. Safety measures included clinical blood and urine analyses and adverse event recordings. In comparisons of the two groups, the mean change in RB staining score from baseline was significantly lower in the VApal group at 2 and 4 weeks ( P <0.05 and P <0.01, respectively). Furthermore, the fluorescein clearance rate (fluorescein staining score) was significantly higher in the VApal group at 4 weeks ( P <0.05). The VApal group showed a significant improvement in blurred vision at 1 and 2 weeks ( P <0.01 and P <0.05, respectively), and the mean change in the total score for subjective symptoms from baseline was significantly lower in the VApal group at 1 week ( P <0.05). In before- and after-intervention comparisons, the fluorescein and RB staining scores showed improvement in both groups. Improvement was noted for 11 subjective symptoms in the VApal group and for seven symptoms in the placebo group. No significant differences in adverse events and reactions were found between the groups. VApal ophthalmic solution (500 IU/mL) is safe and effective for the treatment of patients with dry eye.

Enhancement of Skin Permeation and Skin Immunization of Ovalbumin Antigen via Microneedles.

PubMed

Pamornpathomkul, Boonnada; Rojanarata, Theerasak; Opanasopit, Praneet; Ngawhirunpat, Tanasait

2017-10-01

The purpose of this study was to evaluate the use of different types of microneedles and doses of ovalbumin antigen for in vitro skin permeation and in vivo immunization. In vitro skin permeation experiments and confocal laser scanning microscopy revealed that hollow microneedles had a superior enhancing effect on skin permeation compared with a solid microneedle patch and untreated skin by efficiently delivering ovalbumin-fluorescein conjugate into the deep skin layers. The flux and cumulative amount of ovalbumin-fluorescein conjugate at 8 h after administering with various conditions could be ranked as follows: hollow MN; high dose > medium dose > low dose > MN patch; high dose > medium dose > low dose > untreated skin; high dose > medium dose > low dose > without ovalbumin-fluorescein conjugate. As the dose of ovalbumin-fluorescein conjugate was increased to 500 μg, the antigen accumulated in the skin to a greater extent, as evidenced by the increasing green fluorescence intensity. When the hollow microneedle was used for the delivery of ovalbumin into the skin of mice, it was capable of inducing a stronger immunoglobulin G immune response than conventional subcutaneous injection at the same antigen dose. Immunoglobulin G levels in the hollow MN group were 5.7, 11.6, and 13.3 times higher than those of the subcutaneous injection group for low, medium, and high doses, respectively. Furthermore, the mice immunized using the hollow microneedle showed no signs of skin infection or pinpoint bleeding. The results suggest that the hollow MN is an efficient device for delivering the optimal dose of antigen via the skin for successful immunization.

Interaction and photodegradation characteristics of fluorescein dye in presence of ZnO nanoparticles.

PubMed

Bardhan, Munmun; Mandal, Gopa; Ganguly, Tapan

2011-04-01

The interaction between xanthene dye Fluorescein (Fl) and zinc oxide (ZnO) nanoparticles is investigated under physiological conditions. From the analysis of the steady state and time resolved spectroscopic studies in aqueous solution static mode is found to be responsible in the mechanism of fluorescence quenching of the dye Fl in presence of ZnO. ZnO nanoparticles are used as photocatalyst in order to degrade Fl dye. At pH 7, a maximum degradation efficiency of 44.4% of the dye has been achieved in presence of ZnO as a nanophotocatalyst and the photodegradation follows second-order kinetics.

Injection by hydrostatic pressure in conjunction with electrokinetic force on a microfluidic chip.

PubMed

Gai, Hongwei; Yu, Linfen; Dai, Zhongpeng; Ma, Yinfa; Lin, Bingcheng

2004-06-01

A simple method was developed for injecting a sample on a cross-form microfluidic chip by means of hydrostatic pressure combined with electrokinetic forces. The hydrostatic pressure was generated simply by adjusting the liquid level in different reservoirs without any additional driven equipment such as a pump. Two dispensing strategies using a floating injection and a gated injection, coupled with hydrostatic pressure loading, were tested. The fluorescence observation verified the feasibility of hydrostatic pressure loading in the separation of a mixture of fluorescein sodium salt and fluorescein isothiocyanate. This method was proved to be effective in leading cells to a separation channel for single cell analysis.

Hypersensitivity to contrast media and dyes.

PubMed

Brockow, Knut; Sánchez-Borges, Mario

2014-08-01

This article updates current knowledge on hypersensitivity reactions to diagnostic contrast media and dyes. After application of a single iodinated radiocontrast medium (RCM), gadolinium-based contrast medium, fluorescein, or a blue dye, a hypersensitivity reaction is not a common finding; however, because of the high and still increasing frequency of those procedures, patients who have experienced severe reactions are nevertheless frequently encountered in allergy departments. Evidence on allergologic testing and management is best for iodinated RCM, limited for blue dyes, and insufficient for fluorescein. Skin tests can be helpful in the diagnosis of patients with hypersensitivity reactions to these compounds. Copyright © 2014 Elsevier Inc. All rights reserved.

[Preparation and vitality detection of protoplast in Salvia miltiorrhiza Bunge].

PubMed

Zhu, Nan; Liu, Jun; Zhang, Xinyu; Dong, Juan'e

2014-10-01

We prepared protoplasts from Salvia miltiorrhiza Bunge suspension culture cells. Then, the protoplasts' vitality and functions were tested by fluorescein diacetate staining method and Fluo-3/AM flourescent probe. The optimal condition of protoplast isolation was Cellulase R-10 1.5%, Pectinase Y-23 0.3%, Macerozyme R-10 0.5%, 40 r/min 12 h, 600 r/min 5 min, and the protoplasts yield was 1.1x10(6) cells/g FW, the vitality was more than 95% by using fluorescein diacetate staining method. It has been confirmed that calcium fluorescent probe Fluo-3/AM can be successfully loaded into protoplasts.

Ultra-widefield fluorescein angiography reveals retinal phlebitis in Susac's syndrome.

PubMed

Klufas, Michael A; Dinkin, Marc J; Bhaleeya, Swetangi D; Chapman, Kristin O; Riley, Claire S; Kiss, Szilárd

2014-01-01

A 23-year-old woman with history of headaches and auditory changes presented with acute-onset visual field loss in the right eye. The combination of multiple retinal branch artery occlusions of the right eye on funduscopic examination, characteristic white matter lesions in the corpus callosum on magnetic resonance imaging, and hearing loss on audiometric testing led to a diagnosis of Susac's syndrome. Ultra-widefield fluorescein angiography revealed involvement of the retinal veins, which has not been previously reported with this condition. Additionally, ultra-widefield indocyanine green angiography demonstrated changes in the choroidal circulation, which are controversial in this syndrome. Copyright 2014, SLACK Incorporated.

The Modification of Fluorescein Angiography and Its Applications in Age-Related Macular Degeneration and Polypoidal Choroidal Vasculopathy.

PubMed

Peng, Qing; Chen, Yutong; Hua, Rui

2018-06-07

To establish a novel retinal angiography method, red-free angiography (RFA), to investigate retinal changes in age-related macular degeneration (AMD) and polypoidal choroidal vasculopathy (PCV). Following the venous phase of fundus fluorescein angiography (FFA), the detection mode was switched to red-free reflectance to acquire RFA images using the same parameters. RFA showed subretinal fluid, polyps, and outer retinal tubulation, with a higher definition than the FFA and red-free reflectance results. The absorption coefficients in RFA provided more detailed images for AMD and PCV diagnosis. RFA is therefore a promising approach to supplement FFA. © 2018 S. Karger AG, Basel.

Effect of Sodium Fluorescein and Plating Medium on Recovery of Irradiated Escherichia coli and Serratia marcescens from Aerosols

PubMed Central

Dorsey, Emerson L.; Berendt, Richard F.; Neff, Everett L.

1970-01-01

Irradiation of aerosols of either Escherichia coli or Serratia marcescens with simulated solar (xenon) radiation caused a significant decrease in viability. When sodium fluorescein was employed to determine the physical loss of organisms from the aerosol, an additional adverse effect upon survival was noted. The decay curves indicated that at least two mechanisms of inactivation were operative, one due to aerosolization, the other to irradiation. After collection from aerosols, both species of microorganisms grew better on blood agar base than on Casitone agar, but this finding did not appear to be related to the effect of irradiation. PMID:4922085

Neovascular glaucoma in a patient with X-linked juvenile retinoschisis.

PubMed

Zuo, Chengguo; Chen, Changzheng; Xing, Yiqiao; Du, Lei

2005-09-01

To report the rubeosis iridis and neovascular glaucoma findings in one patient of X-linked juvenile retinoschisis(XLRS). Color fundus photography, fluorescein angiography (FFA), OCT and B-scan were performed in a patient with X-linked juvenile retinoschisis complicated with neovascular glaucoma. Color fundus photography, fluorescein angiography (FFA), OCT and B-scan unveiled a rare condition of XLRS complicated with neovascular glaucoma. XLRS may complicate with neovascular glaucoma. It is necessary to test OCT, FFA, ERG and carefully examine the fundus of the follow eye when it comes to uncertain neovascular glaucoma of youth and child. And only in this way, can we exclude XLRS.

Effects of chronic kidney disease on liver transport: quantitative intravital microscopy of fluorescein transport in the rat liver.

PubMed

Ryan, Jennifer C; Dunn, Kenneth W; Decker, Brian S

2014-12-15

Clinical studies indicate that hepatic drug transport may be altered in chronic kidney disease (CKD). Uremic solutes associated with CKD have been found to alter the expression and/or activity of hepatocyte transporters in experimental animals and in cultured cells. However, given the complexity and adaptability of hepatic transport, it is not clear whether these changes translate into significant alterations in hepatic transport in vivo. To directly measure the effect of CKD on hepatocyte transport in vivo, we conducted quantitative intravital microscopy of transport of the fluorescent organic anion fluorescein in the livers of rats following 5/6th nephrectomy, an established model of CKD. Our quantitative analysis of fluorescein transport showed that the rate of hepatocyte uptake was reduced by ∼20% in 5/6th nephrectomized rats, consistent with previous observations of Oatp downregulation. However, the overall rate of transport into bile canaliculi was unaffected, suggesting compensatory changes in Mrp2-mediated secretion. Our study suggests that uremia resulting from 5/6th nephrectomy does not significantly impact the overall hepatic clearance of an Oatp substrate. Copyright © 2014 the American Physiological Society.

The tear turnover and tear clearance tests - a review.

PubMed

Garaszczuk, Izabela K; Montes Mico, Robert; Iskander, D Robert; Expósito, Alejandro Cerviño

2018-03-01

The aim is to provide a summary of methods available for the assessment of tear turnover and tear clearance rates. The review defines tear clearance and tear turnover and describes their implication for ocular surface health. Additionally, it describes main types of techniques for measuring tear turnover, including fluorescein tear clearance tests, techniques utilizing electromagnetic spectrum and tracer molecule and novel experimental techniques utilizing optical coherence tomography and fluorescein profilometry. Areas covered: Internet databases (PubMed, Science Direct, Google Scholar) and most frequently cited references were used as a principal resource of information on tear turnover rate and tear clearance rate, presenting methodologies and equipment, as well as their definition and implications for the anterior eye surface health and function. Keywords used for data-search were as follows: tear turnover, tear clearance, fluorescein clearance, scintigraphy, fluorophotometry, tear flow, drainage, tear meniscus dynamics, Krehbiel flow and lacrimal functional unit. Expert commentary: After decades, the topic of tear turnover assessment has been reintroduced. Recently, new techniques have been developed to propose less invasive, less time consuming and simpler methodologies for the assessment of tear dynamics that have the potential to be utilized in clinical practice.

Dual fluorescence/contactless conductivity detection for microfluidic chip.

PubMed

Liu, Cui; Mo, Yun-yan; Chen, Zuan-guang; Li, Xiang; Li, Ou-lian; Zhou, Xie

2008-07-28

A new dual detection system for microchip is reported. Both fluorescence detector (FD) and contactless conductivity detector (CCD) were combined together and integrated on a microfluidic chip. They shared a common detection position and responded simultaneously. A blue light-emitting diode was used as excitation source and a small planar photodiode was used to collect the emitted fluorescence in fluorescence detection, which made the device more compact and portable. The coupling of the fluorescence and contactless conductivity modes at the same position of a single separation channel enhanced the detection characterization of sample and offered simultaneous detection information of both fluorescent and charged specimen. The detection conditions of the system were optimized. K(+), Na(+), fluorescein sodium, fluorescein isothiocyanate (FITC) and FITC-labeled amino acids were used to evaluate the performance of the dual detection system. The limits of detection (LOD) of FD for fluorescein Na(+), FITC, FITC-labeled arginine (Arg), glycine (Gly) and phenylalanine (Phe) were 0.02micromolL(-1), 0.05micromolL(-1), 0.16micromolL(-1), 0.15micromolL(-1), 0.12micromolL(-1) respectively, and the limits of detection (LOD) of CCD achieved 0.58micromolL(-1) and 0.39micromolL(-1) for K(+) and Na(+) respectively.

[Fluorescein angiography and optical coherence tomography findings in central fundus of myopic patients].

PubMed

Avetisov, S E; Budzinskaya, M V; Zhabina, O A; Andreeva, I V; Plyukhova, A A; Kobzova, M V; Musaeva, G M

2015-01-01

Myopia prevalence grows alike in many countries, including Russia, regardless of geographical and population conditions. to assess fundus changes in myopic patients at different ocular axial lengths by means of modern diagnostic tools. The study enrolled 97 patients (194 eyes) aged 45 ± 20.17 years with myopia of different degrees. Besides a standard ophthalmic examination, all patients underwent fundus fluorescein angiography and optical coherence tomography. The occurrence of retinal pigment epithelium (RPE) atrophy (diffuse or focal) has been shown to increase with increasing ocular axial length. Only 27 eyes (28.1%) appeared intact. As myopia progression implies axial growth of the eye, it is associated with a more severe decrease in choroid, RPE, and photoreceptor layer thicknesses: the longer the anterior-posterior axis, the thinner the above mentioned fundus structures. Age-related changes in the fundus are also likely to be more pronounced in longer axes. Myopic traction maculopathy, which in our case appeared the main cause of increased retinal thickness, was diagnosed in 105 eyes, "outer" macular retinoschisis--in 40 eyes. Thus, modern diagnostic tools, such as fluorescein angiography and optical coherence tomography, enable objective assessment of the central fundus.

Successful treatment of toxoplasmosis-associated choroidal neovascular lesions with bevacizumab and antiparasitic therapy.

PubMed

Lin, Chun-Ju; Chen, San-Ni; Hwang, Jiunn-Feng; Hu, Pei-Shin

2011-01-01

To report the effects of oral trimethoprim/sulfamethoxazole and intravitreal bevacizumab injection in the treatment of ocular toxoplasmosis-associated choroidal neovascular lesions (CNV). This was a noncomparative, nonrandomized, consecutive case series. All eyes with ocular toxoplasmosis-associated CNV received one intravitreal bevacizumab injection under the coverage of oral trimethoprim/sulfamethoxazole. The changes in best-corrected visual acuity were recorded. Serial fundus photography, fluorescein angiography, optical coherence tomography, and indocyanine green angiography were performed to measure the treatment efficacy. Three eyes of two patients with a history of ocular toxoplasmosis had active CNV demonstrated by fluorescein angiography and optical coherence tomography. Each was treated with oral trimethoprim/sulfamethoxazole and one intravitreal bevacizumab injection. Best-corrected visual acuity, fundus photographs, fluorescein angiography, optical coherence tomography, and indocyanine green angiography all showed favorable results. No ocular or systemic complications were noted. In all three eyes, the CNV subsided and vision improved. Oral trimethoprim/sulfamethoxazole is an effective and less expensive antibiotic against Toxoplasma gondii. Intravitreal bevacizumab injection appears to be a well-tolerated treatment for toxoplasmosis-associated CNV and has the potential as an adjuvant therapy to improve final vision. More cases and further studies are required.

Agonist-induced conformational changes in the G-protein-coupling domain of the β2 adrenergic receptor

PubMed Central

Ghanouni, Pejman; Steenhuis, Jacqueline J.; Farrens, David L.; Kobilka, Brian K.

2001-01-01

The majority of extracellular physiologic signaling molecules act by stimulating GTP-binding protein (G-protein)-coupled receptors (GPCRs). To monitor directly the formation of the active state of a prototypical GPCR, we devised a method to site specifically attach fluorescein to an endogenous cysteine (Cys-265) at the cytoplasmic end of transmembrane 6 (TM6) of the β2 adrenergic receptor (β2AR), adjacent to the G-protein-coupling domain. We demonstrate that this tag reports agonist-induced conformational changes in the receptor, with agonists causing a decline in the fluorescence intensity of fluorescein-β2AR that is proportional to the biological efficacy of the agonist. We also find that agonists alter the interaction between the fluorescein at Cys-265 and fluorescence-quenching reagents localized to different molecular environments of the receptor. These observations are consistent with a rotation and/or tilting of TM6 on agonist activation. Our studies, when compared with studies of activation in rhodopsin, indicate a general mechanism for GPCR activation; however, a notable difference is the relatively slow kinetics of the conformational changes in the β2AR, which may reflect the different energetics of activation by diffusible ligands. PMID:11353823

A compactly integrated laser-induced fluorescence detector for microchip electrophoresis.

PubMed

Li, Hai-Fang; Lin, Jin-Ming; Su, Rong-Guo; Uchiyama, Katsumi; Hobo, Toshiyuki

2004-06-01

A simple and easy-to-use integrated laser-induced fluorescence detector for microchip electrophoresis was constructed and evaluated. The fluid channels and optical fiber channels in the glass microchip were fabricated using standard photolithographic techniques and wet chemical etching. A 473 nm diode-pumped laser was used as the excitation source, and the collimation and collection optics and mirrors were discarded by using a multimode optical fiber to couple the excitation light straight into the microchannel and placing the microchip directly on the top of the photomultiplier tube. A combination of filter systems was incorporated into a poly(dimethylsiloxane) layer, which was reversibly sealed to the bottom of the microchip to eliminate the scattering excitation light reaching to the photomultiplier tube. Fluorescein/calcein samples were taken as model analytes to evaluate the performance with respect to design factors. The detection limits were 0.05 microM for fluorescein and 0.18 microM for calcein, respectively. The suitability of this simple detector for fluorescence detection was demonstrated by baseline separation of fluorescein isothiocyanate (FITC)-labeled arginine, phenylalanine, and glycine and FITC within 30 s at separation length of 3.8 cm and electrical field strength of 600 V/cm.

Effect of antioxidant oxidation potential in the oxygen radical absorption capacity (ORAC) assay.

PubMed

Bisby, Roger H; Brooke, Rachel; Navaratnam, Suppiah

2008-06-01

The "oxygen radical absorption capacity" (ORAC) assay (Ou, B., Hampsch-Woodill, M., Prior, R.L. (2001). Development and validation of an improved oxygen radical absorbance capacity assay using fluorescein as the fluorescent probe. Journal of Agricultural and Food Chemistry 49, 4619-4626) is widely employed to determine antioxidant content of foods and uses fluorescein as a probe for oxidation by peroxyl radicals. Kinetic modeling of the ORAC assay suggests that the lag phase for loss of fluorescence results from equilibrium between antioxidant and fluorescein radicals and the value of the equilibrium constant determines the shape of the lag phase. For an efficient antioxidant this constitutes a "repair" reaction for fluoresceinyl radicals and produces a well defined lag phase. The lag phase becomes less marked with increasing oxidation potential of the antioxidant. Pulse radiolysis confirms that fluoresceinyl radicals are rapidly (k∼10(9)dm(3)mol(-1)s(-1)) reduced by Trolox C, a water soluble vitamin E analogue. ORAC assays of phenols with varying oxidation potentials suggest that it might be employed to obtain an estimate of the redox potential of antioxidants within food materials. Copyright © 2007 Elsevier Ltd. All rights reserved.

Determination of total antioxidant capacity by oxygen radical absorbance capacity (ORAC) using fluorescein as the fluorescence probe: First Action 2012.23.

PubMed

Ou, Boxin; Chang, Tony; Huang, Dejian; Prior, Ronald L

2013-01-01

An improved method for the measurement of oxygen radical absorbance capacity (ORAC) was developed and validated using fluorescein (3',6'-dihydroxyspiro[isobenzofuran-1[3H], 9'[9H]-xanthen]-3-one) as a new fluorescence probe (ORAC(FL)). Randomly methylated beta-cyclodextrin (RMCD) was introduced as the water-solubility enhancer for lipophilic antioxidants. 7% RMCD (w/v) in 50% acetone-H2O mixture sufficiently solubilized vitamin E compounds and other lipophilic phenolic antioxidants in 75 mM phosphate buffer (pH 7.4). Results indicated that fluorescein shows excellent photostability under the plate reader conditions. This ORAC(FL) was validated through linearity, precision, accuracy, and ruggedness. The validation results demonstrated that the ORACFL assay is reliable and robust. The mean of intraday and interday CVs were <15%; for hydrophilic ORAC, LOD and LOQ are 5 and 6.25 microM, respectively; for lipophilic ORAC, LOD and LOQ are 6.25 and 12.5 microM, respectively. It is concluded that unlike other popular methods, the ORAC(FL) assay provides a direct measure of total antioxidant capacity against the peroxyl radicals.

Cochlear microdialysis for quantification of dexamethasone and fluorescein entry into scala tympani during round window administration.

PubMed

Hahn, Hartmut; Kammerer, Bernd; DiMauro, Andre; Salt, Alec N; Plontke, Stefan K

2006-02-01

Before new drugs for the treatment of inner ear disorders can be studied in controlled clinical trials, it is important that their pharmacokinetics be established in inner ear fluids. Microdialysis allows drug levels to be measured in perilymph without the volume disturbances and potential cerebrospinal fluid contamination associated with fluid sampling. The aims of this study were to show: (i) that despite low recovery rates from miniature dialysis probes, significant amounts of drug are removed from small fluid compartments, (ii) that dialysis sampling artifacts can be accounted for using computer simulations and (iii) that microdialysis allows quantification of the entry rates through the round window membrane (RWM) into scala tympani (ST). Initial experiments used microdialysis probes in small compartments in vitro containing sodium fluorescein. Stable concentrations were observed in large compartments (1000 microl) but significant concentration declines were observed in smaller compartments (100, 10 and 5.6 microl) comparable to the size of the inner ear. Computer simulations of these experiments closely approximated the experimental data. In in vivo experiments, sodium fluorescein 10 mg/ml and dexamethasone-dihydrogen-phosphate disodium salt 8 mg/ml were simultaneously applied to the RWM of guinea pigs. Perilymph concentration in the basal turn of ST was monitored using microdialysis. The fluorescein concentration reached after 200 min application (585+/-527 microg/ml) was approximately twice that of dexamethasone phosphate (291+/-369 microg/ml). Substantial variation in concentrations was found between animals by approximately a factor of 34 for fluorescein and at least 41 for dexamethasone phosphate. This is, to a large extent, thought to be the result of the RWM permeability varying in different animals. It was not caused by substance analysis variations, because two different analytic methods were used and the concentration ratio between the two substances remained nearly constant across the experiments and because differences were apparent for the repeated samples obtained in each animal. Interpretation of the results using computer simulations allowed RWM permeability to be quantified. It also demonstrated, however, that cochlear clearance values could not be reliably obtained with microdialysis because of the significant contribution of dialysis to clearance. The observed interanimal variation, e.g., in RWM permeability, is likely to be clinically relevant to the local application of drugs in patients.

Cochlear Microdialysis for Quantification of Dexamethasone and Fluorescein Entry into Scala Tympani During Round Window Administration

PubMed Central

Hahn, Hartmut; Kammerer, Bernd; DiMauro, Andre; Salt, Alec N.; Plontke, Stefan K.

2006-01-01

Before new drugs for the treatment of inner ear disorders can be studied in controlled clinical trials, it is important that their pharmacokinetics be established in inner ear fluids. Microdialysis allows drug levels to be measured in perilymph without the volume disturbances and potential cerebrospinal fluid contamination associated with fluid sampling. The aims of this study were to show: (i) that despite low recovery rates from miniature dialysis probes, significant amounts of drug are removed from small fluid compartments, (ii) that dialysis sampling artifacts can be accounted for using computer simulations and (iii) that microdialysis allows quantification of the entry rates through the round window membrane (RWM) into scala tympani (ST). Initial experiments used microdialysis probes in small compartments in vitro containing sodium fluorescein. Stable concentrations were observed in large compartments (1000 μl) but significant concentration declines were observed in smaller compartments (100, 10 and 5.6 μl) comparable to the size of the inner ear. Computer simulations of these experiments closely approximated the experimental data. In in vivo experiments, sodium fluorescein 10 mg/ml and dexamethasone-dihydrogen-phosphate disodium salt 8 mg/ml were simultaneously applied to the RWM of guinea pigs. Perilymph concentration in the basal turn of ST was monitored using microdialysis. The fluorescein concentration reached after 200 min application (585 ± 527 μg/ml) was approximately twice that of dexamethasone phosphate (291 ± 369 μg/ml). Substantial variation in concentrations was found between animals by approximately a factor of 34 for fluorescein and at least 41 for dexamethasone phosphate. This is, to a large extent, thought to be the result of the RWM permeability varying in different animals. It was not caused by substance analysis variations, because two different analytic methods were used and the concentration ratio between the two substances remained nearly constant across the experiments and because differences were apparent for the repeated samples obtained in each animal. Interpretation of the results using computer simulations allowed RWM permeability to be quantified. It also demonstrated, however, that cochlear clearance values could not be reliably obtained with microdialysis because of the significant contribution of dialysis to clearance. The observed interanimal variation, e.g., in RWM permeability, is likely to be clinically relevant to the local application of drugs in patients. PMID:16442251

Optic disk findings in hypervitaminosis A.

PubMed

Marcus, D F; Turgeon, P; Aaberg, T M; Wiznia, R A; Wetzig, P C; Bovino, J A

1985-07-01

Three cases of papilledema secondary to chronic excessive vitamin A intake are presented, and the optic disk changes are documented with intravenous fluorescein angiography. Two of the three patients reported in this study were symptomatic with blurred vision and systemic complaints. The symptoms of blurred vision and systemic complaints disappeared within one week, and papilledema resolved over several months after discontinuance of vitamin A. The fluorescein angiographic changes observed in the optic disk of patients with hypervitaminosis A are similar to those associated with other known causes of papilledema. Since vitamin A is a nonprescription drug, and its indiscriminate use is potentially great, any history of vitamin ingestion should be elicited during the evaluation of papilledema.

Ultra-wide-field and autofluorescence imaging of choroidal dystrophies.

PubMed

Yuan, Alex; Kaines, Andrew; Jain, Atul; Reddy, Shantan; Schwartz, Steven D; Sarraf, David

2010-10-28

The authors retrospectively identified 2 cases of gyrate atrophy, 3 cases of choroideremia, and 1 case of the carrier state of choroideremia who underwent ultra-wide-field fundus photography and fluorescein angiography. The findings were studied and compared to standard fundus photography and fluorescein angiography. Gyrate atrophy demonstrated a diffuse confluent extent of chorioretinal atrophy extending from the anterior to the posterior pole to the periphery. Choroideremia demonstrated a patchy irregular pattern of chorioretinal atrophy extending from the posterior pole to the periphery. Peripheral reticular degeneration without chorioretinal atrophy was appreciated in the carrier state. Ultra-wide-field imaging of these choroidal dystrophies demonstrated distinctive patterns that may aid in their identification and diagnosis. Copyright 2010, SLACK Incorporated.

Study on the ternary mixed ligand complex of palladium(II)-aminophylline-fluorescein sodium by resonance Rayleigh scattering, second-order scattering and frequency doubling scattering spectrum and its analytical application.

PubMed

Chen, Peili; Liu, Shaopu; Liu, Zhongfang; Hu, Xiaoli

2011-01-01

The interaction between palladium(II)-aminophylline and fluorescein sodium was investigated by resonance Rayleigh scattering, second-order scattering and frequency doubling scattering spectrum. In pH 4.4 Britton-Robinson (BR) buffer medium, aminophylline (Ami) reacted with palladium(II) to form chelate cation([Pd(Ami)]2+), which further reacted with fluorescein sodium (FS) to form ternary mixed ligand complex [Pd(Ami)(FS)2]. As a result, resonance Rayleigh scattering (RRS), second-order scattering (SOS) and frequency doubling scattering spectrum (FDS) were enhanced. The maximum scattering wavelengths of [Pd(Ami)(FS)2] were located at 300 nm (RRS), 650 nm (SOS) and 304 nm (FDS). The scattering intensities were proportional to the Ami concentration in a certain range and the detection limits were 7.3 ng mL(-1) (RRS), 32.9 ng mL(-1) (SOS) and 79.1 ng mL(-1) (FDS), respectively. Based on it, the new simple, rapid, and sensitive scattering methods have been proposed to determine Ami in urine and serum samples. Moreover, the formation mechanism of [Pd(Ami)(FS)2] and the reasons for enhancement of RRS were fully discussed. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Polarization of fluorescein fluorescence in single cells.

PubMed

Udkoff, R; Norman, A

1979-01-01

Measurement of fluorescence polarization (P) gives information about the immediate environment of the fluorescent molecule. We used a flow polarimeter to investigate the factors influencing P of fluorescein in mammalian cells to determine whether such measurements are useful for characterizing heterogeneous cell populations. Fluorescein was introduced into cells by incubation with FDA. Measurements of the intensity of fluorescence (TI) and polarization (P) revealed an unexpected dependence: P decreased with increasing intensity of fluorescence. This may be accounted for by the classical model of the binding of small molecules to protein in which P is dependent on the ratio bound to unbound molecules. We have been able to estimate the quenching due to binding and construct a Scatchard plot. We estimated a wavelength shift from in vitro data consistent with the dependence of P on wavelength seen in our cell work. Generally, the distributions of P are symmetrical. Photon statistics broadens the P distribution of dim cells. However, structure does develop in the P distribution when the cells are deprived of calcium or incubated in the cold. This appears as a shoulder on the P distribution or resolves into two peaks. Calcium deprivation may differentially affect a subpopulation of cells whose significance remains to be explored in various cell types.

Crystallization of dicalcium phosphate dihydrate with presence of glutamic acid and arginine at 37 °C.

PubMed

Li, Chengfeng; Ge, Xiaolu; Li, Guochang; Bai, Jiahai; Ding, Rui

2014-08-01

The formations of non-metabolic stones, bones and teeth were seriously related to the morphology, size and surface reactivity of dicalcium phosphate dihydrate (DCPD). Herein, a facile biomimetic mineralization method with presence of glutamic acid and arginine was employed to fabricate DCPD with well-defined morphology and adjustable crystallite size. In reaction solution containing more arginine, crystallization of DCPD occurred with faster rate of nucleation and higher density of stacked layers due to the generation of more OH(-) ions after hydrolysis of arginine at 37 °C. With addition of fluorescein or acetone, the consumption of OH(-) ions or desolvation reaction of Ca(2+) ions was modulated, which resulted in the fabrication of DCPD with adjustable crystallite sizes and densities of stacked layers. In comparison with fluorescein-loading DCPD, dicalcium phosphate anhydrate was prepared with enhanced photoluminescence properties due to the reduction of self-quenching effect and regular arrangement of encapsulated fluorescein molecules. With addition of more acetone, DCPD was prepared with smaller crystallite size via antisolvent crystallization. The simulated process with addition of amino acids under 37 °C would shed light on the dynamic process of biomineralization for calcium phosphate compounds. Copyright © 2014 Elsevier B.V. All rights reserved.

The Efficacy and Safety of Topical Rocuronium Bromide to Induce Bilateral Mydriasis in Hispaniolan Amazon Parrots ( Amazona ventralis ).

PubMed

Baine, Katherine; Hendrix, Diane V H; Kuhn, Sonia E; Souza, Marcy J; Jones, Michael P

2016-03-01

The efficacy and safety of topically applied rocuronium in Hispaniolan Amazon parrots ( Amazona ventralis ) was assessed in a group of 10 adult birds. A complete ophthalmic examination (including Schirmer tear test, ocular reflexes, applanation tonometry, fluorescein staining, and slit-lamp biomicroscopy) was performed, and rocuronium bromide (0.15 mg in both eyes) was administered. Pupillary light reflex (PLR) and pupillary diameter were recorded in a darkened room at the following time points: 0, 10, 20, 30, 40, 50, 60, 80, 100, 120, 140, 160, 180, 200, 220, 240, 300, and 360 minutes, and 24 hours. Fluorescein staining in both eyes was performed at 24 hours. By 10 minutes, PLR was absent in all birds (at 5 minutes, 8 birds; at 10 minutes, remaining 2 birds). Pupil diameter differed significantly from baseline at all time points. Additionally, PLR was decreased in 7/10 birds at 360 minutes and normal in all birds at 24 hours. Superficial corneal ulceration was observed at 24 hours in the left eye of 2/10 of the birds after fluorescein stain application. This study demonstrated that rocuronium bromide was an effective mydriatic agent in Hispaniolan Amazon parrots with rapid onset and prolonged duration of action.

Specialized moisture retention eyewear for evaporative dry eye.

PubMed

Waduthantri, Samanthila; Tan, Chien Hua; Fong, Yee Wei; Tong, Louis

2015-05-01

To evaluate the suitablity of commercially available moisture retention eyewear for treating evaporative dry eye. Eleven patients with evaporative dry eyes were prescibed moisture retention eyewear for 3 months in addition to regular lubricant eye drops. Frequency and severity of dry eye symptoms, corneal fluorescein staining and tear break up time (TBUT) were evaluated at baseline and 3-month post-treatment. Main outcome measure was global symptom score (based on severity and frequency of dry eye symptoms on a visual analog scale) and secondary outcomes were changes in sectoral corneal fluorescein staining and tear break up time (TBUT) from pre-treatment level. There was a significant improvement in dry eye symptoms after using moisture retention eyewear for 3 months (p < 0.05). Corneal fluorescein staining in all five zones of the cornea in both eyes improved significantly (p < 0.05). There was no significant improvement in TBUT. Patients used ocular lubricants less frequently (p < 0.05) compared to the commencement of the study. Patients found moisture retention eyewear to be useful in relieving dry eye symptoms in windy, air-conditioned environments or when doing vision-related daily tasks. This study shows that moisture retention eyewear might be a valuable adjunct in management of evaporative dry eye and this new design of commercially available eyewear could have a good acceptability rate.

Liposomes Size Engineering by Combination of Ethanol Injection and Supercritical Processing.

PubMed

Santo, Islane Espirito; Campardelli, Roberta; Albuquerque, Elaine Cabral; Vieira De Melo, Silvio A B; Reverchon, Ernesto; Della Porta, Giovanna

2015-11-01

Supercritical fluid extraction using a high-pressure packed tower is proposed not only to remove the ethanol residue from liposome suspensions but also to affect their size and distribution leading the production of nanosomes. Different operating pressures, temperatures, and gas to liquid ratios were explored and ethanol was successfully extracted up to a value of 400 ppm; liposome size and distribution were also reduced by the supercritical processing preserving their integrity, as confirmed by Z-potential data and Trasmission Electron Microscopy observations. Operating at 120 bar and 38°C, nanosomes with a mean diameter of about 180 ± 40 nm and good storage stability were obtained. The supercritical processing did not interfere on drug encapsulation, and no loss of entrapped drug was observed when the water-soluble fluorescein was loaded as a model compound. Fluorescein encapsulation efficiency was 30% if pure water was used during the supercritical extraction as processing fluid; whereas an encapsulation efficiency of 90% was obtained if the liposome suspension was processed in water/fluorescein solution. The described technology is easy to scale up to an industrial production and merge in one step the solvent extraction, liposome size engineering, and an excellent drug encapsulation in a single operation unit. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

Single Lipid Molecule Dynamics on Supported Lipid Bilayers with Membrane Curvature.

PubMed

Cheney, Philip P; Weisgerber, Alan W; Feuerbach, Alec M; Knowles, Michelle K

2017-03-15

The plasma membrane is a highly compartmentalized, dynamic material and this organization is essential for a wide variety of cellular processes. Nanoscale domains allow proteins to organize for cell signaling, endo- and exocytosis, and other essential processes. Even in the absence of proteins, lipids have the ability to organize into domains as a result of a variety of chemical and physical interactions. One feature of membranes that affects lipid domain formation is membrane curvature. To directly test the role of curvature in lipid sorting, we measured the accumulation of two similar lipids, 1,2-Dihexadecanoyl- sn -glycero-3-phosphoethanolamine (DHPE) and hexadecanoic acid (HDA), using a supported lipid bilayer that was assembled over a nanopatterned surface to obtain regions of membrane curvature. Both lipids studied contain 16 carbon, saturated tails and a head group tag for fluorescence microscopy measurements. The accumulation of lipids at curvatures ranging from 28 nm to 55 nm radii was measured and fluorescein labeled DHPE accumulated more than fluorescein labeled HDA at regions of membrane curvature. We then tested whether single biotinylated DHPE molecules sense curvature using single particle tracking methods. Similar to groups of fluorescein labeled DHPE accumulating at curvature, the dynamics of single molecules of biotinylated DHPE was also affected by membrane curvature and highly confined motion was observed.

Protective effect of melatonin in the diabetic rat retina.

PubMed

Mehrzadi, Saeed; Motevalian, Manijeh; Rezaei Kanavi, Mozhgan; Fatemi, Iman; Ghaznavi, Habib; Shahriari, Mansoor

2018-03-01

Diabetic retinopathy (DR) is one of the most common and serious microvascular complications of diabetes. The aim of this study was to evaluate the effects of melatonin (MEL) on retinal injury in diabetic rats. In this study, 21 rats were randomly divided into three groups: control, diabetic, and diabetic + MEL. Streptozotocin was used to induce diabetes at a dose of 50 mg/kg, i.p., and blood glucose was measured to choose the diabetic rats for the study. MEL (20 mg/kg) was given orally for 7 weeks in diabetic rats starting 1 week after induction of diabetes. After 8 weeks, the groups were compared in terms of mean scores of fluorescein leakage, using fluorescein angiography. Reactive oxygen species (ROS) and malondialdehyde (MDA) levels were estimated in retina using commercially available assays. Structural changes in retinas were evaluated by light microscopy. Results showed that diabetes significantly increased the mean scores of fluorescein leakage, and MDA and ROS levels compared to control group. Treatment of the diabetic rats with MEL for 7 weeks prevented the alterations induced by diabetes in comparison with the diabetic control group.Based on these findings, it can be concluded that MEL might have beneficial effects in prevention of DR. © 2018 Société Française de Pharmacologie et de Thérapeutique.

Research of the fluorescence detection apparatus for nutrients

NASA Astrophysics Data System (ADS)

Wang, Yu; Yan, Huimin; Ni, Xuxiang; Xu, Xiaoyi; Chen, Shibing

2015-10-01

The research of the multifunctional analyzer of Clinical Nutrition, which integrates the absorbance, luminescence, fluorescence and other optical detection methods, can overcome the functional limitations of a single technology on human nutrition analysis, and realize a rapid and accurate analysis of the nutrients. This article focuses on the design of fluorescence detection module that uses a photomultiplier tube(PMT) to detect weak fluorescence, and utilizes the single photon counting method to measure the fluorescence intensity, and then according to the relationship between the fluorescent marker and fluorescence intensity, the concentration of the analyte can be derived. Using fluorescein isothiocyanate(FITC, the most widely used fluorescein currently)to mark antibodies in the experiment, therefore, according to the maximum absorption wavelength and the maximum emission wavelength of the fluorescein isothiocyanate, to select the appropriate filters to set up the optical path. In addition, the fluorescence detection apparatus proposed in this paper uses an aspherical lens with large numerical aperture, in order to improve the capacity of signal acquisition more effectively, and the selective adoption of flexible optical fiber can realize a compact opto-mechanical structure, which is also conducive to the miniaturization of the device. The experimental results show that this apparatus has a high sensitivity, can be used for the detection and analysis of human nutrition.

Labelling of histone H5 and its interaction with DNA. 1. Histone H5 labelling with fluorescein isothiocyanate.

PubMed

Favazza, M; Lerho, M; Houssier, C

1990-06-01

Histone H5 has been labelled with fluorescein isothiocyanate (FITC) with particular attention to the reaction conditions (pH, reaction time and input FITC/H5 molar ratio) and to the complete elimination of non-covalently bound dye. We preferred to use reaction conditions which yielded non-specific uniform labelling rather than specific alpha-NH2 terminal labelling, in order to obtain higher sensitivity in further studies dealing with the detection of perturbation at the binding sites of H5 on DNA. FITC-labelled H5 was further characterized by absorption and circular dichroism spectroscopy, and the fluorescein probe titrated in the 4-8 pH range. The structural integrity of H5 was found to be preserved after labelling. The positive electrostatic potential of the environment in which the FITC probe is embedded in the arginine/lysine-rich tails of H5 is believed to be responsible for the drop of pK of 1 unit found for H5-FITC as compared to free FITC. For the globular part of H5, the pK of covalently-bound FITC was only slightly lowered; this is a consequence of the much lower content in positively-charged amino-acid side chains in this region.

Application of the fiber-optic perfusion fluorometer to absorption and exsorption studies in hairless mouse skin.

PubMed

Shackleford, J M; Yielding, K L

1987-09-01

This study was undertaken to test the fiber-optic perfusion fluorometer as a direct means of evaluating skin absorption and exsorption in hairless mice. Skin-barrier compromise was accomplished in the absorption experiments by application of dimethyl sulfoxide to the skin surface or by partial removal of the stratum corneum with sticky tape. Absorbed fluorescein was measured easily in unanesthetized control (skin-barrier intact) and experimental mice. Unabsorbed chemical did not fluoresce 15 minutes after application, although it was present on the surface of the skin as a dry powder. The time course of fluorescein elimination from the skin was related to a rapid phase (vascular removal) and a slow phase (reservoir entrapment). In the exsorption experiments the fluorescein was injected intraperitoneally. Back skin on the right side was swabbed with either dimethyl sulfoxide or 1% capsaicin in alcohol prior to the injections, and differences in skin fluorescence on the left (control) and right sides were recorded. One application of dimethyl sulfoxide or capsaicin increased the level of skin exsorption. Three applications of dimethyl sulfoxide almost doubled the amount of exsorbed dye, whereas three applications of the capsaicin inhibited the exsorption process. It was concluded that the fiber-optic perfusion fluorometer provides an excellent technique in support of other methods of investigating the skin.

Long-term Rebamipide and Diquafosol in Two Cases of Immune-Mediated Dry Eye

PubMed Central

Yamane, Mio; Ogawa, Yoko; Fukui, Masaki; Kamoi, Mizuka; Saijo-Ban, Yumiko; Yaguchi, Saori; Mukai, Shin; Kawakita, Tetsuya; Simmura, Shigeto; Tsubota, Kazuo

2015-01-01

ABSTRACT Purpose Two new drugs with mucin-inducing and secretion-promotive effects, rebamipide and diquafosol, were recently approved as topical dry-eye treatments. We report two cases in which the long-term use of mucin-inducing eye drops improved chronic ocular graft-versus-host disease (cGVHD)–related dry eye and ocular cicatricial pemphigoid (OCP)-like disease. Case Reports Case 1. A 61-year-old woman had cGVHD-related dry eye that resisted traditional medications. Next, we use topical diquafosol in addition to conventional treatments. The patient used diquafosol for 6 months without experiencing any side effects. The symptoms, including dry-eye sensation, ocular pain, foreign body sensation, and photophobia, as well as ocular surface findings including fluorescein and rose bengal scores and tear break-up time (TBUT), partly improved. To further improve the clinical signs and symptoms and decrease chronic inflammation, rebamipide was added to diquafosol. The symptoms, TBUT, and fluorescein and rose bengal scores markedly improved after long-term dual treatment without any side effects for 6 months. Case 2. A 77-year-old woman had OCP-like disease with dry eye. The patient did not improve using the currently available conventional treatments. Next, we use topical rebamipide in addition to conventional treatments. Symptoms including asthenopia, dry-eye sensation, ocular pain, and dull sensation, as well as fluorescein and rose bengal scores and TBUT, partly improved. Specifically, functional visual acuity was markedly improved after commencement of rebamipide. To further improve the clinical signs and symptoms and increase tear film stability and tear film volume, diquafosol was added to rebamipide. The combination of diquafosol and rebamipide worked for the patient. Improvements were seen in several symptoms, fluorescein and rose bengal scores, Schirmer test value, and TBUT without any side effects for 12 months. Conclusions Long-term treatment with topical rebamipide and diquafosol can improve dry eye in patients with cGVHD or OCP-like disease. PMID:25785527

Long-term rebamipide and diquafosol in two cases of immune-mediated dry eye.

PubMed

Yamane, Mio; Ogawa, Yoko; Fukui, Masaki; Kamoi, Mizuka; Saijo-Ban, Yumiko; Yaguchi, Saori; Mukai, Shin; Kawakita, Tetsuya; Simmura, Shigeto; Tsubota, Kazuo

2015-04-01

Two new drugs with mucin-inducing and secretion-promotive effects, rebamipide and diquafosol, were recently approved as topical dry-eye treatments. We report two cases in which the long-term use of mucin-inducing eye drops improved chronic ocular graft-versus-host disease (cGVHD)-related dry eye and ocular cicatricial pemphigoid (OCP)-like disease. Case 1. A 61-year-old woman had cGVHD-related dry eye that resisted traditional medications. Next, we use topical diquafosol in addition to conventional treatments. The patient used diquafosol for 6 months without experiencing any side effects. The symptoms, including dry-eye sensation, ocular pain, foreign body sensation, and photophobia, as well as ocular surface findings including fluorescein and rose bengal scores and tear break-up time (TBUT), partly improved. To further improve the clinical signs and symptoms and decrease chronic inflammation, rebamipide was added to diquafosol. The symptoms, TBUT, and fluorescein and rose bengal scores markedly improved after long-term dual treatment without any side effects for 6 months. Case 2. A 77-year-old woman had OCP-like disease with dry eye. The patient did not improve using the currently available conventional treatments. Next, we use topical rebamipide in addition to conventional treatments. Symptoms including asthenopia, dry-eye sensation, ocular pain, and dull sensation, as well as fluorescein and rose bengal scores and TBUT, partly improved. Specifically, functional visual acuity was markedly improved after commencement of rebamipide. To further improve the clinical signs and symptoms and increase tear film stability and tear film volume, diquafosol was added to rebamipide. The combination of diquafosol and rebamipide worked for the patient. Improvements were seen in several symptoms, fluorescein and rose bengal scores, Schirmer test value, and TBUT without any side effects for 12 months. Long-term treatment with topical rebamipide and diquafosol can improve dry eye in patients with cGVHD or OCP-like disease.

Meibomian Gland Dysfunction Determines the Severity of the Dry Eye Conditions in Visual Display Terminal Workers

PubMed Central

Dong, Nuo; Yang, Fan; Lin, Zhirong; Shang, Xumin; Li, Cheng

2014-01-01

Objective To explore meibomian gland dysfunction (MGD) may determine the severity of dry eye conditions in visual display terminal (VDT) workers. Methodology Prospective, case-control study carried out in China.106 eyes of 53 patients (VDT work time >4 hour per day) were recruited as the Long time VDT group; 80 eyes of 40 control subjects (VDT work time ≤4 hour per day) served as the Short time VDT group. A questionnaire of Ocular Surface Disease Index (OSDI) and multiple tests were performed. Three dry eye tests: tear film breakup time (BUT), corneal fluorescein staining, Schirmer I test; and three MGD parameters: lid margin abnormality score, meibum expression assessment (meibum score), and meibomian gland dropout degree (meiboscore) using Keratograph 5 M. Principal Findings OSDI and corneal fluorescein score were significantly higher while BUT was dramatically shorter in the long time VDT group than the short time VDT group. However, the average of Schirmer tear volumes was in normal ranges in both groups. Interestingly, the three MGD parameters were significantly higher in the long time VDT group than the short time one (P<0.0001). When 52 eyes with Schirmer <10 mm and 54 eyes with Schirmer ≥10 mm were separated from the long time VDT workers, no significant differences were found between the two subgroups in OSDI, fluorescein staining and BUT, as well as the three MGD parameters. All three MGD parameters were positively correlated with VDT working time (P<0.0001) and fluorescein scores (P<0.0001), inversely correlated with BUT (P<0.05), but not correlated with Schirmer tear volumes in the VDT workers. Conclusions Our findings suggest that a malfunction of meibomian glands is associated with dry eye patients in long term VDT workers with higher OSDI scores whereas some of those patients presenting a normal tear volume. PMID:25144638

Tear-Film Evaporation Rate from Simultaneous Ocular-Surface Temperature and Tear-Breakup Area.

PubMed

Dursch, Thomas J; Li, Wing; Taraz, Baseem; Lin, Meng C; Radke, Clayton J

2018-01-01

A corneal heat-transfer model is presented to quantify simultaneous measurements of fluorescein tear-breakup area (TBA) and ocular-surface temperature (OST). By accounting for disruption of the tear-film lipid layer (TFLL), we report evaporation rates through lipid-covered tear. The modified heat-transfer model provides new insights into evaporative dry eye. A quantitative analysis is presented to assess human aqueous tear evaporation rate (TER) through intact TFLLs from simultaneous in vivo measurement of time-dependent infrared OST and fluorescein TBA. We interpret simultaneous OST and TBA measurements using an extended heat-transfer model. We hypothesize that TBAs are ineffectively insulated by the TFLL and therefore exhibit higher TER than does that for a well-insulting TFLL-covered tear. As time proceeds, TBAs increase in number and size, thereby increasing the cornea area-averaged TER and decreasing OST. Tear-breakup areas were assessed from image analysis of fluorescein tear-film-breakup video recordings and are included in the heat-transfer description of OST. Model-predicted OSTs agree well with clinical experiments. Percent reductions in TER of lipid-covered tear range from 50 to 95% of that for pure water, in good agreement with literature. The physical picture of noninsulating or ruptured TFLL spots followed by enhanced evaporation from underlying cooler tear-film ruptures is consistent with the evaporative-driven mechanism for local tear rupture. A quantitative analysis is presented of in vivo TER from simultaneous clinical measurement of transient OST and TBA. The new heat-transfer model accounts for increased TER through expanding TBAs. Tear evaporation rate varies strongly across the cornea because lipid is effectively missing over tear-rupture troughs. The result is local faster evaporation compared with nonruptured, thick lipid-covered tear. Evaporative-driven tear-film ruptures deepen to a thickness where fluorescein quenching commences and local salinity rises to uncomfortable levels. Mitigation of tear-film rupture may therefore reduce dry eye-related symptoms.

Angiography reveals novel features of the retinal vasculature in healthy and diabetic mice.

PubMed

McLenachan, Samuel; Magno, Aaron Len; Ramos, David; Catita, Joana; McMenamin, Paul G; Chen, Fred Kuanfu; Rakoczy, Elizabeth Piroska; Ruberte, Jesus

2015-09-01

The mouse retina is a commonly used animal model for the study of pathogenesis and treatment of blinding retinal vascular diseases such as diabetic retinopathy. In this study, we aimed to characterize normal and pathological variations in vascular anatomy in the mouse retina using fluorescein angiography visualized with scanning laser ophthalmoscopy and optical coherence tomography (SLO-OCT). We examined eyes from C57BL/6J wild type mice as well as the Ins2(Akita) and Akimba mouse models of diabetic retinopathy using the Heidelberg Retinal Angiography (HRA) and OCT system. Angiography was performed on three focal planes to examine distinct vascular layers. For comparison with angiographic data, ex vivo analyses, including Indian ink angiography, histology and 3D confocal scanning laser microscopy were performed in parallel. All layers of the mouse retinal vasculature could be readily visualized during fluorescein angiography by SLO-OCT. Blood vessel density was increased in the deep vascular plexus (DVP) compared with the superficial vascular plexus (SVP). Arteriolar and venular typologies were established and structural differences were observed between venular types. Unexpectedly, the hyaloid artery was found to persist in 15% of C57BL/6 mice, forming anastomoses with peripheral retinal capillaries. Fluorescein leakage was easily detected in Akimba retinae by angiography, but was not observed in Ins2(Akita) mice. Blood vessel density was increased in the DVP of 6 month old Ins2(Akita) mice, while the SVP displayed reduced branching in precapillary arterioles. In summary, we present the first comprehensive characterization of the mouse retinal vasculature by SLO-OCT fluorescein angiography. Using this clinical imaging technique, we report previously unrecognized variations in C57BL/6J vascular anatomy and novel features of vascular retinopathy in the Ins2(Akita) mouse model of diabetes. Crown Copyright © 2015. Published by Elsevier Ltd. All rights reserved.

Fluorescein sodium-guided resection of cerebral metastases-an update.

PubMed

Höhne, Julius; Hohenberger, Christoph; Proescholdt, Martin; Riemenschneider, Markus J; Wendl, Christina; Brawanski, Alexander; Schebesch, Karl-Michael

2017-02-01

Cerebral metastasis (CM) is the most common malignancy affecting the brain. In patients eligible for surgery, complete tumor removal is the most important predictor of overall survival and neurological outcome. The emergence of surgical microscopes fitted with a fluorescein-specific filter have facilitated fluorescein-guided microsurgery and identification of tumor tissue. In 2012, we started evaluating fluorescein (FL) with the dedicated microscope filter in cerebral metastases (CM). After describing the treatment results of our first 30 patients, we now retrospectively report on 95 patients. Ninety-five patients with CM of different primary cancers were included (47 women, 48 men, mean age, 60 years, range, 25-85 years); 5 mg/kg bodyweight of FL was intravenously injected at induction of anesthesia. A YELLOW 560-nm filter (Pentero 900, ZEISS Meditec, Germany) was used for microsurgical tumor resection and resection control. The extent of resection (EOR) was assessed by means of early postoperative contrast-enhanced MRI and the grade of fluorescent staining as described in the surgical reports. Furthermore, we evaluated information on neurological outcome and surgical complications as well as any adverse events. Ninety patients (95%) showed bright fluorescent staining that markedly enhanced tumor visibility. Five patients (5%); three with adenocarcinoma of the lung, one with melanoma of the skin, and one with renal cell carcinoma) showed insufficient FL staining. Thirteen patients (14%) showed residual tumor tissue on the postoperative MRI. Additionally, the MRI of three patients did not confirm complete resection beyond doubt. Thus, gross-total resection had been achieved in 83% (n = 79) of patients. No adverse events were registered during the postoperative course. FL and the YELLOW 560-nm filter are safe and feasible tools for increasing the EOR in patients with CM. Further prospective evaluation of the FL-guided technique in CM-surgery is in planning.

Predicting success with silicone-hydrogel contact lenses in new wearers.

PubMed

Best, Nigel; Drury, Laura; Wolffsohn, James S

2013-10-01

to evaluate changes in tear metrics and ocular signs induced by six months of silicone-hydrogel contact lens wear and the difference in baseline characteristics between those who successfully continued in contact lens wear compared to those that did not. Non-invasive Keratograph, Tearscope and fluorescein tear break-up times (TBUTs), tear meniscus height, bulbar and limbal hyperaemia, lid-parallel conjunctival folds (LIPCOF), phenol red thread, fluorescein and lissamine-green staining, and lid wiper epitheliopathy were measured on 60 new contact lens wearers fitted with monthly silicone-hydrogels (average age 36±14 years, 40 females). Symptoms were evaluated by the Ocular Surface Disease Index (OSDI). After six months full time contact lens wear the above metrics were re-measured on those patients still in contact lens wear (n=33). The initial measurements were also compared between the group still wearing lenses after six months and those who had ceased lens wear (n=27). There were significant changes in tear meniscus height (p=0.031), bulbar hyperaemia (p=0.011), fluorescein TBUT (p=0.027), corneal (p=0.007) and conjunctival (p=0.009) staining, LIPCOF (p=0.011) and lid wiper epitheliopathy (p=0.002) after six months of silicone-hydrogel wear. Successful wearers had a higher non-invasive (17.0±8.2s vs 12.0±5.6s; p=0.001) and fluorescein (10.7±6.4s vs 7.5±4.7s; p=0.001) TBUT than drop-outs, although OSDI (cut-off 4.2) was also a strong predictor of success. Silicone-hydrogel lenses induced significant changes in the tear film and ocular surface as well as lid margin staining. Wettability of the ocular surface is the main factor affecting contact lens drop-out. Copyright © 2013 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

Effect of Povidone Iodine 5% on the Cornea, Vision, and Subjective Comfort.

PubMed

Ridder, William H; Oquindo, Caren; Dhamdhere, Kavita; Burke, James

2017-07-01

To determine the effects of povidone iodine 5% (Betadine 5%) applied to the eye on visual function, corneal integrity, and subjective complaints. Ten subjects were chosen to participate in this study (average age 40.6 ± 19.6; age range 22-68). LogMAR acuity, contrast sensitivity, corneal fluorescein staining, and subjective complaints were measured before and after 60 μl of Betadine 5% or saline were applied to one eye (eligibility visit, baseline, 5, 30, and 60 minutes and 4 and 24 hours post-application). Contrast sensitivity at 14 cpd was determined with a spatial two-alternative, forced choice procedure (Beethoven software). The National Eye Institute (NEI) grid pattern was used to grade corneal staining with sodium fluorescein. To avoid the detrimental effects of sodium fluorescein administration on functional vision, the corneal staining baseline was measured at the eligibility visit. Subjective complaints were monitored using the Schein dry eye questionnaire. The data were analyzed with an ANOVA. Saline administration did not alter logMAR acuity, contrast sensitivity, corneal fluorescein staining, or subjective complaints at any visit. Betadine administration significantly reduced the logMAR acuity, compared to baseline and the saline data, at the 30- and 60-minute visits (all P values <.05). The contrast sensitivity was significantly reduced compared to the baseline and the saline results at 5, 30, and 60 minutes after Betadine application (all P values <.05). The Betadine significantly increased the Schein questionnaire score, compared both to baseline and the saline data, at the 1- and 4-hour visits (all P values <.05). Total corneal staining and the maximum NEI sector staining were significantly different from baseline and from the saline results at every visit (all P values <.05). Betadine 5% application significantly decreases epithelial integrity of the cornea, decreases functional vision, and increases subjective complaints.

The Blood-Retinal Barrier in the Management of Retinal Disease: EURETINA Award Lecture.

PubMed

Cunha-Vaz, José

2017-01-01

Retinal diseases are the main causes of blindness in the Western world. Diabetic retinopathy and age-related macular degeneration continue to increase in prevalence and as main causes of vision loss. Intravitreal anti-VEGF and steroid injections have raised new expectations for their successful treatment. These agents act by stabilizing the blood-retinal barrier (BRB). Our group defined the BRB by identifying for the first time the tight junctions that unite retinal endothelial cells and are the basis for the inner BRB, an observation later confirmed in retinal pigment epithelial cells and in brain vessels. A major role of active transport processes was also identified. Today, the BRB is understood to play a fundamental role in retinal function in both health and disease. Retinal edema, an ubiquitous manifestation of retinal disease, is directly associated with breakdown of the BRB and with vision loss. In its most common form (i.e., vasogenic edema), due to breakdown of the BRB, Starling's law of capillary filtration may be used to interpret the mechanisms of fluid accumulation in the retina. The main factors involved in the development of retinal edema are BRB permeability, capillary hydrostatic pressure, tissue hydrostatic pressure, tissue osmotic pressure, and plasma osmotic pressure. In the clinical environment, breakdown of the BRB has been identified by fluorescein angiography and vitreous fluorometry, requiring the intravenous administration of fluorescein. An OCT-based method, OCT-Leakage, recently introduced by our group is capable of noninvasively identifying and quantifying sites of alteration of the BRB by mapping areas of lower-than-normal optical reflectivity, thus reflecting changes in the retinal extracellular fluid. We found good correspondence between the location of increased areas of low optical reflectivity identified by OCT-Leakage and the main sites of leakage on fluorescein angiography. Furthermore, with OCT-Leakage the areas of abnormal fluid accumulation can be identified in specific retinal layers, clearly offering more information than previously obtained with fluorescein angiography. OCT angiography has become available, replacing much of the information yielded by fluorescein angiography in a noninvasive manner. However, OCT angiography cannot visualize the leakage, i.e., the alteration of the BRB. OCT-Leakage is able to identify the locations of increases in extracellular fluid in the different layers of the retina. The complementarity of these 2 methods is of potential great interest for the diagnosis and management of several retinal diseases in which the presence and amount of fluid, as a marker of severity and activity, is paramount to treatment and management decisions in clinical practice. © 2017 S. Karger AG, Basel.

Meridional lenticular astigmatism associated with bilateral concurrent uveal metastases in renal cell carcinoma.

PubMed

Priluck, Joshua C; Grover, Sandeep; Chalam, Kv

2012-01-01

To demonstrate a case illustrating meridional lenticular astigmatism as a result of renal cell carcinoma uveal metastases. Case report with images. Clinical findings and diagnostic testing of a patient with acquired meridional lenticular astigmatism are described. The refraction revealed best-corrected visual acuity of 20/20-1 OD (-2.50 + 0.25 × 090) and 20/50 OS (-8.25 + 3.25 × 075). Bilateral concurrent renal cell carcinoma metastases to the choroid and ciliary body are demonstrated by utilizing ultrasonography, ultrawidefield fluorescein angiography, and unique spectral-domain optical coherence tomography. Metastatic disease should be included in the differential of acquired astigmatism. Spectral-domain optical coherence tomography, ultrawidefield fluorescein angiography, and ultrasonography have roles in delineating choroidal metastases.

Decolorization and degradation of xanthene dyes by a white rot fungus, Coriolus versicolor.

PubMed

Itoh, Kiyoharu; Yatome, Chizuko

2004-01-01

The decolorization of six xanthene dyes (conc. 100 microM) by a white rot fungus, Coriolus versicolor (C. versicolor), was investigated in liquid culture. The decolorization of Fluorescein, 4-Aminofluorescein, and 5-Aminofluorescein by the fungus was 85.0, 95.0, and 91.9% after 14 days incubation, respectively. However, no decolorization of Rhodamine B, Rhodamine 123 hydrate, and Rhodamine 6G was observed. The first three dyes also were decolorized with cell-free extracts from C. versicolor. The decolorization activity was 10.2, 6.7, and 7.2 microM min(-1)mg(-1), respectively. Thin layer chromatography (TLC) analyses indicated that degradation of Fluorescein was occurring with the detection of three degradation products.

Injection-depth-locking axial motion guided handheld micro-injector using CP-SSOCT.

PubMed

Cheon, Gyeong Woo; Huang, Yong; Kwag, Hye Rin; Kim, Ki-Young; Taylor, Russell H; Gehlbach, Peter L; Kang, Jin U

2014-01-01

This paper presents a handheld micro-injector system using common-path swept source optical coherence tomography (CP-SSOCT) as a distal sensor with highly accurate injection-depth-locking. To achieve real-time, highly precise, and intuitive freehand control, the system used graphics processing unit (GPU) to process the oversampled OCT signal with high throughput and a smart customized motion monitoring control algorithm. A performance evaluation was conducted with 60-insertions and fluorescein dye injection tests to show how accurately the system can guide the needle and lock to the target depth. The evaluation tests show our system can guide the injection needle into the desired depth with 4.12 um average deviation error while injecting 50 nl of fluorescein dye.

On the barrier properties of the cornea: a microscopy study of the penetration of fluorescently labeled nanoparticles, polymers, and sodium fluorescein.

PubMed

Mun, Ellina A; Morrison, Peter W J; Williams, Adrian C; Khutoryanskiy, Vitaliy V

2014-10-06

Overcoming the natural defensive barrier functions of the eye remains one of the greatest challenges of ocular drug delivery. Cornea is a chemical and mechanical barrier preventing the passage of any foreign bodies including drugs into the eye, but the factors limiting penetration of permeants and nanoparticulate drug delivery systems through the cornea are still not fully understood. In this study, we investigate these barrier properties of the cornea using thiolated and PEGylated (750 and 5000 Da) nanoparticles, sodium fluorescein, and two linear polymers (dextran and polyethylene glycol). Experiments used intact bovine cornea in addition to bovine cornea de-epithelialized or tissues pretreated with cyclodextrin. It was shown that corneal epithelium is the major barrier for permeation; pretreatment of the cornea with β-cyclodextrin provides higher permeation of low molecular weight compounds, such as sodium fluorescein, but does not enhance penetration of nanoparticles and larger molecules. Studying penetration of thiolated and PEGylated (750 and 5000 Da) nanoparticles into the de-epithelialized ocular tissue revealed that interactions between corneal surface and thiol groups of nanoparticles were more significant determinants of penetration than particle size (for the sizes used here). PEGylation with polyethylene glycol of a higher molecular weight (5000 Da) allows penetration of nanoparticles into the stroma, which proceeds gradually, after an initial 1 h lag phase.

DELIVERY OF WATER-SOLUBLE DRUGS USING ACOUSTICALLY-TRIGGERED, PERFLUOROCARBON DOUBLE EMULSIONS

PubMed Central

Fabiilli, Mario L.; Lee, James A.; Kripfgans, Oliver D.; Carson, Paul L.; Fowlkes, J. Brian

2010-01-01

Purpose Ultrasound can be used to release a therapeutic payload encapsulated within a perfluorocarbon (PFC) emulsion via acoustic droplet vaporization (ADV), a process whereby the PFC phase is vaporized and the agent is released. ADV-generated microbubbles have been previously used to selectively occlude blood vessels in vivo. The coupling of ADV-generated drug delivery and occlusion has therapeutically, synergistic potentials. Methods Micron-sized, water-in-PFC-in-water (W1/PFC/W2) emulsions were prepared in a two-step process using perfluoropentane (PFP) or perfluorohexane (PFH) as the PFC phase. Fluorescein or thrombin was contained in the W1 phase. Results Double emulsions containing fluorescein in the W1 phase displayed a 5.7±1.4 fold and 8.2±1.3 fold increase in fluorescein mass flux, as measured using a Franz diffusion cell, after ADV for the PFP and PFH emulsions, respectively. Thrombin was stably retained in four out of five double emulsions. For three out of five formulations tested, the clotting time of whole blood decreased, in a statistically significant manner (p < 0.01), when incubated with thrombin-loaded emulsions exposed to ultrasound compared to emulsions not exposed to ultrasound. Conclusions ADV can be used to spatially and temporally control the delivery of water-soluble compounds formulated in PFC double emulsions. Thrombin release could extend the duration of ADV-generated, microbubble occlusions. PMID:20872050

Features of central serous chorioretinopathy presenting at a tertiary care hospital in Lahore.

PubMed

Jamil, Ahmad Zeeshan; Mirza, Khurram Azam; Qazi, Zaheer Uddin Aqil; Iqbal, Wasim; Khaliq, Javed; Fawad-ur-Rahman; Ahmed, Arslan

2013-04-01

To evaluate the clinical, angiographic and optical coherence tomographic features of central serous chorioretinopathy in patients presenting at a tertiary care centre in Lahore. The observational study was conducted at the Layton Rehmatulla Benevolent Trust Eye and Cancer Hospital Lahore from July 15, 2010 to December 15, 2011. Patients who had received prior treatment for the condition and allergy to fluorescein were excluded. There were 86 eyes of 64 adult patients with central serous chorioretinopathy. The following data was recorded: history, signs and symptoms, best corrected visual acuity, fundus fluorescein angiography, and central macular thickness measurement with optical coherence tomography. Data was analyzed using SPSS 17. Mean age of patients who presented during the study duration was 39.52 +/- 8.85 years.There were 53 (82.8%) males and 11(17.2%) females. Of the total, 42 (65.6%) cases had unilateral and 22 (34.4%) cases had bilateral involvement. Chronic central serous chorioretinopathy was seen in 27 (42.2%) cases while 37 (57.8%) cases were acute presentations. Retinal pigment epithelial detachment was observed in 29 (45.3%) cases. On fundus fluorescein angiography, there were 62 (72.1%) eyes that showed ink blot pattern. Median visual acuity at presentation was 0.25. Median central macular thickness at presentation was 550.5micro. Central serous chorioretinopathy in the study sample was associated with pigment epithelial detachment, bilateral involvement, and presence of systemic diseases.

Phototoxic effects of commercial photographic flash lamp on rat eyes.

PubMed

Inoue, Makoto; Shinoda, Kei; Ohde, Hisao; Tezuka, Keiji; Hida, Tetsuo

2006-11-01

To determine whether exposure of the cornea and retina of rats to flashes from a commercial photographic flash lamp is phototoxic. Sprague-Dawley rats were exposed to 10, 100, or 1,000 flashes of the OPTICAM 16M photographic flash lamp (Fujikoeki, Japan) placed 0.1, 1, or 3 m from the eyes. Corneal damage was assessed by a fluorescein staining score, and the retinal damage by eletroretinography (ERG) and histology before and 24 h after exposure. Exposure of the eyes to 1,000 flashes at 0.1 m increased the fluorescein staining score significantly (P = 0.009, the Mann-Whitney test). Scanning electron microscopy (SEM) of the cornea showed a detachment of the epithelial cells from the surface after this exposure. The amplitude of the a-wave was decreased significantly by 23.0% (P = 0.026) of the amplitude before the exposure, and the b-wave by 19.7% (P = 0.0478) following 1,000 flashes at 0.1 m but not by the other exposures. TUNEL-positive cells were present in the outer nuclear layer only after the extreme exposure, but no significant decrease in retinal thickness was seen under any condition. The fluorescein staining score and ERGs recovered to control levels within 1 week. Light exposure to a photographic flash lamp does not induce damage to the cornea and retina except when they are exposed to 1,000 flashes at 0.1 m.

[The clinical investigation of peripheral retinal changes of anterior uveitis].

PubMed

Guo, Chunying; Qiao, Lijun; Yang, Liu

2015-10-01

To observe the peripheral retinal changes of anterior uveitis patients using ultra-wide field fluorescein angiography. Retrospective case series study. Thirty-three eyes of 20 patients diagnosed as anterior uveitis from September to November 2013 in the ophthalmic clinic of Peking University First Hospital was included. All the patients underwent routine ocular examination, followed by ultra-wide field Fluorescein. There were 13 male patients and 7 female patients, aged 23-66, an average of 46 years old. The anterior uveitis recurrent in both eyes in 13 patients and only one eye in 7 patients. Uveitis duration ranging from months to 30 years. Twenty eyes were at the active stage of the anterior uveitis while 13 eyes in the quiescent stage. Peripheral retinal vessels leakage was detected in 57.6% (19/33) of the eyes, among which in eyes with active disease the rate of peripheral retinal vessels leakage was 75% (15/20), and in eyes with quiescent disease the rate was 30.8% (4/13). Optic disk hyperfluorescence was found in 6 of 20 eyes with active disease. Cystoid macular edema was found in 4 eyes. In addition, peripheral retinal hyperfluorescence dot and zone were detected in 2 eyes. Peripheral retinal vessels were detected to have inflammative leakage in patients of anterior uveitis by ultra-wide field fluorescein agiography. Peripheral retinal inflammation may still exist even in the quiescent stage of the anterior uveitis.

Vascular changes on fluorescein angiography of premature infants with low risk of retinopathy of prematurity after high oxygen exposure.

PubMed

Martinez-Castellanos, Maria Ana; Velez-Montoya, Raul; Price, Kenneth; Henaine-Berra, Andree; García-Aguirre, Gerardo; Morales-Canton, Virgilio; Cernichiaro-Espinosa, Linda Alejandra

2017-01-01

To describe a wide array of peripheral vascular changes using fluorescein angiography in preterm neonates, without high risk characteristics for developing retinopathy of prematurity, that were exposed to high oxygen concentration. Retrospective, two center, case series. Newborns at two different hospitals with ≥1500 g or gestational age of ≥32 weeks, fluorescein angiography performed, and with high oxygen exposure without adequate control were included. 294 infants diagnosed with ROP were analyzed. Only 28 eyes from 14 patients with peripheral vascular abnormalities in older and heavier babies were included. Two distinct type of peripheral vascular changes were observed: group 1 or non-proliferative: areas of capillary non-perfusion along with widespread arteriovenous shunting between adjacent primary vessels, tortuosity of primary vessels, abnormal budding of tertiary vessels and capillaries, abnormal capillary tufts and absence of foveal avascular zone; group 2 or proliferative: all of the characteristics of group 1 plus leakage of dye from the boundary between perfused and non-perfused retina and/or optic disc. Peripheral vascular abnormalities different from retinopathy of prematurity are observed in older than 32 weeks of gestational age, and heavier than 1500 g babies. This makes the authors classify these patients as having a disease caused solely by oxygen dysregulation at the neonatal intensive care unit similarly to the oxygen induced retinopathy in experimental studies.

On tear film breakup (TBU): dynamics and imaging.

PubMed

Braun, Richard J; Driscoll, Tobin A; Begley, Carolyn G; King-Smith, P Ewen; Siddique, Javed I

2018-06-13

We report the results of some recent experiments to visualize tear film dynamics. We then study a mathematical model for tear film thinning and tear film breakup (TBU), a term from the ocular surface literature. The thinning is driven by an imposed tear film thinning rate which is input from in vivo measurements. Solutes representing osmolarity and fluorescein are included in the model. Osmolarity causes osmosis from the model ocular surface, and the fluorescein is used to compute the intensity corresponding closely to in vivo observations. The imposed thinning can be either one-dimensional or axisymmetric, leading to streaks or spots of TBU, respectively. For a spatially-uniform (flat) film, osmosis would cease thinning and balance mass lost due to evaporation; for these space-dependent evaporation profiles TBU does occur because osmolarity diffuses out of the TBU into the surrounding tear film, in agreement with previous results. The intensity pattern predicted based on the fluorescein concentration is compared with the computed thickness profiles; this comparison is important for interpreting in vivo observations. The non-dimensionalization introduced leads to insight about the relative importance of the competing processes; it leads to a classification of large vs small TBU regions in which different physical effects are dominant. Many regions of TBU may be considered small, revealing that the flow inside the film has an appreciable influence on fluorescence imaging of the tear film.

Confocal laser-induced fluorescence detector for narrow capillary system with yoctomole limit of detection.

PubMed

Weaver, Mitchell T; Lynch, Kyle B; Zhu, Zaifang; Chen, Huang; Lu, Joann J; Pu, Qiaosheng; Liu, Shaorong

2017-04-01

Laser-induced fluorescence (LIF) detectors for low-micrometer and sub-micrometer capillary on-column detection are not commercially available. In this paper, we describe in details how to construct a confocal LIF detector to address this issue. We characterize the detector by determining its limit of detection (LOD), linear dynamic range (LDR) and background signal drift; a very low LOD (~70 fluorescein molecules or 12 yoctomole fluorescein), a wide LDR (greater than 3 orders of magnitude) and a small background signal drift (~1.2-fold of the root mean square noise) are obtained. For detecting analytes inside a low-micrometer and sub-micrometer capillary, proper alignment is essential. We present a simple protocol to align the capillary with the optical system and use the position-lock capability of a translation stage to fix the capillary in position during the experiment. To demonstrate the feasibility of using this detector for narrow capillary systems, we build a 2-μm-i.d. capillary flow injection analysis (FIA) system using the newly developed LIF prototype as a detector and obtain an FIA LOD of 14 zeptomole fluorescein. We also separate a DNA ladder sample by bare narrow capillary - hydrodynamic chromatography and use the LIF prototype to monitor the resolved DNA fragments. We obtain not only well-resolved peaks but also the quantitative information of all DNA fragments. Copyright © 2016 Elsevier B.V. All rights reserved.

Gross, Histologic, and Computed Tomographic Anatomy of the Lacrimal System of Snakes

PubMed Central

Souza, Nicole M.; Maggs, David J.; Park, Shin Ae; Puchalski, Sarah; Reilly, Christopher M.; Paul-Murphy, Joanne; Murphy, Christopher J.

2014-01-01

Objective To describe the lacrimal system of snakes using contrast micro-computed tomography (micro-CT) with 3-dimensional reconstruction, fluorescein passage (“Jones”) testing, histology, and gross dissection. Animals studied One Royal Python and 19 snake cadavers representing 10 species. Procedures Direct observation following injection of fluorescein into the subspectacular space, micro-CT following injection of 3 contrast agents into the subspectacular space, gross dissection following injection of latex into the subspectacular space, and histopathology. Results Injection of fluorescein confirmed patency but not course of the lacrimal duct. Barium enabled clear visualization of the lacrimal duct whereas two iodinated contrast agents proved inadequate. Collectively, micro-CT, anatomic dissections, and histology suggest tears are produced by a single, large, serous, retrobulbar gland, released into the subspectacular space via several ductules, and drained through a single punctum originating in the ventronasal subspectacular space and the lacrimal duct taking one of 3 routes of variable tortuosity before opening into the oral cavity in close association with the opening of the duct of the vomeronasal organ. Conclusions The ophidian lacrimal duct has a generally tortuous course and the details of its anatomy is species variable. The tortuous course of the duct likely predisposes snakes to duct occlusion and must be considered when planning medical and surgical interventions in snakes with pseudobuphthalmos and subspectacular abscessation. PMID:24862081

Fluorescence chemodosimeter for dopamine based on the inner filter effect of the in situ generation of silver nanoparticles and fluorescent dye

NASA Astrophysics Data System (ADS)

Uppa, Yuwapon; Ngamdee, Kessarin; Promarak, Vinich; Ngeontae, Wittaya

2018-07-01

A new strategy for the sensitive and selective detection of dopamine (DA) was proposed. The chemodosimeter design was based on the measurement of the fluorescent quenching of fluorescein dye caused by the in situ generation of silver nanoparticles (AgNPs). The AgNPs can be simply generated by a reaction between DA and Ag+ in the presence of polymethacrylic acid (PMAA). In addition, the generated AgNPs possess the maximum surface plasmon resonance (SPR) at 440 nm and an increase in the SPR intensity with an increasing DA concentration. Basically, fluorescein dye can emit the fluorescent intensity maximum at 513 nm with excitation at 487 nm. Thus, fluorescent quenching was achieved due to an inner filter effect from the overlap between the excitation spectrum of the fluorescein dye and the SPR spectrum of the generated AgNPs. The degree of fluorescent quenching linearly depends on the number of generated AgNPs that can be directly related to the concentration of DA. The proposed chemodosimeter can be used to detect DA in a working linear concentration range of 1.0-5.0 μM at a detection limit of 10.6 nM. This chemodosimeter was successfully applied to determine DA in a real urine sample and a dopamine injection formulation with satisfactory results.

Corneal Abrasions

MedlinePlus

... the doctor looks at the eye under a light that is filtered cobalt blue. The fluorescein causes the abrasion to glow bright green under the light. The doctor also might do a standard ophthalmic ...

Fluorescein eye stain

MedlinePlus

Abnormal results may point to: Abnormal tear production (dry eye) Blocked tear duct Corneal abrasion (a scratch on ... object in eye ) Infection Injury or trauma Severe dry eye associated with arthritis (keratoconjunctivitis sicca)

Multifocal choroiditis - an unusual finding in Crohns disease.

PubMed

Vianna, R N G; Ozdal, P C; Deschnes, J

To report a patient with Crohns disease and acute decreased vision in one eye secondary to multifocal choroiditis and serous retinal detachment. A complete ocular examination, including fluorescein angiography, was performed. Fundus biomicroscopy disclosed multifocal, deep, discretely elevated yellowish lesions at the posterior pole of the affected eye. Fluorescein angiographic study of these lesions revealed early hypofluorescence followed by late yperfluorescence. Subtenonian injection of corticosteroids rapidly induced remission of the choroidal lesions. Chorioretinal involvement in patients with Crohns disease may or may not be related to reactivation of this disorder. Therefore, even patients without gastrointestinal symptoms who present with posterior segment inflammation must be informed of this. The chorioretinal inflammatory lesions do seem to respond promptly to periocular injection of corticosteroids. (Eur J Ophthalmol 2004; 14: 345-9).

Fluorescein isothiocyanate and rhodamine B dye encapsulated mesoporous SiO2 for applications of blue LED excited white LED

NASA Astrophysics Data System (ADS)

Das, Sourav; Manam, J.

2018-05-01

In this work, the fluorescein isothiocyanate (FITC) and rhodamine B (RhB) dyes were encapsulated in mesoporous silica nanoparticles (MSNp). The MSNp-FITC-RhB nanohybrids phosphor showed a dichromatic PL emission at green region and orange region when excited at 460 nm. A Forster Resonance Energy Transfer (FRET) was observed from FITC to RhB. The materials were further characterized by XRD, FTIR, TEM, and temperature dependent photoluminescence. The CIE coordinates were tuned from greenish yellow to the orange region and quantum yield was reached 52.04% based on FRET. So by combining the MSNp-FITC-RhB nanohybrids phosphor with the blue LED chip, the white light emission with flexible Color Correlated Temperature and improved Color Rendering Index can be obtained.

Meridional lenticular astigmatism associated with bilateral concurrent uveal metastases in renal cell carcinoma

PubMed Central

Priluck, Joshua C; Grover, Sandeep; Chalam, KV

2012-01-01

Purpose To demonstrate a case illustrating meridional lenticular astigmatism as a result of renal cell carcinoma uveal metastases. Methods Case report with images. Results Clinical findings and diagnostic testing of a patient with acquired meridional lenticular astigmatism are described. The refraction revealed best-corrected visual acuity of 20/20–1 OD (−2.50 + 0.25 × 090) and 20/50 OS (−8.25 + 3.25 × 075). Bilateral concurrent renal cell carcinoma metastases to the choroid and ciliary body are demonstrated by utilizing ultrasonography, ultrawidefield fluorescein angiography, and unique spectral-domain optical coherence tomography. Conclusions Metastatic disease should be included in the differential of acquired astigmatism. Spectral-domain optical coherence tomography, ultrawidefield fluorescein angiography, and ultrasonography have roles in delineating choroidal metastases. PMID:23152663

Resonant Mode-hopping Micromixing

PubMed Central

Jang, Ling-Sheng; Chao, Shih-Hui; Holl, Mark R.; Meldrum, Deirdre R.

2009-01-01

A common micromixer design strategy is to generate interleaved flow topologies to enhance diffusion. However, problems with these designs include complicated structures and dead volumes within the flow fields. We present an active micromixer using a resonating piezoceramic/silicon composite diaphragm to generate acoustic streaming flow topologies. Circulation patterns are observed experimentally and correlate to the resonant mode shapes of the diaphragm. The dead volumes in the flow field are eliminated by rapidly switching from one discrete resonant mode to another (i.e., resonant mode-hop). Mixer performance is characterized by mixing buffer with a fluorescence tracer containing fluorescein. Movies of the mixing process are analyzed by converting fluorescent images to two-dimensional fluorescein concentration distributions. The results demonstrate that mode-hopping operation rapidly homogenized chamber contents, circumventing diffusion-isolated zones. PMID:19551159

[Bilateral abnormalities in central serous chorioretinopathy seen in optical coherence tomography, ultra-widefield fluorescein angiography and microperimetry--case report].

PubMed

Tylus, Magdalena; Święch-Zubilewicz, Anna; Dolar-Szczasny, Joanna; Mackiewicz, Jerzy

2015-01-01

Central serous chorioretinopathy is a common retinopathy, which is manifested by the idiopathic detachment of the neurosensory retina in the posterior pole, secondary to fluid leakage from choroidal vessels at the level of retinal pigment epithelium. The disease is typically unilateral and affects young men. We present a case of a 48-year old man, admitted to the Department of Vitreo-Retinal Surgery, Medical University in Lublin, reporting vision impairment in his right eye. The bilateral ocular exam followed by optical coherence tomography, ultra-widefield fluorescein angiography and microperimetry revealed bilateral central serous chorioretinopathy. This case presents a bilateral manifestation of central serous chorioretinopathy and emphasizes the role of advanced diagnostic imaging techniques in analyzing retinal function and disease management.

Flow cytometry of HEK 293T cells interacting with polyelectrolyte multilayer capsules containing fluorescein-labeled poly(acrylic acid) as a pH sensor.

PubMed

Reibetanz, Uta; Halozan, David; Brumen, Milan; Donath, Edwin

2007-06-01

Polyelectrolyte multilayer sensor capsules, 5 microm in diameter, which contained fluorescein-labeled poly(acrylic acid) (PAAAF) as pH-sensitive reporter molecules, were fabricated and employed to explore their endocytotic uptake into HEK 293T cells by flow cytometry. The percentage of capsules residing in the endolysosomal compartment was estimated from the fluorescence intensity decrease caused by acidification. Capsules attached to the extracellular surface of the plasma membrane were identified by trypan blue quenching. The number of capsules in the cytoplasm was rather small, being below the detection limit of the method. The advantages of polyelectrolyte multilayer capsules are that the fluorophore is protected from interaction with cellular compartments and that the multilayer can be equipped with additional functions.

Certain tricyclic and pentacyclic-hetero nitrogen rhodol dyes

DOEpatents

Haugland, Richard P.; Whitaker, James E.

1993-01-01

Novel fluorescent dyes based on the rhodol structure are provided. The new reagents contain functional groups capable of forming a stable fluorescent product with functional groups typically found in biomolecules or polymers including amines, phenols, thiols, acids, aldehydes and ketones. Reactive groups in the rhodol dyes include activated esters, isothiocyanates, amines, hydrazines, halides, acids, azides, maleimides, aldehydes, alcohols, acrylamides and haloacetamides. The products are detected by their absorbance or fluorescence properties. The spectral properties of the fluorescent dyes are sufficiently similar in wavelengths and intensity to fluorescein or rhodamine derivatives as to permit use of the same equipment. The dyes, however, show less spectral sensitivity to pH in the physiological range than does fluorescein, have higher solubility in non-polar solvents and have improved photostability and quantum yields.

Effect of individual SCFA on the epithelial barrier of sheep rumen under physiological and acidotic luminal pH conditions.

PubMed

Greco, Gabriele; Hagen, Franziska; Meißner, Svenja; Shen, Zanming; Lu, Zhongyan; Amasheh, Salah; Aschenbach, Jörg R

2018-02-15

The objective of this study was to investigate whether individual short-chain fatty acids (SCFA) have a different potential to either regulate the formation of the ruminal epithelial barrier (REB) at physiological pH or to damage the REB at acidotic ruminal pH. Ruminal epithelia of sheep were incubated in Ussing chambers on their mucosal side in buffered solutions (pH 6.1 or 5.1) containing no SCFA (control), 30 mM of either acetate, propionate or butyrate, or 100 mM acetate. Epithelial conductance (Gt), short-circuit current (Isc), and fluorescein flux rates were measured over 7 h. Thereafter, mRNA and protein abundance, as well as localization of the tight junction proteins claudin (Cldn)-1, -4, -7, and occludin were analyzed. At pH 6.1, butyrate increased Gt and decreased Isc, with additional decreases in claudin-7 mRNA and protein abundance (each P < 0.05) and disappearance of Cldn-7 immunosignals from the stratum corneum. By contrast, the mRNA abundance of Cldn-1 and/or Cldn-4 were upregulated by 30 mM propionate, 30 mM butyrate, or 100 mM acetate (P < 0.05), however, without coordinated changes in protein abundance. At luminal pH 5.1, neither Gt, Isc, nor TJ protein abundance was altered in the absence of SCFA; only fluorescein flux rates were slightly increased (P < 0.05) and fluorescein signals were no longer restricted to the stratum corneum. The presence of acetate, propionate, or butyrate at pH 5.1 increased fluorescein flux rates and Gt, and decreased Isc (each P < 0.05). Protein abundance of Cldn-1 was decreased in all SCFA treatments but 30 mM butyrate; abundance of Cldn -4 and -7 was decreased in all SCFA treatments but 30 mM acetate; and abundance of occludin was decreased in all SCFA treatments but 30 mM propionate (each P < 0.05). Immunofluorescence staining of SCFA-treated tissues at pH 5.1 showed disappearance of Cldn-7, discontinuous pattern for Cldn-4 and blurring of occludin and Cldn-1 signals in tight junction complexes. The fluorescein dye appeared to freely diffuse into deeper cell layers. The strongest increase in Gt and consistent decreases in the abundance and immunosignals of tight junction proteins were observed with 100 mM acetate at pH 5.1. We conclude that SCFA may contribute differently to the REB formation at luminal pH 6.1 with possible detrimental effects of butyrate at 30 mM concentration. At luminal pH 5.1, all SCFA elicited REB damage with concentration appearing more critical than SCFA species.

Bovine and Porcine Transscleral Solute Transport: Influence of Lipophilicity and the Choroid–Bruch’s Layer

PubMed Central

Cheruvu, Narayan P. S.; Kompella, Uday B.

2012-01-01

Purpose To determine the influence of the choroid–Bruch’s layer and solute lipophilicity on in vitro transscleral drug permeability in bovine and porcine eyes. Methods The in vitro permeability of two VEGF inhibitory drugs, budesonide and celecoxib, which are lipophilic and neutral at physiologic pH, and of three marker solutes, 3H-mannitol (hydrophilic, neutral), sodium fluorescein (hydrophilic, anionic), and rhodamine 6G (lipophilic, cationic), were determined across freshly excised scleras, with or without the underlying choroid–Bruch’s layer. Select studies were performed using porcine sclera with and without choroid–Bruch’s layer. Neural retina was removed by exposure of the eyecup to isotonic buffer and wherever required, the retinal pigment epithelial (RPE) layer of the preparation was disrupted and removed by exposure to hypertonic buffer. Because of the poor solubility of celecoxib and budesonide, permeability studies were conducted with 5% wt/vol of hydroxypropyl-β-cyclodextrin (HPβCD). For other solutes, permeability studies were conducted, with and without HPβCD. Partitioning of the solutes into bovine sclera and choroid–Bruch’s layer was also determined. Results The calculated log (distribution coefficient) values were −2.89, −0.68, 2.18, 3.12, and 4.02 for mannitol, sodium fluorescein, budesonide, celecoxib, and rhodamine 6G, respectively. Removal of RPE was confirmed by transmission electron microscopy and differences in the transport of mannitol. The order of the permeability coefficients (Papp) across sclera and sclera–choroid–Bruch’s layers in bovine and porcine models was 3H-mannitol > fluorescein > budesonide > celecoxib > rhodamine 6G, with HPβCD, and 3H-mannitol > fluorescein > rhodamine 6G, without HPβCD. The presence of choroid–Bruch’s layer reduced the bovine scleral permeability by 2-, 8-, 16-, 36-, and 50-fold and porcine tissue permeability by 2-, 7-, 15-, 33-, and 40-fold, respectively, for mannitol, sodium fluorescein, budesonide, celecoxib, and rhodamine 6G. The partition coefficients measured in bovine tissues correlated positively with the log (distribution coefficient) and exhibited a trend opposite that of transport. The partition coefficient ratio of bovine choroid–Bruch’s layer to sclera was ~1, 1.5, 1.7, 2, and 3.5, respectively, for the solutes, as listed earlier. Conclusions The choroid–Bruch’s layer is a more significant barrier to drug transport than is sclera. It hinders the transport of lipophilic solutes, especially a cationic solute, more than hydrophilic solutes and in a more dramatic way than does sclera. The reduction in transport across this layer directly correlates with solute binding to the tissue. Understanding the permeability properties of sclera and underlying layers would be beneficial in designing better drugs for transscleral delivery. PMID:17003447

Slit-lamp exam

MedlinePlus

... eyes, especially the eyelids, cornea, conjunctiva, sclera, and iris. Often a yellow dye (fluorescein) is used to ... lens on the front of the eye) Eyelids Iris (colored part of the eye between the cornea ...

Fluorescein angiography

MedlinePlus

... other retinopathy High blood pressure Inflammation or edema Macular degeneration Microaneurysms -- enlargement of capillaries in the retina Tumors ... Eye Problems Read more Eye Diseases Read more Macular Degeneration Read more A.D.A.M., Inc. is ...

Exploring the energy landscape of antibody-antigen complexes: protein dynamics, flexibility, and molecular recognition.

PubMed

Thielges, Megan C; Zimmermann, Jörg; Yu, Wayne; Oda, Masayuki; Romesberg, Floyd E

2008-07-08

The production of antibodies that selectively bind virtually any foreign compound is the hallmark of the immune system. While much is understood about how sequence diversity contributes to this remarkable feat of molecular recognition, little is known about how sequence diversity impacts antibody dynamics, which is also expected to contribute to molecular recognition. Toward this goal, we examined a panel of antibodies elicited to the chromophoric antigen fluorescein. On the basis of isothermal titration calorimetry, we selected six antibodies that bind fluorescein with diverse binding entropies, suggestive of varying contributions of dynamics to molecular recognition. Sequencing revealed that two pairs of antibodies employ homologous heavy chains that were derived from common germline genes, while the other two heavy chains and all six of the light chains were derived from different germline genes and are not homologous. Interestingly, more than half of all the somatic mutations acquired during affinity maturation among the six antibodies are located in positions unlikely to contact fluorescein directly. To quantify and compare the dynamics of the antibody-fluorescein complexes, three-pulse photon echo peak shift and transient grating spectroscopy were employed. All of the antibodies exhibited motions on three distinct time scales, ultrafast motions on the <100 fs time scale, diffusive motions on the picosecond time scale, and motions that occur on time scales longer than nanoseconds and thus appear static. However, the exact frequency of the picosecond time scale motion and the relative contribution of the different motions vary significantly among the antibody-chromophore complexes, revealing a high level of dynamic diversity. Using a hierarchical model, we relate the data to features of the antibodies' energy landscapes as well as their flexibility in terms of elasticity and plasticity. In all, the data provide a consistent picture of antibody flexibility, which interestingly appears to be correlated with binding entropy as well as with germline gene use and the mutations introduced during affinity maturation. The data also provide a gauge of the dynamic diversity of the antibody repertoire and suggest that this diversity might contribute to molecular recognition by facilitating the recognition of the broadest range of foreign molecules.

Smokestack leak in central serous chorioretinopathy.

PubMed

Bujarborua, Dhiren; Nagpal, Pran N; Deka, Manab

2010-03-01

To study the demography, various morphological patterns and fluid dynamics of the smokestack leak by fluorescein angiography (FA) in central serous chorioretinopathy (CSC). Part I (clinical): review of the medical records and angiographic documents of 69 consecutive cases of CSC with smokestack leak. Part II (experimental): documentation of the movement of various concentrations of fluorescein dye due to convection currents in a laboratory model that roughly represents a closed chamber similar to that of CSC in human eyes. The clinical study (Part I) revealed that 14.40% of 479 consecutive cases had smokestack leak, of which 70% occurred in first acute episode (p-value: <0.001), 27.14% in acute recurrent episodes (50% fresh leak) and 2.85% in chronic stage. Patients were predominantly male (84.05%) with a median age of 34.00 +/- 8.14 years. The median symptom duration excluding the chronic cases was 15 +/- 34.28 days. This type of leak was mostly (48.57%) seen in medium-sized CSC, and the majority were in the parafoveal superonasal quadrant (31.42%). The ascending type of leak was predominant (94.28%). In four eyes, an atypical pattern and in two eyes more than one smokestack leak were seen within the same detached area. The experimental study (Part II) demonstrated that fluid containing a low concentration of fluorescein ascended due to convection currents, whereas highly concentrated dye descended. The clinical study revealed smokestack leaks to be significantly more common in a primary acute episode, and they usually develop in the early part of the acute phase of the disease (average duration 15 +/- 34.28 days). Rarely, this type of leak can occur in the chronic stage, and multiple leaks may develop in the same detached space. The various patterns of dye movement due to convection currents in the experimental model resembled the dye movement in certain cases of CSC of the present series. The experimental study also hinted at the probability of drainage of unbound fluorescein molecules along with protein-laden heavy fluid in downward spread of the leak.

Sodium Fluorescein-Guided Resection under the YELLOW 560 nm Surgical Microscope Filter in Malignant Gliomas: Our First 38 Cases Experience.

PubMed

Zhang, Ningning; Tian, Hailong; Huang, Dezhang; Meng, Xianbing; Guo, Wenqiang; Wang, Chaochao; Yin, Xin; Zhang, Hongying; Jiang, Bin; He, Zheng; Wang, Zhigang

2017-01-01

Sodium fluorescein (FL) had been safely used in fluorescence-guided microsurgery for imaging various brain tumors. Under the YELLOW 560 nm surgical microscope filter, low-dose FL as a fluorescent dye helps in visualization. Our study investigated the safety and efficacy of this innovative technique in malignant glioma (MG) patients. 38 patients suffering from MGs confirmed by pathology underwent FL-guided resection under YELLOW 560 nm surgical microscope filter. We retrospectively analyzed the clinical characters, microsurgery procedure, extent of resection, pathology of MGs, progression-free survival (PFS), and overall survival (OS). Thirty-eight patients had MGs (10 WHO grade III, 28 WHO grade IV). With YELLOW 560 nm surgical microscope filter combined with neuronavigation, sodium fluorescein-guided gross total resection (GTR) was achieved in 35 (92.1%) patients and subtotal resection in 3 (7.69%). The sensitivity and specificity of FL were 94.4% and 88.6% regardless of radiographic localization. Intraoperatively, 10 biopsies (10/28 FL[+]) showed "low" or "high" fluorescence in non-contrast-enhancement region and are also confirmed by pathology. Our data showed 6-month PFS of 92.3% and median survival of 11 months. FL-guided resection of MGs under the YELLOW 560 nm surgical microscope filter combined with neuronavigation was safe and effective, especially in non-contrast-MRI regions. It is feasible for improving the extent of resection in MGs especially during emergency cases.

Poly(amido)amine dendrimers generation 4.0 (PAMAM G4) reduce blood hyperglycaemia and restore impaired blood-brain barrier permeability in streptozotocin diabetes in rats.

PubMed

Karolczak, Kamil; Rozalska, Sylwia; Wieczorek, Marek; Labieniec-Watala, Magdalena; Watala, Cezary

2012-10-15

We hypothesized that BBB is impaired in rat model of streptozotocin-induced diabetes and can be sealed by poly(amido)amine dendrimers G4.0 (PAMAM G4), which reveal anti-glycation activity. The BBB permeabilization was monitored in rats with the 60-day streptozotocin-diabetes and non-diabetic animals, using three fluorescent dyes (given intraperitoneally) differing in molecular weight: fluorescein, fluorescein isothiocyanate (FITC)-dextran and Evans blue. All animals were administered for 2 months with either PAMAM G4 dendrimer or placebo. The fluorescence intensities of the injected fluorescent markers were recorded in the homogenates of selected brain regions. The highest accumulations of the used fluorescent dyes were observed for fluorescein, predominantly in thalamus, hippocampus, frontal cortex, striatum and cerebellum. FITC-dextran leaked to much smaller extent, however, higher permeabilization for FITC-dextran was revealed in pons-medulla oblongata, frontal and parietal cortex of diabetic compared to control animals. Evans blue leaked very slowly into striatum and pons-medulla oblongata in diabetic rats. The treatment of diabetic animals with PAMAM G4 significantly reduced blood glucose concentration and hallmarks of late diabetic complications, compared to non-treated diabetic animals. PAMAM G4 significantly reduced diabetes-induced permeabilization of BBB, which remained in line with the reduced blood glucose and the amelioration of the biochemical hallmarks of severe hyperglycaemia. Copyright © 2012 Elsevier B.V. All rights reserved.

A novel bioerodible deep scleral lamellar cyclosporine implant for uveitis.

PubMed

Gilger, Brian C; Salmon, Jacklyn H; Wilkie, David A; Cruysberg, Lars P J; Kim, Jonghyeon; Hayat, Matt; Kim, Hyuncheol; Kim, Stephanie; Yuan, Peng; Lee, Susan S; Harrington, Susan M; Murray, Patrick R; Edelhauser, Henry F; Csaky, Karl G; Robinson, Michael R

2006-06-01

To determine the feasibility, safety, and effectiveness of an episcleral or deep scleral lamellar sustained release cyclosporine (CsA) device in a naturally occurring animal model of uveitis. A two-compartment perfusion chamber was used to assess in vitro human and equine scleral permeability of fluorescein, dexamethasone-fluorescein, or CsA. A biodegradable, matrix-reservoir CsA implant was designed, and release rates of CsA were determined in vitro. Tissue CsA levels were measured in eyes with the implant. Horses with equine recurrent uveitis (ERU) received episcleral or deep scleral lamellar CsA implants and were monitored for up to 3 years. Dexamethasone-fluorescein and CsA penetrated the in vitro equine sclera poorly; however, low but detectable levels of CsA were detected intraocularly in vivo. The implant placed episclerally failed to control inflammatory episodes in ERU. CsA implants placed in the deep sclera adjacent to the suprachoroidal space resulted in high levels of CsA in most ocular tissues. In clinical equine patients with ERU, frequency of uveitic flare-ups was significantly decreased after implantation of a deep scleral lamellar CsA implant. Diffusion of CsA across the sclera from the episcleral space was not a feasible method of drug delivery to the equine eye. However, placing a deep scleral lamellar CsA implant adjacent to the suprachoroidal space was effective in achieving therapeutic ocular drug concentrations and controlling uveitis in horses with ERU.

Acute choroidal ischemia associated with toxoplasmic retinochoroiditis.

PubMed

Khairallah, Moncef; Yahia, Salim Ben; Zaouali, Sonia; Jenzeri, Salah; Attia, Sonia; Messaoud, Riadh

2007-09-01

To describe eight patients with active toxoplasmic retinochoroiditis (RC) who had features suggestive of acute choroidal ischemia. A retrospective review of the clinical records of 23 consecutive patients with acute toxoplasmic RC was performed. All patients underwent detailed ophthalmic examination at presentation and throughout follow-up, including dilated biomicroscopic fundus examination, fundus photography, fluorescein angiography, and indocyanine green (ICG) angiography. Of 23 patients, 8 (34.8%) had a large area of retinal whitening surrounding a small focus of RC. Fluorescein as well as ICG angiography showed a well demarcated geographic area of early choroidal hypofluorescence that extended beyond the clinical borders of the white retinal lesion, particularly by ICG angiography. Associated findings for these 8 patients included old retinochoroidal scars (7 [87.5%]), serous retinal detachment (3 [37.5%]), retinal hemorrhages (1 [12.5%]), and multiple satellite dark dots by ICG angiography (6 [75%]). Seven of eight patients were treated using a combination of antitoxoplasmic drugs and corticosteroids. All findings seen at the acute stage resolved in 2 weeks to 6 weeks. A small atrophic retinochoroidal scar replaced the active toxoplasmic lesion and was surrounded with mild or moderate retinal pigment epithelium changes that were associated with decreased final visual acuity in 2 patients (25%). Patients with toxoplasmic RC may develop features suggestive of choroidal ischemia that can result in a transient or permanent decrease in vision. Choroidal ischemia can only be suspected clinically, and fluorescein angiography and ICG angiography are required to establish the definitive diagnosis.

Phosphate-modified TiO2 nanoparticles for selective detection of dopamine, levodopa, adrenaline, and catechol based on fluorescence quenching.

PubMed

Wu, Hsin-Pin; Cheng, Tian-Lu; Tseng, Wei-Lung

2007-07-03

For the first time, an aqueous solution, comprising 6-nm phosphate-modified titanium dioxide (P-TiO2) nanoparticles (NPs) and fluorescein, has been used for sensing dopamine (DA), levodopa (L-DOPA), adrenaline, and catechol. The complexes obtained by means of chelation of surface Ti(IV) ions with an enediol group exhibit strong absorption at 428 nm; thus, they can be designed as efficient quenchers for fluorescein. The fluorescence of a fluorescein solution containing 1.4 mM P-TiO2 NPs at pH 8.0 decreases if the solution comprises DA, L-DOPA, adrenaline, and catechol, but not noradrenaline, ascorbic acid, and salicylic acid. We consider that P-TiO2 NPs have a number of advantages over bare TiO2 NPs, such as ease of preparation, high selectivity, and high stability. By measuring fluorescence quenching, the limits of detection at a signal-to-noise ratio of 3 are calculated as 33.5, 81.8, 20.3, and 92.1 nM for DA, L-DOPA, adrenaline, and catechol, respectively. In contrast, UV-vis absorption reveals the relatively poor sensitivity of these compounds. We have validated the applicability of our method by means of analyses of DA in urine samples. High-performance liquid chromatography in combination with an electrochemical cell has been used to further confirm our results. We believe that this approach has great potential for diagnostic purposes.

Automated Detection of Leakage in Fluorescein Angiography Images with Application to Malarial Retinopathy

PubMed Central

Zhao, Yitian; J. C. MacCormick, Ian; G. Parry, David; Leach, Sophie; A. V. Beare, Nicholas; P. Harding, Simon; Zheng, Yalin

2015-01-01

The detection and assessment of leakage in retinal fluorescein angiogram images is important for the management of a wide range of retinal diseases. We have developed a framework that can automatically detect three types of leakage (large focal, punctate focal, and vessel segment leakage) and validated it on images from patients with malarial retinopathy. This framework comprises three steps: vessel segmentation, saliency feature generation and leakage detection. We tested the effectiveness of this framework by applying it to images from 20 patients with large focal leak, 10 patients with punctate focal leak, and 5,846 vessel segments from 10 patients with vessel leakage. The sensitivity in detecting large focal, punctate focal and vessel segment leakage are 95%, 82% and 81%, respectively, when compared to manual annotation by expert human observers. Our framework has the potential to become a powerful new tool for studying malarial retinopathy, and other conditions involving retinal leakage. PMID:26030010

Automated detection of leakage in fluorescein angiography images with application to malarial retinopathy.

PubMed

Zhao, Yitian; MacCormick, Ian J C; Parry, David G; Leach, Sophie; Beare, Nicholas A V; Harding, Simon P; Zheng, Yalin

2015-06-01

The detection and assessment of leakage in retinal fluorescein angiogram images is important for the management of a wide range of retinal diseases. We have developed a framework that can automatically detect three types of leakage (large focal, punctate focal, and vessel segment leakage) and validated it on images from patients with malarial retinopathy. This framework comprises three steps: vessel segmentation, saliency feature generation and leakage detection. We tested the effectiveness of this framework by applying it to images from 20 patients with large focal leak, 10 patients with punctate focal leak, and 5,846 vessel segments from 10 patients with vessel leakage. The sensitivity in detecting large focal, punctate focal and vessel segment leakage are 95%, 82% and 81%, respectively, when compared to manual annotation by expert human observers. Our framework has the potential to become a powerful new tool for studying malarial retinopathy, and other conditions involving retinal leakage.

Alkylation of Staurosporine to Derive a Kinase Probe for Fluorescence Applications.

PubMed

Disney, Alexander J M; Kellam, Barrie; Dekker, Lodewijk V

2016-05-06

The natural product staurosporine is a high-affinity inhibitor of nearly all mammalian protein kinases. The labelling of staurosporine has proven effective as a means of generating protein kinase research tools. Most tools have been generated by acylation of the 4'-methylamine of the sugar moiety of staurosporine. Herein we describe the alkylation of this group as a first step to generate a fluorescently labelled staurosporine. Following alkylation, a polyethylene glycol linker was installed, allowing subsequent attachment of fluorescein. We report that this fluorescein-staurosporine conjugate binds to cAMP-dependent protein kinase in the nanomolar range. Furthermore, its binding can be antagonised with unmodified staurosporine as well as ATP, indicating it targets the ATP binding site in a similar fashion to native staurosporine. This reagent has potential application as a screening tool for protein kinases of interest. © 2015 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

Batch-Versuche zur Bestimmung der Sorption und Reaktionskinetik von fluoreszierenden Tracern

NASA Astrophysics Data System (ADS)

Vaitl, Tobias; Wohnlich, Stefan

2018-06-01

For many tracer experiments, prior determination of interaction between solid medium and used tracers is of major interest in order to achieve efficient, economic and successful field experiments. In the present study, three different types of batch experiments were performed with three fluorescent dyes (Na-Fluorescein, Amidorhodamin G and Tinopal CBS-X) and three different rock types (sandstone, claystone and limestone), to determine distribution coefficients and reaction kinetics. All three rock types were analysed for organic carbon content, specific surface area and mineralogical composition to identify the main sorption mechanisms. For all tracers, different sorption properties were found depending on the type of rock. The strongest sorption was observed for Tinopal CBS-X in contact with claystone. Only Na-Fluorescein showed sorption (albeit limited) in contact with the sandstones. The investigated limestones indicated a high sorption for the tracer Tinopal CBS-X. Regarding reaction kinetics, in most cases, thermodynamic equilibrium conditions were reached after two weeks.

Ultra-Widefield Fluorescein Angiography in Intermediate Uveitis.

PubMed

Laovirojjanakul, Wipada; Acharya, Nisha; Gonzales, John A

2017-10-17

To examine associations between pattern of vascular leakage on ultrawide-field fluorescein angiography (UWFFA) and visual acuity, cystoid macular edema (CME), and inflammatory activity in intermediate uveitis. Single center cross-sectional, retrospective review of medical records, spectral domain optical coherence tomography (SD-OCT) and angiographic images of intermediate uveitis patients who underwent UWFFA over a 12-month period. Forty-one eyes from 24 patients were included. Twelve eyes (29%) exhibited peripheral leakage, 26 eyes (64%) had diffuse leakage and three eyes (7%) had no leakage. Diffuse leakage was associated with 0.2 logMAR worse visual acuity than peripheral leakage (p = 0.02). There was no statistically significant difference in the odds of having CME when diffuse leakage was compared to peripheral leakage. UWFFA identifies retinal vascular pathology in intermediate uveitis not present on clinical examination. Diffuse retinal vascular leakage was associated with worse visual acuity when compared to peripheral and no leakage patterns.

Decompensation of a congenital retinal macrovessel with arteriovenous communications induced by repetitive rollercoaster rides.

PubMed

Beatty, S; Goodall, K; Radford, R; Lavin, M J

2000-10-01

To describe a congenital retinal venous macrovessel that communicates with a cilioretinal artery and a retinal artery, and to report how this vascular anomaly decompensated as a result of repetitive rollercoaster rides. Case report with serial fundus photography and fluorescein angiography. After a short period of intensive rollercoaster rides, a 19-year-old woman complained of reduced vision in one eye. Funduscopy and fluorescein angiography revealed a venous congenital retinal macrovessel with arteriovenous communications, and retinal exudation was visible at the termination of the anomalous vessel. Exudation resolved, and acuity recovered after a period of avoidance of rollercoaster rides. This case represents the first report of a retinal artery and a cilioretinal artery communicating with a congenital retinal macrovessel, and it suggests that such patients are at increased risk of retinal vascular decompensation if involved in activities associated with changes in g-forces, such as bungee jumping or rollercoaster rides.

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

Kamel, R.S.; Landon, J.; Smith, D.S.

Nortriptyline derivatives prepared by reaction with fluorescein isothiocyanate or conjugation to N-acetyl-L-histidine were radioiodinated and the products purified with Sephadex LH-20 columns to obtain two novel nortriptyline radioligands. Antisera were raised in rabbits by immunization with nortriptyline conjugated to succinylated ovine albumin. By use of the iodinated fluorescein derivative we developd a liquid-phase second-antibody radioimmunoassay that gives results correlating closely (r = 0.98) with those by an established radioimmunoassay of similar specificity in the assay of apparent total amitriptyline and its metabolite nortriptyline in serum or plasma from patients being treated with these drugs. With the iodinated N-acetyl-L-histidin derivative wemore » developed a magnetizable solid-phase second-antibody radioimmunoassay. The cross reactivities of amitriptyline and nortriptyline could be made equal by performing the assay at pH 9.0, which makes it possible to measure true total active drug concentrations in patients receiving amitriptyline.« less

Altered blood-brain barrier permeability and its effect on the distribution of Evans blue and sodium fluorescein in the rat brain applied by intracarotid injection.

PubMed

Kozler, P; Pokorný, J

2003-01-01

The aim was to study the blood-brain permeability according to the distribution in the rat brain of Evans blue (EB) and sodium fluorescein (NaFl) administered by an intracarotid injection. Eighteen animals were divided into six groups according to the state of the blood-brain barrier (BBB) at the moment when the dyes were being applied. In the first two groups, the BBB was intact, in groups 3 and 4 the barrier had been opened osmotically prior to the application of the dyes, and in groups 5 and 6 a cellular edema was induced by hyperhydration before administration of the dyes. The intracellular and extracellular distribution of the dyes was studied by fluorescence microscopy. The histological picture thus represented the morphological correlate of the way BBB permeability had been changed before the application of the dyes.

Chemical reactivation of fluorescein isothiocyanate immunofluorescence-labeled resin-embedded samples

NASA Astrophysics Data System (ADS)

Li, Longhui; Rao, Gong; Lv, Xiaohua; Chen, Ruixi; Cheng, Xiaofeng; Wang, Xiaojun; Zeng, Shaoqun; Liu, Xiuli

2018-02-01

Resin embedding is widely used and facilitates microscopic imaging of biological tissues. In contrast, quenching of fluorescence during embedding process hinders the application of resin embedding for imaging of fluorescence-labeled samples. For samples expressing fluorescent proteins, it has been demonstrated that the weakened fluorescence could be recovered by reactivating the fluorophore with alkaline buffer. We extended this idea to immunofluorescence-labeling technology. We showed that the fluorescence of pH-sensitive fluorescein isothiocyanate (FITC) was quenched after resin embedding but reactivated after treating by alkaline buffer. We observed 138.5% fluorescence preservation ratio of reactivated state, sixfold compared with the quenched state in embedding resin, which indicated its application for fluorescence imaging of high signal-to-background ratio. Furthermore, we analyzed the chemical reactivation mechanism of FITC fluorophore. This work would show a way for high-resolution imaging of immunofluorescence-labeled samples embedded in resin.

An Enzyme-Responsive "Turn-on" Fluorescence Polymeric Superamphiphile as a Potential Visualizable Phosphate Prodrug Delivery Vehicle.

PubMed

Yang, Xi; Shen, Shihong; Guo, Li; Tan, Jidong; Lei, Henxin; Wu, Jianghan; Zhao, Lei; Xiong, Tao; Wu, Youshen; Cheng, Yilong; Zhang, Yanfeng

2018-06-01

The development of inexpensive and highly efficient enzyme-responsive polymers has significantly contributed to targeted drug delivery systems. Here, a superamphiphile with a capability of fluorescent dissociation sensing is designed. It is constructed with negatively charged adenosine 5'-triphosphate (ATP) and negatively charged fluorescein diphosphate (FDP), which are used as fluorescence detection, and a cationic diblock copolymer methoxy-poly(ethylene glycol) 113 -b-poly(2-dimethyl-aminoethyl methacrylate) 70 . Upon addition of calf intestinal alkaline phosphatase, the superamphiphile disintegrates, presumably due to the enzymatic hydrolysis of ATP. This process is accompanied by an increase in the fluorescence emission intensity of fluorescein owing to the hydrolysis of FDP. The in vitro application of the superamphiphile is also proven. Thus, the "turn-on" fluorescence of the superamphiphile serves as a real-time module for detection of the disintegration of superamphiphile. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

STUDIES ON INFLUENZA INFECTION IN FERRETS BY MEANS OF FLUORESCEIN-LABELLED ANTIBODY

PubMed Central

Liu, Ch'ien

1955-01-01

By means of fluorescein-labelled antibody staining, specific influenza viral antigens were seen in both the cytoplasm and the nucleus of infected ciliated epithelial cells covering the nasal turbinates of infected ferrets. Initially, only a small portion of the nasal epithelium showed fluorescence, and no appreciable abnormality of the cells could be detected by hematoxylin and eosin staining. The fluorescence soon spread to involve the entire epithelium, followed by desquamation coinciding with the onset of manifest illness. Pneumonia was seen in some of the infected ferrets, and in them, viral antigens were found in the bronchial epithelium and in the mediastinal lymph nodes. A rise of viral infectivity titer paralleled the observed spread of viral antigens. Many desquamated nasal epithelial cells and macrophages containing antigen were present in nasal smears. The finding would seem to offer a method for the rapid specific diagnosis of influenza infection. PMID:14367687

Identification of mycobacterial surface proteins released into subcellular compartments of infected macrophages.

PubMed

Beatty, W L; Russell, D G

2000-12-01

Considerable effort has focused on the identification of proteins secreted from Mycobacterium spp. that contribute to the development of protective immunity. Little is known, however, about the release of mycobacterial proteins from the bacterial phagosome and the potential role of these molecules in chronically infected macrophages. In the present study, the release of mycobacterial surface proteins from the bacterial phagosome into subcellular compartments of infected macrophages was analyzed. Mycobacterium bovis BCG was surface labeled with fluorescein-tagged succinimidyl ester, an amine-reactive probe. The fluorescein tag was then used as a marker for the release of bacterial proteins in infected macrophages. Fractionation studies revealed bacterial proteins within subcellular compartments distinct from mycobacteria and mycobacterial phagosomes. To identify these proteins, subcellular fractions free of bacteria were probed with mycobacterium-specific antibodies. The fibronectin attachment protein and proteins of the antigen 85-kDa complex were identified among the mycobacterial proteins released from the bacterial phagosome.

Development of hydrogel TentaGel shell-core beads for ultrahigh throughput solution-phase screening of encoded OBOC combinatorial small molecule libraries.

PubMed

Baek, Hyoung Gee; Liu, Ruiwu; Lam, Kit S

2009-01-01

The one-bead one-compound (OBOC) combinatorial library method enables the rapid generation and screening of millions of discrete chemical compounds on beads. Most of the OBOC screening methods require the library compounds to remain tethered to the bead during screening process. Methods have also been developed to release library compounds from immobilized beads for in situ solution phase or "lawn" assays. However, this latter approach, while extremely powerful, is severely limited by the lack of suitable solid supports for such assays. Here, we report on the development of a novel hydrogel TentaGel shell-core (HTSC) bead in which hydrogel is grafted onto the polystyrene-based TentaGel (TG) bead as an outer shell (5-80 mum thick) via free radical surface-initiated polymerization. This novel shell-core bilayer resin enables the preparation of encoded OBOC combinatorial small molecule libraries, such that the library compounds reside on the highly hydrophilic outer layer and the coding tags reside in the polystyrene-based TG core. Using fluorescein as a model small molecule compound, we have demonstrated that fluorescein molecules that have been linked covalently to the hydrogel shell via a disulfide bond could readily diffuse out of the hydrogel layer into the bead surrounding after reduction with dithiothreitol. In contrast, under identical condition, the released fluorescein molecules remained bound to unmodified TG bead. We have prepared an encoded OBOC small molecule library on the novel shell-core beads and demonstrated that the beads can be readily decoded.

Changes in rabbit corneal innervation induced by the topical application of benzalkonium chloride.

PubMed

Chen, Wensheng; Zhang, Zhenhao; Hu, Jiaoyue; Xie, Hui; Pan, Juxin; Dong, Nuo; Liu, Zuguo

2013-12-01

To investigate the effect of benzalkonium chloride (BAK) on corneal nerves. Fifty-four adult New Zealand Albino rabbits were randomly divided into 3 groups. BAK at concentrations of 0.005%, 0.01%, or 0.02% was applied once daily to 1 eye of each rabbit for 9 days. The contralateral untreated eyes were used as controls. Corneal mechanical sensitivity, aqueous tear production, tear break-up time (BUT), fluorescein, and Rose Bengal staining scores were compared with those of control values on days 3, 6, and 9. Corneal whole mounts were immunostained with a specific antitubulin βIII antibody to label nerve fibers. Epithelial superficial nerve terminal, subbasal, and stromal nerve fiber densities were quantified. The structure of the central cornea was examined by means of in vivo confocal microscopy on day 9. The topical application of BAK resulted in lower corneal sensitivity and higher Rose Bengal staining scores on day 3, whereas there were no significant changes in the BUT, Schirmer, and corneal fluorescein scores. Decreased nerve densities in superficial and subbasal layers were observed in BAK-treated eyes on days 3 and 6, respectively. The eyes treated with 0.02% BAK exhibited significantly reduced Schirmer scores, BUT, and stromal nerve fiber density, and increased fluorescein staining scores on day 9. Corneal superficial epithelial cell size was significantly larger in all BAK-treated eyes compared with that in control eyes. The topical application of BAK can quickly cause corneal hypoesthesia without tear deficiency. Changes in corneal innervation significantly correlate with BAK-induced ocular surface changes.

Semifluorinated Alkane Eye Drops for Treatment of Dry Eye Disease Due to Meibomian Gland Disease.

PubMed

Steven, Philipp; Augustin, Albert J; Geerling, Gerd; Kaercher, Thomas; Kretz, Florian; Kunert, Kathleen; Menzel-Severing, Johannes; Schrage, Norbert; Schrems, Wolfgang; Krösser, Sonja; Beckert, Michael; Messmer, Elisabeth M

2017-11-01

Meibomian gland disease is generally accepted as the leading cause for evaporative dry eye disease (DED). In a previous study, perfluorohexyloctane, a semifluorinated alkane, has been demonstrated to significantly increase tear film breakup time and to reduce corneal fluorescein staining in patients with evaporative DED, thereby vastly reducing dry eye-related symptoms. This study was set up to evaluate perfluorohexyloctane in a larger population of patients with Meibomian gland dysfunction. Seventy-two patients with Meibomian gland disease and associated dry eye received 1 drop of perfluorohexyloctane 4 times daily during an observational, prospective, multicenter, 6-8-week study. Clinical assessment included best-corrected visual acuity, intraocular pressure, Schirmer test I, tear film breakup time, anterior and posterior blepharitis assessment, number of expressible Meibomian glands, meibum quality and quantity, ocular surface fluorescein staining, lid margin and symptom assessment, and Ocular Surface Disease Index (OSDI © ). From the 72 patients recruited, 61 completed the trial per protocol. Nine patients did not apply the medication as recommended and 2 patients were lost to follow-up. Tear film breakup time, corneal and conjunctival fluorescein staining, number of expressible Meibomian glands, and severity of anterior and posterior blepharitis significantly improved after 6-8 weeks of perfluorohexyloctane application. In addition, symptoms improved as demonstrated by a significant decrease of OSDI-values from 37 (±13) to 26 (±16). In concordance with previous findings, 6-8 weeks of topical application of perfluorohexyloctane significantly improves clinical signs of Meibomian gland disease and associated mild to moderate DED.

Evaluation of a Novel Artificial Tear in the Prevention and Treatment of Dry Eye in an Animal Model.

PubMed

She, Yujing; Li, Jinyang; Xiao, Bing; Lu, Huihui; Liu, Haixia; Simmons, Peter A; Vehige, Joseph G; Chen, Wei

2015-11-01

To evaluate effects of a novel multi-ingredient artificial tear formulation containing carboxymethylcellulose (CMC) and hyaluronic acid (HA) in a murine dry eye model. Dry eye was induced in mice (C57BL/6) using an intelligently controlled environmental system (ICES). CMC+HA (Optive Fusion™), CMC-only (Refresh Tears(®)), and HA-only (Hycosan(®)) artificial tears and control phosphate-buffered saline (PBS) were administered 4 times daily and compared with no treatment (n = 64 eyes per group). During regimen 1 (prevention regimen), mice were administered artificial tears or PBS for 14 days (starting day 0) while they were exposed to ICES, and assessed on days 0 and 14. During regimen 2 (treatment regimen), mice exposed to ICES for 14 days with no intervention were administered artificial tears or PBS for 14 days (starting day 14) while continuing exposure to ICES, and assessed on days 0, 14, and 28. Corneal fluorescein staining and conjunctival goblet cell density were measured. Artificial tear-treated mice had significantly better outcomes than control groups on corneal staining and goblet cell density (P < 0.01). Mice administered CMC+HA also showed significantly lower corneal fluorescein staining and higher goblet cell density, compared with CMC (P < 0.01) and HA (P < 0.05) in both regimens 1 and 2. The artificial tear formulation containing CMC and HA was effective in preventing and treating environmentally induced dry eye. Improvements observed for corneal fluorescein staining and conjunctival goblet cell retention suggest that this combination may be a viable treatment option for dry eye disease.

Dynamic Loading of Immature Epiphyseal Cartilage Pumps Nutrients out of Vascular Canals

PubMed Central

Albro, Michael B.; Banerjee, Rajan E.; Li, Roland; Oungoulian, Sevan R.; Chen, Bo; del Palomar, Amaya P.; Hung, Clark T.; Ateshian, Gerard A.

2011-01-01

The potential influence of mechanical loading on transvascular transport in vascularized soft tissues has not been explored extensively. This experimental investigation introduced and explored the hypothesis that dynamic mechanical loading can pump solutes out of blood vessels and into the surrounding tissue, leading to faster uptake and higher solute concentrations than could otherwise be achieved under unloaded conditions. Immature epiphyseal cartilage was used as a model tissue system, with fluorescein (332 Da), dextran (3, 10 and 70 kDa) and transferrin (80 kDa) as model solutes. Cartilage disks were either dynamically loaded (±10% compression over a 10% static offset strain, at 0.2 Hz) or maintained unloaded in solution for up to 20 hours. Results demonstrated statistically significant solute uptake in dynamically loaded (DL) explants relative to passive diffusion (PD) controls for all solutes except unbound fluorescein, as evidenced by the DL:PD concentration ratios after 20 hours (1.0 ± 0.2, 2.4 ± 1.1, 6.1 ± 3.3, 9.0 ± 4.0, and 5.5±1.6 for fluorescein, 3, 10, and 70 kDa dextran, and transferrin). Significant uptake enhancements were also observed within the first 30 seconds of loading. Termination of dynamic loading produced dissipation of enhanced solute uptake back to PD control values. Confocal images confirmed that solute uptake occurred from cartilage canals into their surrounding extracellular matrix. The incidence of this loading-induced transvascular solute pumping mechanism may significantly alter our understanding of the interaction of mechanical loading and tissue metabolism. PMID:21481875

PCR-enzyme-linked immunosorbent assay and partial rRNA gene sequencing: a rational approach to identifying mycobacteria.

PubMed Central

Patel, S; Yates, M; Saunders, N A

1997-01-01

A PCR-enzyme-linked immunosorbent assay (ELISA) for amplification and rapid identification of mycobacterial DNA coding for 16S rRNA was developed. The PCR selectively targeted and amplified part of the 16S rRNA gene from all mycobacteria while simultaneously labelling one strand of the amplified product with a 5' fluorescein-labelled primer. The identity of the labelled strand was subsequently determined by hybridization to a panel of mycobacterial species-specific capture probes, which were immobilized via their 5' biotin ends to a streptavidin-coated microtiter plate. Specific hybridization of a 5' fluorescein-labelled strand to a species probe was detected colorimetrically with an anti-fluorescein enzyme conjugate. The assay was able to identify 10 Mycobacterium spp. A probe able to hybridize to all Mycobacterium species (All1) was also included. By a heminested PCR, the assay was sensitive enough to detect as little as 10 fg of DNA, which is equivalent to approximately three bacilli. The assay was able to detect and identify mycobacteria directly from sputa. The specificities of the capture probes were assessed by analysis of 60 mycobacterial strains corresponding to 18 species. Probes Avi1, Int1, Kan1, Xen1, Che1, For1, Mal1, Ter1, and Gor1 were specific. The probe Tbc1 cross-hybridized with the Mycobacterium terrae amplicon. Analysis of 35 strains tested blind resulted in 34 strains being correctly identified. This method could be used for rapid identification of early cultures and may be suitable for the detection and concurrent identification of mycobacteria within clinical specimens. PMID:9276419

Fluorescein diacetate (FDA) and its analogue as substrates for Pi-class glutathione S-transferase (GSTP1) and their biological application.

PubMed

Fujikawa, Yuuta; Nampo, Taiki; Mori, Masaya; Kikkawa, Manami; Inoue, Hideshi

2018-03-01

Pi class glutathione S-transferase (GSTP1) is highly expressed in various cancerous cells and pre-neoplastic legions, where it is involved in apoptotic resistance or metabolism of several anti-tumour chemotherapeutics. Therefore, GSTP1 is a marker of malignant and pre-malignant cells and is a promising target for visualization and drug development. Here we demonstrate that fluorescein diacetate (FDA), a fluorescent probe used for vital staining, is a fluorescently activated by esterolytic activity of human GSTP1 (hGSTP1) selectively among various cytosolic GSTs. Fluorescence activation of FDA susceptible to GST inhibitors was observed in MCF7 cells exogenously overexpressing hGSTP1, but not in cells overexpressing hGSTA1 or hGSTM1. Inhibitor-sensitive fluorescence activation was also observed in several cancer cell lines endogenously expressing GSTP1, suggesting that GSTP1 is involved in FDA esterolysis in these cells. Among the FDA derivatives examined, FOMe-Ac, the acetyl ester of fluorescein O-methyl ether, was found to be a potential reporter for GSH-dependent GSTP1 activity as well as for carboxylesterase activity. Since GSTP1 is highly expressed in various types of cancer cells compared to their normal counterparts, improving the fluorogenic substrates to be more selective to the esterolysis activity of GSTP1 rather than carboxylesterases should lead to development of tools for detecting GSTP1-overexpressing cancer cells and investigating the biological functions of GSTP1. Copyright © 2017 Elsevier B.V. All rights reserved.

A Simple Strategy in Avulsion Flap Injury: Prediction of Flap Viability Using Wood's Lamp Illumination and Resurfacing with a Full-thickness Skin Graft.

PubMed

Lim, Hyoseob; Han, Dae Hee; Lee, Il Jae; Park, Myong Chul

2014-03-01

Extensive degloving injuries of the extremities usually result in necrosis of the flap, necessitating comprehensive skin grafting. Provided there is a sufficient tool to evaluate flap viability, full-thickness skin can be used from a nonviable avulsed flap. We used a Wood's lamp to determine the viability of avulsed flaps in the operation field after intravenous injection of fluorescein dye. We experienced 13 cases during 16 months. Fifteen minutes after the intravenous injection of fluorescein dye, the avulsed skin flaps were examined and non-fluorescent areas were marked under Wood's lamp illumination. The marked area was defatted for full-thickness skin grafting. The fluorescent areas were sutured directly without tension. The non-fluorescent areas were covered by defatted skin. Several days later, there was soft tissue necrosis within the flap area. We measured necrotic area and revised the flap. Among all the cases, necrotic area was 21.3% of the total avulsed area. However, if we exclude three cases, one of a carelessly managed patient and two cases of the flaps were inappropriately applied, good results were obtained, with a necrotic area of only 8.4%. Eight patients needed split-thickness skin grafts, and heel pad reconstruction was performed with free flap. A full-thickness skin graft from an avulsed flap is a good method for addressing aesthetic concerns without producing donor site morbidity. Fluorescein dye is a useful, simple, and cost-effective tool for evaluating flap viability. Avulsed flap injuries can be managed well with Wood's lamp illumination and a full-thickness skin graft.

Green fiber lasers: An alternative to traditional DPSS green lasers for flow cytometry

PubMed Central

Telford, William G.; Babin, Sergey A.; Khorev, Serge V.; Rowe, Stephen H.

2009-01-01

Green and yellow diode-pumped solid state (DPSS) lasers (532 and 561 nm) have become common fixtures on flow cytometers, due to their efficient excitation of phycoerythrin (PE) and its tandems, and their ability to excite an expanding array of expressible red fluorescent proteins. Nevertheless, they have some disadvantages. DPSS 532 nm lasers emit very close to the fluorescein bandwidth, necessitating optical modifications to permit detection of fluorescein and GFP. DPSS 561 nm lasers likewise emit very close to the PE detection bandwidth, and also cause unwanted excitation of APC and its tandems, requiring high levels of crossbeam compensation to reduce spectral overlap into the PE tandems. In this paper, we report the development of a new generation of green fiber lasers that can be engineered to emit in the range between 532 and 561 nm. A 550 nm green fiber laser was integrated into both a BD LSR II™ cuvette and FACSVantage DiVa™ jet-in-air cell sorter. This laser wavelength avoided both the fluorescein and PE bandwidths, and provided better excitation of PE and the red fluorescent proteins DsRed and dTomato than a power-matched 532 nm source. Excitation at 550 nm also caused less incidental excitation of APC and its tandems, reducing the need for crossbeam compensation. Excitation in the 550 nm range therefore proved to be a good compromise between 532 and 561 nm sources. Fiber laser technology is therefore providing the flexibility necessary for precisely matching laser wavelengths to our flow cytometry applications. PMID:19777600

Validation of an automated fluorescein method for determining bromide in water

USGS Publications Warehouse

Fishman, M. J.; Schroder, L.J.; Friedman, L.C.

1985-01-01

Surface, atmospheric precipitation and deionized water samples were spiked with ??g l-1 concentrations of bromide, and the solutions stored in polyethylene and polytetrafluoroethylene bottles. Bromide was determined periodically for 30 days. Automated fluorescein and ion chromatography methods were used to determine bromide in these prepared samples. Analysis of the data by the paired t-test indicates that the two methods are not significantly different at a probability of 95% for samples containing from 0.015 to 0.5 mg l-1 of bromide. The correlation coefficient for the same sets of paired data is 0.9987. Recovery data, except for the surface water samples to which 0.005 mg l-1 of bromide was added, range from 89 to 112%. There appears to be no loss of bromide from solution in either type of container.Surface, atmospheric precipitation and deionized water samples were spiked with mu g l** minus **1 concentrations of bromide, and the solutions stored in polyethylene and polytetrafluoroethylene bottles. Bromide was determined periodically for 30 days. Automated fluorescein and ion chromatography methods were used to determine bromide in these prepared samples. Analysis of the data by the paired t-test indicates that the two methods are not significantly different at a probability of 95% for samples containing from 0. 015 to 0. 5 mg l** minus **1 of bromide. The correlation coefficient for the same sets of paired data is 0. 9987. Recovery data, except for the surface water samples to which 0. 005 mg l** minus **1 of bromide was added, range from 89 to 112%. Refs.

Clinical benefits of a new multipurpose disinfecting solution in silicone hydrogel and soft contact lens users.

PubMed

Campbell, Robert; Kame, Gregory; Leach, Norman; Paul, Matthew; White, Eric; Zigler, Lamar

2012-03-01

To evaluate the safety and efficacy of a new multipurpose disinfecting solution (MPDS) containing a diblock copolymer, poly(oxyethylene)-poly(oxybutylene), designed to improve the wetting properties of silicone-hydrogel lenses. This 90-day, randomized, concurrently controlled, double-masked, multisite study involved 573 subjects at 30 investigational sites in the United states. Existing lens wearers were randomly assigned to either regimen 1 (OPTI-FREE PureMoist, a newly developed formulation, Alcon Laboratories Inc., Fort Worth, TX) or regimen 2 (renu fresh, Bausch and Lomb Inc., Rochester, NY). At baseline, days 7, 30, 60, and 90, the subjects recorded subjective evaluations relating to lens acceptability and comfort, wearing time, and rewetting drop frequency, whereas investigators assessed slitlamp findings (including corneal fluorescein staining), lens deposits (modified Rudko), visual acuity, and adverse events. Group IV lenses were collected for ex vivo measurement of lysozyme deposits. Differences favoring regimen 1 were noted on day 90 for subjective evaluations of lens comfort (insertion, removal, overall); lens moisture and freshness; gentle on eye; clear vision; and lens handling (all P<0.004). Corneal fluorescein staining severity and area and lysozyme deposits (group IV lenses) were lower with regimen 1 (P<0.0001). This 90-day study shows that the new MPDS designed for silicone-hydrogel lenses is safe and effective when used by both silicone-hydrogel and traditional soft lens wearers. By improving comfort and decreasing lysozyme deposits and corneal fluorescein staining, the new MPDS enhances patients' wearing experience and helps maintain optimal lens performance.

Treatment of Sjögren's syndrome dry eye using 0.03% tacrolimus eye drop: Prospective double-blind randomized study.

PubMed

Moscovici, Bernardo Kaplan; Holzchuh, Ricardo; Sakassegawa-Naves, Fernando Eiji; Hoshino-Ruiz, Diego Ricardo; Albers, Marcos Bottene Villa; Santo, Ruth Miyuki; Hida, Richard Yudi

2015-10-01

To describe the clinical efficacy of the treatment of Sjögren's syndrome dry eye using 0.03% tacrolimus eye drop. Prospective double-blind randomized study. Institutional outpatient clinic. Forty-eight eyes of twenty-four patients with dry eye related to Sjögren syndrome were enrolled in this study. The patients were randomized in 2 groups: tacrolimus (n=14) and vehicle (n=10) group. The tacrolimus group received a vial containing tacrolimus 0.03% (almond oil as vehicle) and the other group received the almond oil vehicle. All patients were instructed to use the eye drops every 12h in the lower conjunctival sac. Schirmer I test, break-up-time (BUT), corneal fluorescein and Rose Bengal staining scores were evaluated in all patients one day before the treatment (baseline), 7, 14, 28 and 90 days after treatment with the eye drops. The average fluorescein and Rose Bengal scores improved statistically after 7 days of treatment and even more after 90 days. The average Schirmer I and BUT values were unchanged after 7, 14 and 21 days but did show an improvement relative to baseline after 28 days of treatment. Schirmer I, BUT, fluorescein and Rose Bengal did not show any statistical significance in the vehicle group. Topical 0.03% tacrolimus eye drop improved tear stability and ocular surface status in cases of inflammatory or SS-related dry eye. ClinicalTrials.gov Identifier: NCT01850979. Copyright © 2015 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

Antibody-mediated fluorescence enhancement based on shifting the intramolecular dimer<-->monomer equilibrium of fluorescent dyes.

PubMed

Wei, A P; Blumenthal, D K; Herron, J N

1994-05-01

A novel concept is described for directly coupling fluorescence emission to protein-ligand binding. It is based on shifting the intramolecular monomer<-->dimer equilibrium of two fluorescent dyes linked by a short spacer. A 13-residue peptide, recognized by a monoclonal antibody against human chorionic gonadotrophin (hCG), was labeled with fluorescein (F) and tetramethylrhodamine (T) at its N- and C-terminus, respectively. Spectral evidence suggests that when the conjugate is free in solution, F and T exist as an intramolecular dimer. Fluorescence quenching of fluorescein and rhodamine is approximately 98% and approximately 90%, respectively, due to dimerization. When the double-labeled peptide is bound to anti-hCG, however, the rhodamine fluorescence increases by up to 7.8-fold, depending upon the excitation wavelength. This is attributed to the dissociation of intramolecular dimers brought about by conformational changes of the conjugate upon binding. Fluorescein fluorescence, on the other hand, was still quenched because of excited-state energy transfer and residual ground-state interactions. Antibody binding also resulted in a approximately 3.4-fold increase in fluorescence anisotropy of the peptide. These changes in intensity and anisotropy allow direct measurement of antigen-antibody binding with a fluorescence plate reader or a polarization analyzer, without the need for separation steps and labeling antibodies. Because recent advances in peptide technology have allowed rapid and economical identification of antigen-mimicking peptides, the double-labeled peptide approach offers many opportunities for developing new diagnostic assays and screening new therapeutic drugs. It also has many potential applications to techniques involving recombinant antibodies, biosensors, cell sorting, and DNA probes.

Aqueous Angiography–Mediated Guidance of Trabecular Bypass Improves Angiographic Outflow in Human Enucleated Eyes

PubMed Central

Huang, Alex S.; Saraswathy, Sindhu; Dastiridou, Anna; Begian, Alan; Mohindroo, Chirayu; Tan, James C. H.; Francis, Brian A.; Hinton, David R.; Weinreb, Robert N.

2016-01-01

Purpose To assess the ability of trabecular micro-bypass stents to improve aqueous humor outflow (AHO) in regions initially devoid of AHO as assessed by aqueous angiography. Methods Enucleated human eyes (14 total from 7 males and 3 females [ages 52–84]) were obtained from an eye bank within 48 hours of death. Eyes were oriented by inferior oblique insertion, and aqueous angiography was performed with indocyanine green (ICG; 0.4%) or fluorescein (2.5%) at 10 mm Hg. With an angiographer, infrared and fluorescent images were acquired. Concurrent anterior segment optical coherence tomography (OCT) was performed, and fixable fluorescent dextrans were introduced into the eye for histologic analysis of angiographically positive and negative areas. Experimentally, some eyes (n = 11) first received ICG aqueous angiography to determine angiographic patterns. These eyes then underwent trabecular micro-bypass sham or stent placement in regions initially devoid of angiographic signal. This was followed by fluorescein aqueous angiography to query the effects. Results Aqueous angiography in human eyes yielded high-quality images with segmental patterns. Distally, angiographically positive but not negative areas demonstrated intrascleral lumens on OCT images. Aqueous angiography with fluorescent dextrans led to their trapping in AHO pathways. Trabecular bypass but not sham in regions initially devoid of ICG aqueous angiography led to increased aqueous angiography as assessed by fluorescein (P = 0.043). Conclusions Using sequential aqueous angiography in an enucleated human eye model system, regions initially without angiographic flow or signal could be recruited for AHO using a trabecular bypass stent. PMID:27588614

Development of and Clinical Experience with a Simple Device for Performing Intraoperative Fluorescein Fluorescence Cerebral Angiography: Technical Notes.

PubMed

Ichikawa, Tsuyoshi; Suzuki, Kyouichi; Watanabe, Yoichi; Sato, Taku; Sakuma, Jun; Saito, Kiyoshi

2016-01-01

To perform intraoperative fluorescence angiography (FAG) under a microscope without an integrated FAG function with reasonable cost and sufficient quality for evaluation, we made a small and easy to use device for fluorescein FAG (FAG filter). We investigated the practical use of this FAG filter during aneurysm surgery, revascularization surgery, and brain tumor surgery. The FAG filter consists of two types of filters: an excitatory filter and a barrier filter. The excitatory filter excludes all wavelengths except for blue light and the barrier filter passes long waves except for blue light. By adding this FAG filter to a microscope without an integrated FAG function, light from the microscope illuminating the surgical field becomes blue, which is blocked by the barrier filter. We put the FAG filter on the objective lens of the operating microscope correctly and fluorescein sodium was injected intravenously or intra-arterially. Fluorescence (green light) from vessels in the surgical field and the dyed tumor were clearly observed through the microscope and recorded by a memory device. This method was easy and could be performed in a short time (about 10 seconds). Blood flow of small vessels deep in the surgical field could be observed. Blood flow stagnation could be evaluated. However, images from this method were inferior to those obtained by currently commercially available microscopes with an integrated FAG function. In brain tumor surgery, a stained tumor on the brain surface could be observed using this method. FAG could be performed with a microscope without an integrated FAG function easily with only this FAG filter.

Development of and Clinical Experience with a Simple Device for Performing Intraoperative Fluorescein Fluorescence Cerebral Angiography: Technical Notes

PubMed Central

ICHIKAWA, Tsuyoshi; SUZUKI, Kyouichi; WATANABE, Yoichi; SATO, Taku; SAKUMA, Jun; SAITO, Kiyoshi

2016-01-01

To perform intraoperative fluorescence angiography (FAG) under a microscope without an integrated FAG function with reasonable cost and sufficient quality for evaluation, we made a small and easy to use device for fluorescein FAG (FAG filter). We investigated the practical use of this FAG filter during aneurysm surgery, revascularization surgery, and brain tumor surgery. The FAG filter consists of two types of filters: an excitatory filter and a barrier filter. The excitatory filter excludes all wavelengths except for blue light and the barrier filter passes long waves except for blue light. By adding this FAG filter to a microscope without an integrated FAG function, light from the microscope illuminating the surgical field becomes blue, which is blocked by the barrier filter. We put the FAG filter on the objective lens of the operating microscope correctly and fluorescein sodium was injected intravenously or intra-arterially. Fluorescence (green light) from vessels in the surgical field and the dyed tumor were clearly observed through the microscope and recorded by a memory device. This method was easy and could be performed in a short time (about 10 seconds). Blood flow of small vessels deep in the surgical field could be observed. Blood flow stagnation could be evaluated. However, images from this method were inferior to those obtained by currently commercially available microscopes with an integrated FAG function. In brain tumor surgery, a stained tumor on the brain surface could be observed using this method. FAG could be performed with a microscope without an integrated FAG function easily with only this FAG filter. PMID:26597335

Sodium Fluorescein-Guided Resection under the YELLOW 560 nm Surgical Microscope Filter in Malignant Gliomas: Our First 38 Cases Experience

PubMed Central

Tian, Hailong; Huang, Dezhang; Meng, Xianbing; Guo, Wenqiang; Wang, Chaochao; Yin, Xin; Zhang, Hongying; Jiang, Bin; He, Zheng

2017-01-01

Objective Sodium fluorescein (FL) had been safely used in fluorescence-guided microsurgery for imaging various brain tumors. Under the YELLOW 560 nm surgical microscope filter, low-dose FL as a fluorescent dye helps in visualization. Our study investigated the safety and efficacy of this innovative technique in malignant glioma (MG) patients. Patients and Method 38 patients suffering from MGs confirmed by pathology underwent FL-guided resection under YELLOW 560 nm surgical microscope filter. We retrospectively analyzed the clinical characters, microsurgery procedure, extent of resection, pathology of MGs, progression-free survival (PFS), and overall survival (OS). Results Thirty-eight patients had MGs (10 WHO grade III, 28 WHO grade IV). With YELLOW 560 nm surgical microscope filter combined with neuronavigation, sodium fluorescein-guided gross total resection (GTR) was achieved in 35 (92.1%) patients and subtotal resection in 3 (7.69%). The sensitivity and specificity of FL were 94.4% and 88.6% regardless of radiographic localization. Intraoperatively, 10 biopsies (10/28 FL[+]) showed “low” or “high” fluorescence in non-contrast-enhancement region and are also confirmed by pathology. Our data showed 6-month PFS of 92.3% and median survival of 11 months. Conclusion FL-guided resection of MGs under the YELLOW 560 nm surgical microscope filter combined with neuronavigation was safe and effective, especially in non-contrast-MRI regions. It is feasible for improving the extent of resection in MGs especially during emergency cases. PMID:29124069

Intravenous Bevacizumab Causes Regression of Choroidal Neovascularization Secondary to Diseases Other Than Age-related Macular Degeneration

PubMed Central

NGUYEN, QUAN DONG; SHAH, SYED MAHMOOD; HAFIZ, GULNAR; DO, DIANA V.; HALLER, JULIA A.; PILI, ROBERTO; ZIMMER-GALLER, INGRID E.; JANJUA, KASHIF; SYMONS, R. C. ANDREW; CAMPOCHIARO, PETER A.

2016-01-01

PURPOSE To investigate the safety, tolerability, and bioactivity of intravenous infusions of bevacizumab in patients with choroidal neovascularization (CNV) attributable to causes other than age-related macular degeneration. DESIGN Nonrandomized clinical trial. METHODS Ten patients with CNV received infusions of 5 mg/kg of bevacizumab. The primary efficacy outcome measure was change in visual acuity (VA; Early Treatment Diabetic Retinopathy Study letters read at 4 meters) at 24 weeks and secondary measures were changes from baseline in excess foveal thickness (center subfield thickness), area of fluorescein leakage, and area of CNV. RESULTS Infusions were well tolerated and there were no ocular or systemic adverse events. At baseline, median VA was 25.5 letters read at 4 meters (20/80) and median foveal thickness was 346 μm. At the primary endpoint (24 weeks), median VA was 48.5 letters (20/32), representing four lines of improvement from baseline (P = .005), median foveal thickness was 248 μm representing a 72% reduction in excess foveal thickness (P = .007). Four of nine patients had complete elimination of fluorescein leakage, three had near complete elimination (reductions of 91%, 88%, and 87%), two had modest reductions, and one had no reduction. All patients except one showed a reduction in area of CNV with a median reduction of 43%. CONCLUSIONS Despite the small number of patients studied, the marked improvement in VA accompanied by prominent reductions in foveal thickness, fluorescein leakage, and area of CNV suggest a beneficial effect. It may be worthwhile to consider further evaluation of systemic bevacizumab in young patients with CNV. PMID:18054887

Direct Measurement of Acetylesterase in Living Protist Cells1

PubMed Central

Medzon, Edward L.; Brady, Marilyn L.

1969-01-01

The fluorogenic acetylesterase (acetic ester hydrolase EC 3.1.1.6.) substrate, fluorescein diacetate, was used to measure enzyme activity in living protist cells. The visual enzyme assay was done by monitoring fluorochromasia by fluorescent microscopy. Quantitative fluorogenic assays were done by measuring the evolved fluorescein in a fluorometer. Of 59 strains of bacteria, 35 were fluorochromatically positive. Eight of the fluorochromatically negative strains were fluorogenically positive. Of 22 strains of slime molds and fungi, all were fluorochromatically positive. Three out of 12 different algae were fluorochromatically positive. Several unidentified protozoa were also fluorochromatically positive. Four out of six protozoa were fluorochromatically positive. Structures of special interest showing acetylesterase activity were: the growing hyphal tips of fungi, the vacuolated areas of yeast and protozoa, newly formed bacterial spores or immature fungal spores, “mesosome-like” bodies in Bacillus megaterium, and the cell membrane and nuclear region of green algae. Yeast protoplasts and bacterial protoplasts and spheroplasts were fluorochromatically positive when derived from positive cells and negative when derived from negative cells. There was no correlation between the possession of a capsule and acetylesterase activity. There was no effect on the viability of bacterial cells incubated in the presence of fluorescein diacetate. Paraoxon inhibited bacterial and yeast enzyme at 10−5m. Eserine (10−5m) and Paraoxon (10−7m) inhibited B. megaterium enzyme. Sodium acetate at 10−2m did not inhibit bacterial enzyme. The implications of these findings on the location and expression of esterase activity in living cells are discussed. Images PMID:4974398

PREPARATION AND CHARACTERIZATION OF MOLECULARLY IMPRINTED ELECTROPOLYMERIZED CARBON ELECTRODES

EPA Science Inventory

Molecularly imprinted polymers (MIP) selective for fluorescein, rhodamine or 2,4-dichlorophenoxyacetic acid (2,4-D) were electropolymerized onto graphite electrodes using an aqueous solution equimolar in resorsinol/ortho-phenylenediamine and in the presence of the template mole...

Cellular uptake of glucoheptoamidated poly(amidoamine) PAMAM G3 dendrimer with amide-conjugated biotin, a potential carrier of anticancer drugs.

PubMed

Uram, Łukasz; Szuster, Magdalena; Filipowicz, Aleksandra; Zaręba, Magdalena; Wałajtys-Rode, Elżbieta; Wołowiec, Stanisław

2017-01-15

In search for soluble derivatives of PAMAM dendrimers as potential carriers for hydrophobic drugs, the conjugates of PAMAM G3 with biotin, further converted into glycodendrimer with d-glucoheptono-1,4-lactone, were prepared. Polyamidoamine dendrimer (PAMAM) of third generation, G3 was functionalized with four biotin equivalents covalently attached to terminal amine nitrogens via amide bond G3 4B . The remaining 28 amine groups were blocked by glucoheptoamide substituents (gh) to give G3 4B28gh or with one fluorescein equivalent (attached by reaction of G3 4B with fluorescein isothiocyanate, FITC) via thiourea bond as FITC followed by exhaustive glucoheptoamidation to get G3 4B27gh1F . As a control the G3 substituted totally with 32 glucoheptoamide residues, G3 gh and its fluorescein labeled analogue G3 31gh1F were synthesized. The glucoheptoamidation of PAMAM G0 dendrimer with glucoheptono-1,4-lactone was performed in order to fully characterize the 1 H NMR spectra of glucoheptoamidated PAMAM dendrimers and to control the derivatization of G3 with glucoheptono-1,4-lactone. Another two derivatives of G3, namely G3 4B28gh1F' and G3 32ghF' , with ester bonded fluorescein were also obtained. Biological properties of obtained dendrimer conjugates were estimated in vitro with human cell lines: normal fibroblast (BJ) and two cancer glioblastoma (U-118 MG) and squamous carcinoma (SCC-15), including cytotoxicity by reduction of XTT and neutral red (NR) assays. Cellular uptake of dendrimer conjugates was evaluated with confocal microscopy. Obtained results confirmed, that biotinylated bioconjugates have always lower cytotoxicity and 3-4 times higher cellular uptake than non-biotinylated dendrimer conjugates in all cell lines. Comparison of various cell lines revealed different dose-dependent cell responses and the lower cytotoxicity of examined dendrimer conjugates for normal fibroblasts and squamous carcinoma, as compared with much higher cytotoxic effects seen in glioblastoma cell line. Synthetized multi-functional conjugate (G3 4B27gh1F ) is a promising candidate as biocompatible vehicle for hydrophobic molecules used in anticancer therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fluorescence Immunofiltration Assay of Brucella Melitensis.

DTIC Science & Technology

1995-01-01

second urease -labelled antibody directed against fluorescein. The assay system is useful for measuring protein, virus and bacteria in aqueous...binding site for the signal-generating urease -labelled antibody, it is a highly fluorescent molecule and has signal-generating capacity of its own

MULTIMODAL IMAGING IN VORTEX VEIN VARICES.

PubMed

Veronese, Chiara; Staurenghi, Giovanni; Pellegrini, Marco; Maiolo, Chiara; Primavera, Laura; Morara, Mariachiara; Armstrong, Grayson W; Ciardella, Antonio P

2017-03-22

The aim of this study is to describe the clinical presentation of vortex vein varices with multimodal imaging. The authors carried out a retrospective case series of eight patients (7 female, 1 male) with an average age of 60.2 years (min 8, max 84, median 68.5) presenting with vortex vein varices. All patients were evaluated at the Sant'Orsola-Malpighi Hospital, University of Bologna, Bologna, Italy and at Luigi Sacco Hospital, University of Milan, Milan, Italy. Patients underwent complete ophthalmologic examinations, including best corrected visual acuity, intraocular pressure, anterior segment, and fundus examination. Imaging studies, including fundus color photography, near-infrared reflectance imaging, fundus autofluorescence, fluorescein angiography, indocyanine green angiography, and spectral-domain enhanced depth imaging optical coherence tomography were also performed. Ultra-widefield fluorescein angiography and ultra-widefield indocyanine angiography using the Heidelberg Retina Angiograph and the Staurenghi 230 SLO Retina Lens were used to demonstrate the disappearance of all retinal lesions when pressure was applied to the globe. All eight cases initially presented to the emergency room. One patient presented secondary to trauma, two patients presented for suspected hemangioma, whereas the other five were referred to the authors' hospitals for suspected retinal lesions. On examination, retinal abnormalities were identified in all 8 patients, with 7 (87.5%) oculus dexter and 1 (12.5%) oculus sinister, and with 1 (12.5%) inferotemporally, 3 (37.5%) superonasally, 3 (37.5%) inferonasally, and 1 (12.5%) inferiorly. Fundus color photography showed an elevated lesion in seven patients and a nonelevated red lesion in one patient. In all patients, near-infrared reflectance imaging showed a hyporeflective lesion in the periphery of the retina. Fundus autofluorescence identified round hypofluorescent rings surrounding weakly hyperfluorescent lesions in all patients. On fluorescein angiography, all lesions were initially hyperfluorescent with a hypofluorescent ring, with the lesion becoming hyperfluorescent after injection of dye. Indocyanine green angiography demonstrated dilation of the vortex vein ampullae in all patients. Spectral-domain enhanced depth imaging optical coherence tomography demonstrated dilated choroidal vessels and a hyporeflective cavity without subretinal fluid in all patients. Ultra-widefield fluorescein angiography and ultra-widefield indocyanine angiography demonstrated disappearance of all retinal lesions when pressure was applied to the globe. Findings are consistent with the diagnosis of vortex vein varix in all eight patients, with six patients (75%) exhibiting a single varix and two patients (25%) exhibiting a double varix. The diagnosis of vortex vein varices can be confirmed through clinical examination through the use of digital pressure to the globe during ophthalmoscopic examination. Adjunctive multimodal imaging (fundus color photography, near-infrared reflectance imaging, fundus autofluorescence, fluorescein angiography, indocyanine angiography, and spectral-domain enhanced depth imaging optical coherence tomography) was useful in the diagnosis of vortex vein varices in the authors' clinical cases. However, in more challenging clinical cases, the authors' novel use of the ultra-widefield contact lens for application of ocular pressure with a resulting resolution of the varix proved to be a useful and easy diagnostic imaging method for confirming the presence of vortex vein varices.

ORGANOPHOSPHORUS HYDROLASE-BASED ASSAY FOR ORGANOPHOSPHATE PESTICIDES

EPA Science Inventory

We report a rapid and versatile Organophosphorus hydrolase (OPH)-based method for measurement of organophosphates. This assay is based on a substrate-dependent change in pH at the local vicinity of the enzyme. The pH change is monitored using fluorescein isothiocyanate (FITC), ...

Evaluation of diminished microbial contamination in handling of a novel daily disposable flat pack contact lens.

PubMed

Nomachi, Miya; Sakanishi, Kotaro; Ichijima, Hideji; Cavanagh, H Dwight

2013-05-01

To evaluate the efficacy of a novel daily disposable (DD) flat package in regard to microbial contamination on the anterior and posterior surfaces of a contact lens (CL) during handling of the lens for insertion. Four kinds of commercially available general blister-packed daily disposable contact lenses (DD CLs) as controls and a novel Magic 1-day Menicon Flat Pack as a test lens were used for this in vitro study. Lenses were removed from their packages using fingers coated with fluorescein 3 to 5 μm beads or an approximately 7×10(2) to 2×10(3) colony-forming unit (CFU)/mL Staphylococcus aureus suspension. The transfer of fluorescein beads to the surface of the lenses was then observed by fluorescence microscopy. Microbial contamination on the lenses was observed by light microscopy after a 2-day incubation period; and, the number of colonies isolated from the contaminated lenses was determined after 4 days of incubation. The number of fluorescein beads on the Magic lens was significantly less (p<0.05) than that of the general blister-packed control lenses. Adherence of microbial colonies was observed on both inner and outer surfaces of general blister-packed lenses, whereas no colony formation was found on the inner surface of the Magic lens, and the lowest bacterial adherence was observed for the Magic lens. The data demonstrated that placement of the Magic DD lens onto the eye is accompanied by diminished microbial contamination compared with general blister-packed DD CLs. Eye care professionals; however, should instruct patients to comply with intended use of DD CLs to prevent CL-associated microbial keratitis. In all cases, hand washing is mandated prelens insertion.

Color Fundus Photography versus Fluorescein Angiography in Identification of the Macular Center and Zone in Retinopathy of Prematurity

PubMed Central

Patel, Samir N.; Klufas, Michael A.; Ryan, Michael C.; Jonas, Karyn E.; Ostmo, Susan; Martinez-Castellanos, Maria Ana; Berrocal, Audina M.; Chiang, Michael F.; Chan, R.V. Paul

2016-01-01

Purpose To examine the utility of fluorescein angiography (FA) in identification of the macular center and the diagnosis of zone in patients with retinopathy of prematurity (ROP). Design Validity and reliability analysis of diagnostic tools Methods 32 sets (16 color fundus photographs; 16 color fundus photographs paired with the corresponding FA) of wide-angle retinal images obtained from 16 eyes of eight infants with ROP were compiled on a secure web site. 9 ROP experts (3 pediatric ophthalmologists; 6 vitreoretinal surgeons) participated in the study. For each image set, experts identified the macular center and provided a diagnosis of zone. Main Outcome Measures (1) Sensitivity and specificity of zone diagnosis (2) “Computer facilitated diagnosis of zone,” based on precise measurement of the macular center, optic disc center, and peripheral ROP. Results Computer facilitated diagnosis of zone agreed with the expert’s diagnosis of zone in 28/45 (62%) cases using color fundus photographs and in 31/45 (69%) cases using FA. Mean (95% CI) sensitivity for detection of zone I by experts as compared to a consensus reference standard diagnosis when interpreting the color fundus images alone versus interpreting the color fundus photographs and FA was 47% (35.3% – 59.3%) and 61.1% (48.9% – 72.4%), respectively, (t(9) ≥ (2.063), p = 0.073). Conclusions There is a marginally significant difference in zone diagnosis when using color fundus photographs compared to using color fundus photographs and the corresponding fluorescein angiograms. There is inconsistency between traditional zone diagnosis (based on ophthalmoscopic exam and image review) compared to a computer-facilitated diagnosis of zone. PMID:25637180

Flow cytometric detection of human immunodeficiency virus type 1 proviral DNA by the polymerase chain reaction incorporating digoxigenin- or fluorescein-labeled dUTP

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

Yang, Gang; Olson, J.C.; Pu, R.

1995-10-01

Serological assays are routinely used in the laboratory diagnosis of human immunodeficiency virus type-1 (HrV-1) infection, but the polymerase chain reaction (PCR) is ultimately the most sensitive and direct method for establishing definitive diagnosis. As an alternative to the conventional radioactive PCR procedure we have developed and evaluated a pair of rapid nonradioisotopic flow cytometric detection methods. Using heminested PCR we directly incorporated fluorescein-12-dUTP (fluo-dUTP) or digoxigenin-11-dUTP (dig-dUTP) into the PCR-amplicons. The labeled amplicons were hybridized with biotinylated antisense and sense probes, followed by capture of the hybrid DNA using streptavidin-coated beads which were finally analyzed in a flow cytometermore » by (1) direct detection of the fluorescence intensity of the amplicons incorporating fluo-dUTP and (2) immunodetection of the amplicons incorporating dig-dUTP by anti-digoxigenin IgG labeled with fluorescein isothiocyanate (FITC). Although both assays were functionally comparable with radiolabeled probe in reliably detecting as low as five copies of HIV-1 proviral DNA sequences, the immunodetection of dig-dUTP consistently yielded higher mean channel fluorescence and gave a stable signal over an extended period of 12-14 weeks. In testing a panel of 20 pedigreed PBMC specimens from blood donors with or without HIV-1 infection, the results of both flow cytometric assays were identical with those of the conventional radioactive procedure. Therefore, we conclude that the dig-dUTP incorporation in amplicons, hybridization with a pair of sense-antisense biotinylated probes and immunodetection of hybrids by flow cytometric analyses is the nonisotopic method of choice for PCR-diagnosis of HIV-1 infection. 21 refs., 2 figs., 4 tabs.« less

COMPARISON OF THREE TRACER TESTS AT THE RAFT RIVER GEOTHERMAL SITE

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

Earl D Mattson; Mitchell Plummer; Carl Palmer

2011-02-01

Three conservative tracer tests have been conducted through the Bridge Fault fracture zone at the Raft River Geothermal (RRG) site. All three tests were conducted between injection well RRG-5 and production wells RRG-1 (790 m distance) and RRG-4 (740 m distance). The injection well is used during the summer months to provide pressure support to the production wells. The first test was conducted in 2008 using 136 kg of fluorescein tracer. Two additional tracers were injected in 2010. The first 2010 tracer injected was 100 kg fluorescein disodium hydrate salt on June, 21. The second tracer (100 kg 2,6-naphthalene disulfonicmore » acid sodium salt) was injected one month later on July 21. Sampling of the two productions wells is still being performed to obtain the tail end of the second 2010 tracer test. Tracer concentrations were measured using HPLC with a fluorescence detector. Results for the 2008 test, suggest 80% tracer recover at the two production wells. Of the tracer recovered, 85% of tracer mass was recovered in well RRG-4 indicating a greater flow pathway connection between injection well and RRG-4 than RRG-1. Fluorescein tracer results appear to be similar between the 2008 and 2010 tests for well RRG-4 with peak concentrations arriving approximately 20 days after injection despite the differences between the injection rates for the two tests (~950 gpm to 475 gpm) between the 2008 and 2010. The two 2010 tracer tests will be compared to determine if the results support the hypothesis that rock contraction along the flow pathway due to the 55 oC cooler water injection alters the flow through the ~140 oC reservoir.« less

Fluorescein permeability and electrical resistance of human skin during low frequency ultrasound application.

PubMed

Cancel, Limary M; Tarbell, John M; Ben-Jebria, Abdellaziz

2004-09-01

Transdermal drug delivery offers an alternative to injections and oral medication but is limited by the low skin permeability of most drugs. The use of low-frequency ultrasound over long periods of time, typically over an hour, has been shown to enhance skin permeability, a phenomenon referred to as sonophoresis. In this study, we investigated the effects of short time sonication of human skin at 20 kHz and at variable intensities and duty cycles on the dynamics of fluorescein transport across the skin (permeability) as well as the changes in the skin's structural integrity (electrical resistance). We found that a short application of ultrasound enhanced the transport of fluorescein across human skin by a factor in the range of 2-9 for full thickness skin samples and by a factor in the range of 2-28 000 for heat-stripped stratum corneum samples (however, samples with very high (10(3)) enhancement were likely to have been damaged by ultrasound). The electrical resistance of the skin decreased by an average of 20% for full thickness samples and 58% for stratum corneum samples. Increasing the duty cycle from 10 to 60% caused a significant increase in permeability enhancement from 2.3 to 9.1, and an increase in intensity from 8 to 23 mW cm(-2) induced a significant increase in permeability enhancement from 2 to 7.4, indicating a clear dependence of the permeability on both duty cycle and intensity. The increase in solute flux upon ultrasound exposure was immediate, demonstrating for the first time the fast response dynamics of sonophoretic enhancement. In addition, a quantitative analysis of the thermal and convective dispersion effects associated with ultrasound application showed that each contributes significantly to the overall permeability enhancement observed.

The effectiveness and cost-effectiveness of intraoperative imaging in high-grade glioma resection; a comparative review of intraoperative ALA, fluorescein, ultrasound and MRI.

PubMed

Eljamel, M Sam; Mahboob, Syed Osama

2016-12-01

Surgical resection of high-grade gliomas (HGG) is standard therapy because it imparts significant progression free (PFS) and overall survival (OS). However, HGG-tumor margins are indistinguishable from normal brain during surgery. Hence intraoperative technology such as fluorescence (ALA, fluorescein) and intraoperative ultrasound (IoUS) and MRI (IoMRI) has been deployed. This study compares the effectiveness and cost-effectiveness of these technologies. Critical literature review and meta-analyses, using MEDLINE/PubMed service. The list of references in each article was double-checked for any missing references. We included all studies that reported the use of ALA, fluorescein (FLCN), IoUS or IoMRI to guide HGG-surgery. The meta-analyses were conducted according to statistical heterogeneity between studies. If there was no heterogeneity, fixed effects model was used; otherwise, a random effects model was used. Statistical heterogeneity was explored by χ 2 and inconsistency (I 2 ) statistics. To assess cost-effectiveness, we calculated the incremental cost per quality-adjusted life-year (QALY). Gross total resection (GTR) after ALA, FLCN, IoUS and IoMRI was 69.1%, 84.4%, 73.4% and 70% respectively. The differences were not statistically significant. All four techniques led to significant prolongation of PFS and tended to prolong OS. However none of these technologies led to significant prolongation of OS compared to controls. The cost/QALY was $16,218, $3181, $6049 and $32,954 for ALA, FLCN, IoUS and IoMRI respectively. ALA, FLCN, IoUS and IoMRI significantly improve GTR and PFS of HGG. Their incremental cost was below the threshold for cost-effectiveness of HGG-therapy, denoting that each intraoperative technology was cost-effective on its own. Copyright © 2016 Elsevier B.V. All rights reserved.

Fully Automatic Segmentation of Fluorescein Leakage in Subjects With Diabetic Macular Edema

PubMed Central

Rabbani, Hossein; Allingham, Michael J.; Mettu, Priyatham S.; Cousins, Scott W.; Farsiu, Sina

2015-01-01

Purpose. To create and validate software to automatically segment leakage area in real-world clinical fluorescein angiography (FA) images of subjects with diabetic macular edema (DME). Methods. Fluorescein angiography images obtained from 24 eyes of 24 subjects with DME were retrospectively analyzed. Both video and still-frame images were obtained using a Heidelberg Spectralis 6-mode HRA/OCT unit. We aligned early and late FA frames in the video by a two-step nonrigid registration method. To remove background artifacts, we subtracted early and late FA frames. Finally, after postprocessing steps, including detection and inpainting of the vessels, a robust active contour method was utilized to obtain leakage area in a 1500-μm-radius circular region centered at the fovea. Images were captured at different fields of view (FOVs) and were often contaminated with outliers, as is the case in real-world clinical imaging. Our algorithm was applied to these images with no manual input. Separately, all images were manually segmented by two retina specialists. The sensitivity, specificity, and accuracy of manual interobserver, manual intraobserver, and automatic methods were calculated. Results. The mean accuracy was 0.86 ± 0.08 for automatic versus manual, 0.83 ± 0.16 for manual interobserver, and 0.90 ± 0.08 for manual intraobserver segmentation methods. Conclusions. Our fully automated algorithm can reproducibly and accurately quantify the area of leakage of clinical-grade FA video and is congruent with expert manual segmentation. The performance was reliable for different DME subtypes. This approach has the potential to reduce time and labor costs and may yield objective and reproducible quantitative measurements of DME imaging biomarkers. PMID:25634978

Fully automatic segmentation of fluorescein leakage in subjects with diabetic macular edema.

PubMed

Rabbani, Hossein; Allingham, Michael J; Mettu, Priyatham S; Cousins, Scott W; Farsiu, Sina

2015-01-29

To create and validate software to automatically segment leakage area in real-world clinical fluorescein angiography (FA) images of subjects with diabetic macular edema (DME). Fluorescein angiography images obtained from 24 eyes of 24 subjects with DME were retrospectively analyzed. Both video and still-frame images were obtained using a Heidelberg Spectralis 6-mode HRA/OCT unit. We aligned early and late FA frames in the video by a two-step nonrigid registration method. To remove background artifacts, we subtracted early and late FA frames. Finally, after postprocessing steps, including detection and inpainting of the vessels, a robust active contour method was utilized to obtain leakage area in a 1500-μm-radius circular region centered at the fovea. Images were captured at different fields of view (FOVs) and were often contaminated with outliers, as is the case in real-world clinical imaging. Our algorithm was applied to these images with no manual input. Separately, all images were manually segmented by two retina specialists. The sensitivity, specificity, and accuracy of manual interobserver, manual intraobserver, and automatic methods were calculated. The mean accuracy was 0.86 ± 0.08 for automatic versus manual, 0.83 ± 0.16 for manual interobserver, and 0.90 ± 0.08 for manual intraobserver segmentation methods. Our fully automated algorithm can reproducibly and accurately quantify the area of leakage of clinical-grade FA video and is congruent with expert manual segmentation. The performance was reliable for different DME subtypes. This approach has the potential to reduce time and labor costs and may yield objective and reproducible quantitative measurements of DME imaging biomarkers. Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

Targeted administration into the suprachoroidal space using a microneedle for drug delivery to the posterior segment of the eye.

PubMed

Patel, Samirkumar R; Berezovsky, Damian E; McCarey, Bernard E; Zarnitsyn, Vladimir; Edelhauser, Henry F; Prausnitz, Mark R

2012-07-01

This study seeks to determine the intraocular pharmacokinetics of molecules and particles injected into the suprachoroidal space of the rabbit eye in vivo using a hollow microneedle. Suprachoroidal injections of fluorescein and fluorescently tagged dextrans (40 and 250 kDa), bevacizumab, and polymeric particles (20 nm to 10 μm in diameter) were performed using microneedles in New Zealand white rabbits. The fluorescence intensity within the eye was monitored in each animal using an ocular fluorophotometer to determine the distribution of the injected material in the eye over time as compared with intravitreal injection of fluorescein. Fundus photography and histology were performed as well. Molecules and particles injected near the limbus using a microneedle flowed circumferentially around the eye within the suprachoroidal space. By targeting the suprachoroidal space, the concentration of injected materials was at least 10-fold higher in the back of the eye tissues than in anterior tissues. In contrast, intravitreal injection of fluorescein targeted the vitreous humor with no significant selectivity for posterior versus anterior segment tissues. Half-lives in the suprachoroidal space for molecules of molecular weight from 0.3 to 250 kDa ranged from 1.2 to 7.9 hours. In contrast, particles ranging in size from 20 nm to 10 μm remained primarily in the suprachoroidal space and choroid for a period of months and did not clear the eye. No adverse effects of injection into the suprachoroidal space were observed. Injection into the suprachoroidal space using a microneedle offers a simple and minimally invasive way to target the delivery of drugs to the choroid and retina.

Targeted Administration into the Suprachoroidal Space Using a Microneedle for Drug Delivery to the Posterior Segment of the Eye

PubMed Central

Patel, Samirkumar R.; Berezovsky, Damian E.; McCarey, Bernard E.; Zarnitsyn, Vladimir; Edelhauser, Henry F.; Prausnitz, Mark R.

2012-01-01

Purpose. This study seeks to determine the intraocular pharmacokinetics of molecules and particles injected into the suprachoroidal space of the rabbit eye in vivo using a hollow microneedle. Methods. Suprachoroidal injections of fluorescein and fluorescently tagged dextrans (40 and 250 kDa), bevacizumab, and polymeric particles (20 nm to 10 μm in diameter) were performed using microneedles in New Zealand white rabbits. The fluorescence intensity within the eye was monitored in each animal using an ocular fluorophotometer to determine the distribution of the injected material in the eye over time as compared with intravitreal injection of fluorescein. Fundus photography and histology were performed as well. Results. Molecules and particles injected near the limbus using a microneedle flowed circumferentially around the eye within the suprachoroidal space. By targeting the suprachoroidal space, the concentration of injected materials was at least 10-fold higher in the back of the eye tissues than in anterior tissues. In contrast, intravitreal injection of fluorescein targeted the vitreous humor with no significant selectivity for posterior versus anterior segment tissues. Half-lives in the suprachoroidal space for molecules of molecular weight from 0.3 to 250 kDa ranged from 1.2 to 7.9 hours. In contrast, particles ranging in size from 20 nm to 10 μm remained primarily in the suprachoroidal space and choroid for a period of months and did not clear the eye. No adverse effects of injection into the suprachoroidal space were observed. Conclusion. Injection into the suprachoroidal space using a microneedle offers a simple and minimally invasive way to target the delivery of drugs to the choroid and retina. PMID:22669719

Real-time monitoring of hairpin ribozyme kinetics through base-specific quenching of fluorescein-labeled substrates.

PubMed Central

Walter, N G; Burke, J M

1997-01-01

Current methods for evaluating the kinetics of ribozyme-catalyzed reactions rely primarily on the use of radiolabeled RNA substrates, and so require tedious electrophoretic separation and quantitation of reaction products for each data point in any experiment. Here, we report the use of fluorescent substrates for real-time analysis of the time course of reactions of the hairpin ribozyme. Fluorescence of 3' fluorescein-labeled substrates was quenched upon binding to the hairpin ribozyme or its isolated substrate-binding strand (SBS), under conditions of ribozyme or SBS excess. This decrease was accompanied by an increase in anisotropy, and resulted from a base-specific quenching by a guanosine residue added to the 5' end of the SBS, close to fluorescein in the complex. Fluorescence quenching was used to determine rate constants for substrate binding (1.4 x 10(8) M(-1) min(-1)), cleavage (0.15 min(-1)), and substrate dissociation (0.010 min(-1)) by a structurally well-defined ribozyme at 25 degrees C in 50 mM Tris-HCI, pH 7.5, 12 mM MgCl2. These rates are in excellent agreement with those obtained using traditional radioisotopic methods. Measurements of dissociation rates provided physical support for interdomain interactions within the substrate-ribozyme complex. We estimate that 2.1 kcal/mol of additional substrate binding energy is provided by the B domain of the ribozyme. Part of this free energy apparently stems from coaxial stacking of helices in the hinge region between domains, and it is plausible that the remainder might be contributed by direct interactions with loop B. The fluorescence quenching and dequenching methods described here should be readily adaptable to studying a wide variety of RNA interactions and reactions using ribozymes and other model systems. PMID:9085846

[Angiography: safety x economy].

PubMed

Torres, Rogil José de Almeida; Muccioli, Cristina; Casella, Antonio Marcelo Barbante; Regonha, Eduardo; Luchini, Andréa; Weiss, Wilma; Frecceiro, Paulo Roberto

2006-01-01

To evaluate the safety, effectiveness, and cost of angiofluoresceinographic examination by using both the least amount of dye as well as digital equipment along with a laser photocopier. Prospective and comparative study carried out in a group of 70 patients, who underwent an angiographic evaluation with a conventional retinographer injecting 5 ml sodium fluorescein at 10% (control group) as well as a group of 70 patients who underwent an angiographic evaluation with a digital retinographer injecting 2 ml fluorescein at 10% (study group). Arterial pressure, heart rate and oximetry were assessed prior to and after the dye injection. Organic reactions related to the examination were reported. Photograph quality as well as cost between the two techniques were compared. Control group patients showed a greater increase in systolic and diastolic arterial pressure. Heart rate, oximetry measurement and adverse reactions did not show any significant statistical differences between both groups. As for the quality of photographs, a better performance was noticed in the control group. As for the cost, the examination carried out in the study group required lower cost and thus saved around 54.8% per examination in relation to the control group. The examination carried out with a lower dose of fluorescein using digital equipment along with a laser photocopier provided greater stability in the systolic and diastolic arterial pressure. However, it did not have any influence on heart rate, oximetry or adverse reactions such as nausea, vomiting, syncope and rashes. The quality of photographs was poor although they enabled diagnosis as well as therapy follow-up for those who carried out the examination. Moreover, economically the above procedure represented a gain of 66.26%, against 25.81% in relation to the conventional equipment.

Chemiluminescence resonance energy transfer imaging on magnetic particles for single-nucleotide polymorphism detection based on ligation chain reaction.

PubMed

Bi, Sai; Zhang, Zhipeng; Dong, Ying; Wang, Zonghua

2015-03-15

A novel ligation chain reaction (LCR) methodology for single-nucleotide polymorphism (SNP) detection was developed based on luminol-H2O2-horseradish peroxidase (HRP)-mimicking DNAzyme-fluorescein chemiluminescence resonance energy transfer (CRET) imaging on magnetic particles. For LCR, four unique target-complement probes (X and X(⁎), YG and Y(⁎)) for the amplification of K-ras (G12C) were designed by modifying G-quadruplex sequence at 3'-end of YG and fluorescein at 5'-end of Y(⁎). After the LCR, the resulting products of XYG/X(⁎)Y(⁎) with biotin-labeled X(⁎) were captured onto streptavidin-coated magnetic particles (SA-MPs) via specific biotin-SA interaction, which stimulated the CRET reaction from hemin/G-quadruplex-catalyzed luminol-H2O2 CL system to fluorescein. By collecting signals by a cooled low-light CCD, a CRET imaging method was proposed for visual detection and quantitative analysis of SNP. As low as 0.86fM mutant DNA was detected by this assay, and positive mutation detection was achieved with a wild-type to mutant ratio of 10,000:1. This high sensitivity and specificity could be attributed to not only the exponential amplification and excellent discrimination of LCR but also the employment of SA-MPs. SA-MPs ensured the feasibility of the proposed strategy, which also simplified the operations through magnetic separation and separated the reaction and detection procedures to improve sensitivity. The proposed LCR-CRET imaging strategy extends the application of signal amplification techniques to SNP detection, providing a promising platform for effective and high-throughput genetic diagnosis. Copyright © 2014 Elsevier B.V. All rights reserved.

Effects of Punctal Occlusion on Clinical Signs and Symptoms and on Tear Cytokine Levels in Patients with Dry Eye.

PubMed

Tong, Louis; Beuerman, Roger; Simonyi, Susan; Hollander, David A; Stern, Michael E

2016-04-01

To investigate changes in signs, symptoms, and tear cytokines following punctal plug occlusion in patients with dry eye. A single-center study was conducted at Singapore Eye Research Institute. Nonabsorbable punctal plugs were inserted in the lower punctum of both eyes in patients with moderate dry eye. Over 3 weeks, in the more severe eye, dry eye symptoms, fluorescein corneal staining, Schirmer I (without topical anesthesia) test, tear film breakup time (TFBUT), and safety were assessed. Cytokine and matrix metalloproteinase-9 (MMP-9) levels in tear samples were measured. Twenty-nine patients (mean age 49.8 years) with moderate dry eye were evaluated. At baseline, mean (standard deviation) global symptoms score was 53.8 (26.5), Schirmer I test score was 5.1 (2.8) mm, and TFBUT was 2.2 (0.6) seconds. After 3 weeks, punctal occlusion significantly reduced global irritation symptoms score (P<.001) and decreased fluorescein staining in all zones (P<.01) except the inferior zone (P=.42). No significant association between levels of cytokines or MMP-9 and either TFBUT or global irritation symptoms were observed at baseline. Levels of several cytokines and MMP-9 were higher in patients with Schirmer I test scores ≤8 mm at baseline. After 3 weeks of punctal occlusion, no significant changes in overall cytokine or MMP-9 levels were observed. Punctal plug occlusion provided symptomatic relief and reduced fluorescein staining in all except the inferior zone. However, insertion of punctal plugs had minimal effect on tear cytokines and MMP-9 levels, suggesting a need for earlier treatment with anti-inflammatory agents for management of dry eye disease. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Goldmann tonometry tear film error and partial correction with a shaped applanation surface.

PubMed

McCafferty, Sean J; Enikov, Eniko T; Schwiegerling, Jim; Ashley, Sean M

2018-01-01

The aim of the study was to quantify the isolated tear film adhesion error in a Goldmann applanation tonometer (GAT) prism and in a correcting applanation tonometry surface (CATS) prism. The separation force of a tonometer prism adhered by a tear film to a simulated cornea was measured to quantify an isolated tear film adhesion force. Acrylic hemispheres (7.8 mm radius) used as corneas were lathed over the apical 3.06 mm diameter to simulate full applanation contact with the prism surface for both GAT and CATS prisms. Tear film separation measurements were completed with both an artificial tear and fluorescein solutions as a fluid bridge. The applanation mire thicknesses were measured and correlated with the tear film separation measurements. Human cadaver eyes were used to validate simulated cornea tear film separation measurement differences between the GAT and CATS prisms. The CATS prism tear film adhesion error (2.74±0.21 mmHg) was significantly less than the GAT prism (4.57±0.18 mmHg, p <0.001). Tear film adhesion error was independent of applanation mire thickness ( R 2 =0.09, p =0.04). Fluorescein produces more tear film error than artificial tears (+0.51±0.04 mmHg; p <0.001). Cadaver eye validation indicated the CATS prism's tear film adhesion error (1.40±0.51 mmHg) was significantly less than that of the GAT prism (3.30±0.38 mmHg; p =0.002). Measured GAT tear film adhesion error is more than previously predicted. A CATS prism significantly reduced tear film adhesion error bŷ41%. Fluorescein solution increases the tear film adhesion compared to artificial tears, while mire thickness has a negligible effect.

Effects of topical cyclosporine a plus artificial tears versus artificial tears treatment on conjunctival goblet cell density in dysfunctional tear syndrome.

PubMed

Demiryay, Elvan; Yaylali, Volkan; Cetin, Ebru Nevin; Yildirim, Cem

2011-09-01

The aim was to compare the effects of topical cyclosporine A and artificial tears combination with artificial tears alone in patients with dysfunctional tear syndrome (DTS). Forty-two eyes of 42 patients with DTS were enrolled in the study. The inclusion criteria for the study were Schirmer I (without anesthesia) scores below 10 mm/5 min and tear film break-up time (BUT) below 10 sec. The patients were randomly divided into two groups. The study group (22 patients) underwent 0.05% cyclosporine A treatment twice a day and preservative-free artificial tears for four times a day for 4 months. The control group (20 patients) was administered only preservative-free artificial tears four times a day for 4 months. The BUT, Schirmer test scores, corneal fluorescein staining, conjunctival lissamine green staining, and goblet cell density derived by impression cytology were recorded before and after treatment in each group. In the study group, all parameters improved statistically significantly after treatment at the 4-month follow-up compared with the pretreatment values (P<0.001 for all). In the control group, corneal fluorescein staining (P<0.001) and conjunctival lissamine green staining (P=0.014) improved, but BUT and Schirmer scores did not change significantly after treatment. At the end of the 4-month follow-up, the study group demonstrated statistically significantly better BUT (P=0.020), Schirmer scores (P=0.002), goblet cell density (P=0.006), corneal fluorescein staining (P=0.003), and conjunctival lissamine green staining (P=0.017) scores than did the control group. Topical cyclosporine A and artificial tears treatment significantly increases goblet cell density, decreases the signs of DTS, and improves ocular surface health.

DMSO modifies the permeability of the zebrafish (Danio rerio) chorion-implications for the fish embryo test (FET).

PubMed

Kais, B; Schneider, K E; Keiter, S; Henn, K; Ackermann, C; Braunbeck, T

2013-09-15

Since 2007, when REACH came into force, the fish embryo test has received increasing attention as a potential alternative for the acute fish test. Due to its low toxicity and the ability to permeate biological membranes without significant damage to their structural integrity, dimethyl sulfoxide (DMSO) is a commonly used solvent in the fish embryo test. Little is known, however, about the membrane penetration properties of DMSO, the impact of different concentrations of DMSO on the potential barrier function of the zebrafish chorion and on changes in the uptake of chemicals into the embryo. Therefore, in the present study, the fluorescent dyes fluorescein (mol wt 332; Pow 3.4) and 2,7-dichlorofluorescein (mol wt 401; Pow 4.7), both substances with limited water solubility, were used to visualize the uptake into the egg as well as the accumulation in the embryo of the zebrafish depending on different concentrations of DMSO. The distribution of fluorescein within the egg compartments varied with DMSO concentration: When dissolved in 0.01% DMSO, fluorescein did not pass the chorion. In contrast, concentrations ≥ 0.1% DMSO increasingly facilitated the uptake into the perivitelline space. In contrast, the uptake of 2,7-dichlorofluorescein was not substantially increased with rising DMSO concentrations, indicating the importance of factors other than the solvent (e.g. mol wt). With respect to the fish embryo test, results indicate that DMSO may be used without complications as a solvent, however, only at a maximum concentration of 0.01% (0.1 mL/L) as already indicated in the OECD difficult substances paper (OECD, 2000). Copyright © 2013 Elsevier B.V. All rights reserved.

Perilymph pharmacokinetics of markers and dexamethasone applied and sampled at the lateral semi-circular canal.

PubMed

Salt, Alec N; Hartsock, Jared J; Gill, Ruth M; Piu, Fabrice; Plontke, Stefan K

2012-12-01

Perilymph pharmacokinetics was investigated by a novel approach, in which solutions containing drug or marker were injected from a pipette sealed into the perilymphatic space of the lateral semi-circular canal (LSCC). The cochlear aqueduct provides the outlet for fluid flow so this procedure allows almost the entire perilymph to be exchanged. After wait times of up to 4 h the injection pipette was removed and multiple, sequential samples of perilymph were collected from the LSCC. Fluid efflux at this site results from cerebrospinal fluid (CSF) entry into the basal turn of scala tympani (ST) so the samples allow drug levels from different locations in the ear to be defined. This method allows the rate of elimination of substances from the inner ear to be determined more reliably than with other delivery methods in which drug may only be applied to part of the ear. Results were compared for the markers trimethylphenylammonium (TMPA) and fluorescein and for the drug dexamethasone (Dex). For each substance, the concentration in fluid samples showed a progressive decrease as the delay time between injection and sampling was increased. This is consistent with the elimination of substance from the ear with time. The decline with time was slowest for fluorescein, was fastest for Dex, with TMPA at an intermediate rate. Simulations of the experiments showed that elimination occurred more rapidly from scala tympani (ST) than from scala vestibuli (SV). Calculated elimination half-times from ST averaged 54.1, 24.5 and 22.5 min for fluorescein, TMPA and Dex respectively and from SV 1730, 229 and 111 min respectively. The elimination of Dex from ST occurred considerably faster than previously appreciated. These pharmacokinetic parameters provide an important foundation for understanding of drug treatments of the inner ear.

Incidental retinal phototoxicity associated with ingestion of photosensitizing drugs.

PubMed

Mauget-Faÿsse, M; Quaranta, M; Francoz, N; BenEzra, D; Mauget-Fa, M

2001-07-01

to report on the possible correlation between incident retinal phototoxicity and the use of photosensitizing drugs. four patients were examined because of scotomas and visual loss after an incidental exposure to a strong light source. One patient (two eyes) was exposed to standard camera flash; one patient (one eye) had a brief exposure to welding light; one patient (two eyes) underwent uncomplicated phacoemulsifications with intraocular lens implantation. The fourth patient had a severe retinal phototoxicity following a secondary intraocular lens implantation. All four patients underwent a thorough assessment including history of systemic drug use. These patients had ophthalmologic evaluation including: best corrected visual acuity (ETDRS charts), fundus examination, fluorescein and indocyanine green angiographies and were followed for 1 year. on presentation, the mean visual acuity was 7.5/20 (range: 20/400-20/20). Fundus examination disclosed yellow-gray sub-retinal lesions in all affected eyes. Early phase fluorescein angiography showed one or multiple hypofluorescent spots surrounded by a halo of hyperfluorescent window defect. In the late phase, some of these spots leaked the fluorescein dye. Indocyanine green angiography demonstrated hypofluorescent spots throughout with ill-defined borders of hyperfluorescence observed during the late stages. The common finding in these four patients was the fact that they were all taking one or more photosensitizing drugs (hydrochlorothiazide, furosemide, allopurinol, and benzodiazepines). Three of the patients had a full visual recovery a few months after the phototoxicity. The fourth patient remained with a visual acuity of 20/60 12 months after the light exposure. Despite the visual recovery, non-homogeneous retinal pigment epithelial disturbances persisted in all affected eyes. phototoxicity following incidental light exposure may occur in patients taking drugs of photosensitizing potential. Therefore, the thorough history of systemic drug ingestion should be obtained if patients have exposure to strong light sources.

A nanomedicine to treat ocular surface inflammation: performance on an experimental dry eye murine model.

PubMed

Contreras-Ruiz, L; Zorzi, G K; Hileeto, D; López-García, A; Calonge, M; Seijo, B; Sánchez, A; Diebold, Y

2013-05-01

MUC5AC is a glycoprotein with gel-forming properties, whose altered expression has been implicated in the pathogenesis of dry eye disease. The aim of our study was to achieve an efficient in vivo transfection of MUC5AC, restore its normal levels in an inflamed ocular surface and determine whether restored MUC5AC levels improve ocular surface inflammation. Cationized gelatin-based nanoparticles (NPs) loaded with a plasmid coding a modified MUC5AC protein (pMUC5AC) were instilled in healthy and experimental dry eye (EDE) mice. MUC5AC expression, clinical signs, corneal fluorescein staining and tear production were evaluated. Ocular specimens were processed for histopathologic evaluation, including goblet cell count and CD4 immunostaining. Neither ocular discomfort nor irritation was observed in vivo after NP treatment. Expression of modified MUC5AC was significantly higher in ocular surface tissue of pMUC5AC-NP-treated animals than that of controls. In healthy mice, pMUC5AC-NPs had no effect on fluorescein staining or tear production. In EDE mice, both parameters significantly improved after pMUC5AC-NP treatment. Anterior eye segment of treated mice showed normal architecture and morphology with lack of remarkable inflammatory changes, and a decrease in CD4+ T-cell infiltration. Thus, pMUC5AC-NPs were well tolerated and able to induce the expression of modified MUC5A in ocular surface tissue, leading to reduction of the inflammation and, consequently improving the associated clinical parameters, such as tear production and fluorescein staining. These results identify a potential application of pMUC5AC-NPs as a new therapeutic modality for the treatment of dry eye disease.

Therapeutic effects of topical doxycycline in a benzalkonium chloride-induced mouse dry eye model.

PubMed

Zhang, Zhen; Yang, Wen-Zhao; Zhu, Zhen-Zhen; Hu, Qian-Qian; Chen, Yan-Feng; He, Hui; Chen, Yong-Xiong; Liu, Zu-Guo

2014-05-06

We investigated the therapeutic effects and underlying mechanisms of topical doxycycline in a benzalkonium chloride (BAC)-induced mouse dry eye model. Eye drops containing 0.025%, 0.1% doxycycline or solvent were administered to a BAC-induced dry eye model four times daily. The clinical evaluations, including tear break-up time (BUT), fluorescein staining, inflammatory index, and tear volume, were performed on days 0, 1, 4, 7, and 10. Global specimens were collected on day 10 and processed for immunofluorescent staining, TUNEL, and periodic acid-Schiff assay. The levels of inflammatory mediators in the corneas were determined by real-time PCR. The total and phosphorylated nuclear factor-κB (NF-κB) were detected by Western blot. Both 0.025% and 0.1% doxycycline treatments resulted in increased BUT, lower fluorescein staining scores, and inflammatory index on days 4, 7, and 10, while no significant change in tear volume was observed. The 0.1% doxycycline-treated group showed more improvements in decreasing fluorescein staining scores, increasing Ki-67-positive cells, and decreasing TUNEL- and keratin-10-positive cells than other groups. The mucin-filled goblet cells in conjunctivas were increased, and the expression of CD11b and levels of matrix metalloproteinase-9, IL-1β, IL-6, TNF-α, macrophage inflammatory protein-2, and cytokine-induced neutrophil chemoattractant in corneas were decreased in both doxycycline-treated groups. In addition, doxycycline significantly reduced the phosphorylation of NF-κB activated in the BAC-treated corneas. Topical doxycycline showed clinical improvements and alleviated ocular surface inflammation on BAC-induced mouse dry eye, suggesting a potential as an anti-inflammatory agent in the clinical treatment of dry eye.

Optical Coherence Tomography Angiography Features of Iris Racemose Hemangioma in 4 Cases.

PubMed

Chien, Jason L; Sioufi, Kareem; Ferenczy, Sandor; Say, Emil Anthony T; Shields, Carol L

2017-10-01

Optical coherence tomography angiography (OCTA) allows visualization of iris racemose hemangioma course and its relation to the normal iris microvasculature. To describe OCTA features of iris racemose hemangioma. Descriptive, noncomparative case series at a tertiary referral center (Ocular Oncology Service of Wills Eye Hospital). Patients diagnosed with unilateral iris racemose hemangioma were included in the study. Features of iris racemose hemangioma on OCTA. Four eyes of 4 patients with unilateral iris racemose hemangioma were included in the study. Mean patient age was 50 years, all patients were white, and Snellen visual acuity was 20/20 in each case. All eyes had sectoral iris racemose hemangioma without associated iris or ciliary body solid tumor on clinical examination and ultrasound biomicroscopy. By anterior segment OCT, the racemose hemangioma was partially visualized in all cases. By OCTA, the hemangioma was clearly visualized as a uniform large-caliber vascular tortuous loop with intense flow characteristics superimposed over small-caliber radial iris vessels against a background of low-signal iris stroma. The vascular course on OCTA resembled a light bulb filament (filament sign), arising from the peripheral iris (base of light bulb) and forming a tortuous loop on reaching its peak (midfilament) near the pupil (n = 3) or midzonal iris (n = 1), before returning to the peripheral iris (base of light bulb). Intravenous fluorescein angiography performed in 1 eye depicted the iris hemangioma; however, small-caliber radial iris vessels were more distinct on OCTA than intravenous fluorescein angiography. Optical coherence tomography angiography is a noninvasive vascular imaging modality that clearly depicts the looping course of iris racemose hemangioma. Optical coherence tomography angiography depicted fine details of radial iris vessels, not distinct on intravenous fluorescein angiography.

A level-set method for pathology segmentation in fluorescein angiograms and en face retinal images of patients with age-related macular degeneration

NASA Astrophysics Data System (ADS)

Mohammad, Fatimah; Ansari, Rashid; Shahidi, Mahnaz

2013-03-01

The visibility and continuity of the inner segment outer segment (ISOS) junction layer of the photoreceptors on spectral domain optical coherence tomography images is known to be related to visual acuity in patients with age-related macular degeneration (AMD). Automatic detection and segmentation of lesions and pathologies in retinal images is crucial for the screening, diagnosis, and follow-up of patients with retinal diseases. One of the challenges of using the classical level-set algorithms for segmentation involves the placement of the initial contour. Manually defining the contour or randomly placing it in the image may lead to segmentation of erroneous structures. It is important to be able to automatically define the contour by using information provided by image features. We explored a level-set method which is based on the classical Chan-Vese model and which utilizes image feature information for automatic contour placement for the segmentation of pathologies in fluorescein angiograms and en face retinal images of the ISOS layer. This was accomplished by exploiting a priori knowledge of the shape and intensity distribution allowing the use of projection profiles to detect the presence of pathologies that are characterized by intensity differences with surrounding areas in retinal images. We first tested our method by applying it to fluorescein angiograms. We then applied our method to en face retinal images of patients with AMD. The experimental results included demonstrate that the proposed method provided a quick and improved outcome as compared to the classical Chan-Vese method in which the initial contour is randomly placed, thus indicating the potential to provide a more accurate and detailed view of changes in pathologies due to disease progression and treatment.

CRYPTOSPORIDIUM OOCYST RECOVERY IN WATER BY EPA METHOD 1623: EVALUATION OF A MODIFIED IMS DISSOCIATION

EPA Science Inventory

EPA Methods 1622 and 1623 are the benchmarks for detection of Cryptosporidium spp. oocysts in water. These methods consist of filtration, elution, purification by immunomagnetic separation (IMS), and microscopic analysis after staining with a fluorescein isothiocyanate conjugate...

Automated Quantitative Characterization of Retinal Vascular Leakage and Microaneurysms in Ultra-widefield Fluorescein Angiography

PubMed Central

Ehlers, Justis P.; Wang, Kevin; Vasanji, Amit; Hu, Ming; Srivastava, Sunil K.

2017-01-01

Summary Ultra-widefield fluorescein angiography (UWFA) is an emerging imaging modality used to characterize pathology in the retinal vasculature such as microaneurysms (MA) and vascular leakage. Despites its potential value for diagnosis and disease surveillance, objective quantitative assessment of retinal pathology by UWFA is currently limited because it requires laborious manual segmentation by trained human graders. In this report, we describe a novel fully automated software platform, which segments MAs and leakage areas in native and dewarped UWFA images with retinal vascular disease. Comparison of the algorithm to human grader generated gold standards demonstrated significant strong correlations for MA and leakage areas (ICC=0.78-0.87 and ICC=0.70-0.86, respectively, p=2.1×10-7 to 3.5×10-10 and p=7.8×10-6 to 1.3×10-9, respectively). These results suggest the algorithm performs similarly to human graders in MA and leakage segmentation and may be of significant utility in clinical and research settings. PMID:28432113

A compact and low-cost laser induced fluorescence detector with silicon based photodetector assembly for capillary flow systems.

PubMed

Geng, Xuhui; Shi, Meng; Ning, Haijing; Feng, Chunbo; Guan, Yafeng

2018-05-15

A compact and low-cost laser induced fluorescence (LIF) detector based on confocal structure for capillary flow systems was developed and applied for analysis of Her2 protein on single Hela cells. A low-power and low-cost 450 nm laser diode (LD) instead of a high quality laser was used as excitation light source. A compact optical design together with shortened optical path length improved the optical efficiency and detection sensitivity. A superior silicon based photodetector assembly was used for fluorescence detection instead of a photomultiplier (PMT). The limit of detection (LOD) for fluorescein sodium was 3 × 10 -12 M or 165 fluorescein molecules in detection volume measured on a homemade capillary electroosmotic driven (EOD)-LIF system, which was similar to commercial LIFs. Compared to commercial LIFs, the whole volume of our LIF was reduced to 1/2-1/3, and the cost was less than 1/3 of them. Copyright © 2018 Elsevier B.V. All rights reserved.

Fluorescein: a Photo-CIDNP Sensitizer Enabling Hyper-Sensitive NMR Data Collection in Liquids at Low Micromolar Concentration

PubMed Central

Okuno, Yusuke; Cavagnero, Silvia

2016-01-01

Photochemically induced dynamic nuclear polarization (photo-CIDNP) is a powerful approach for sensitivity enhancement in NMR spectroscopy. In liquids, inter-molecular photo-CIDNP depends on the transient bimolecular reaction between photoexcited dye and sample of interest. Hence the extent of polarization is sample-concentration dependent. This study introduces fluorescein (FL) as a photo-CIDNP dye whose performance is exquisitely tailored to data collection at extremely low sample concentrations. The photo-CIDNP resonance intensities of tryptophan in the presence of either FL or FMN (i.e., the routinely employed flavin mononucleotide photosensitizer) in the liquid state show that FL yields superior sensitivity and enables rapid data collection down to an unprecedented 1 micromolar concentration. This result was achieved on a conventional spectrometer operating at 14.1 Tesla, and equipped with a room-temperature probe (i.e., non-cryogenic). Kinetic simulations show that the excellent behavior of FL arises from its long excited-state triplet lifetime and superior photostability relative to conventional photo-CIDNP sensitizers. PMID:26744790

Improved salvage of complicated microvascular transplants monitored with quantitative fluorometry.

PubMed

Whitney, T M; Lineaweaver, W C; Billys, J B; Siko, P P; Buncke, G M; Alpert, B S; Oliva, A; Buncke, H J

1992-07-01

Quantitative fluorometry has been used to monitor circulation in transplanted toes and cutaneous flaps in our unit since 1982. Analysis of 177 uncomplicated transplants monitored by quantitative fluorometry shows that this technique has low false indication rates for arterial occlusion (0.6 percent of patients) and venous occlusion (6.2 percent of patients). None of these patients was reexplored because of a false monitor reading, and except for single abnormal sequences, monitoring appropriately indicated intact circulation throughout the postoperative period. Quantitative fluorometry has correctly indicated vascular complications in 21 (91.3 percent) of 23 transplants over an 8-year period. The salvage rate (85.7 percent) of the fluorescein-monitored reexplored transplants was significantly higher than the salvage rates of similar reexplored transplants not monitored with fluorescein and of reexplored muscle flaps (which cannot be monitored with the fluorometer used at this unit). These clinical data indicate that quantitative fluorometry is a valid and useful postoperative monitor for transplanted toes and cutaneous flaps.

Fluorescein angiography of the iris in anterior segment pigment dispersal syndrome.

PubMed Central

Gillies, W E; Tangas, C

1986-01-01

The results are presented of fluorescein angiography of the iris in 11 patients with anterior segment pigment dispersal syndrome. These show a general hypovascularity of the iris with fine neovascularisation at the pupil margin and the peripupillary area. Hypoplasia of the iris stroma was also present in many cases. When the condition was virtually unilateral, the vascular changes were present though less marked in the relatively unaffected eye. It is postulated that the anterior segment pigment dispersal syndrome is secondary to a congenital mesodermal deficiency of the iris stroma with hypovascularity of the iris, which forms a poor support tissue for the pigment epithelium of the iris, resulting in shedding of pigment granules particularly in the region of the attachment of the dilator muscle to the pigment epithelium. The condition may be hereditary. Because of the hypovascularity the mesodermal hypoplasia may be progressive, but pigment release may diminish in later life with treatment, with consequent diminution of pupil activity. Images PMID:2421760

Honey-Induced Protein Stabilization as Studied by Fluorescein Isothiocyanate Fluorescence

PubMed Central

Abdul Kadir, Habsah; Tayyab, Saad

2013-01-01

Protein stabilizing potential of honey was studied on a model protein, bovine serum albumin (BSA), using extrinsic fluorescence of fluorescein isothiocyanate (FITC) as the probe. BSA was labelled with FITC using chemical coupling, and urea and thermal denaturation studies were performed on FITC-labelled BSA (FITC-BSA) both in the absence and presence of 10% and 20% (w/v) honey using FITC fluorescence at 522 nm upon excitation at 495 nm. There was an increase in the FITC fluorescence intensity upon increasing urea concentration or temperature, suggesting protein denaturation. The results from urea and thermal denaturation studies showed increased stability of protein in the presence of honey as reflected from the shift in the transition curve along with the start point and the midpoint of the transition towards higher urea concentration/temperature. Furthermore, the increase in ΔG D H2O and ΔG D 25°C in presence of honey also suggested protein stabilization. PMID:24222758

The effect of oral acetazolamide on cystoid macular edema in hydroxychloroquine retinopathy: a case report.

PubMed

Hong, Eun Hee; Ahn, Seong Joon; Lim, Han Woong; Lee, Byung Ro

2017-07-12

Hydroxychloroquine (HCQ) retinopathy can accompany other retinal complications such as cystoid macular edema (CME), which leads to central visual loss. We report a case of CME with HCQ retinopathy that improved with the use of oral acetazolamide, and discussed the possible mechanisms of CME in HCQ retinopathy using multimodal imaging modalities. A 62-year-old patient with systemic lupus erythematosus (SLE) and HCQ retinopathy developed bilateral CME with visual decline. Fluorescein angiography (FA) showed fluorescein leakage in the macular and midperipheral area. After treatment with oral acetazolamide (250 mg/day) for one month, CME was completely resolved, best corrected visual acuity (BCVA) improved from 20/50 to 20/25, and FA examination showed decreased dye leakage in the macular and midperipheral areas. In cases of vision loss in HCQ retinopathy, it is important to consider not only progression of maculopathy, but also development of CME, which can be effectively treated with oral acetazolamide.

Fluorescein-labeled β-Glucosidase as a Bacterial Stain

PubMed Central

Pital, Abe; Janowitz, Sheldon L.; Hudak, Charles E.; Lewis, Evelyn E.

1967-01-01

Fluorescein isothiocyanate-labeled β-glucosidase was used as a simple staining reagent with selected gram-positive and gram-negative organisms. Staining in situ appeared to be dependent on the presence of accessible glycosidic-type linkages in the bacterial cell wall. Extensive wall damage or lysis did not occur when stained cells were suspended in washing and mounting solutions. The apparent specificity of labeled enzyme for wall substance was tested by blocking reactions, staining of isolated cell walls, and failure to stain substances lacking appropriate glycosidic linkages. Severe cell wall lesions were produced after prolonged contact with labeled enzyme, and this phenomenon may also be related to staining specificity. Gram-negative organisms and spores were poorly stained unless protected glycopeptide substrate was previously exposed by treatment of cells with thioglycolic acid or dilute alkaline sodium hypochlorite solution. A potential for staining tissues and cell lines may also exist. Some possible applications of labeled enzymes are briefly discussed. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:4169543

Evaluation of retinoic acid ophthalmic emulsion in dry eye.

PubMed

Selek, H; Unlü, N; Orhan, M; Irkeç, M

2000-01-01

An oil in water emulsion of 0.01% all-trans-retinoic acid (tretinoin) was prepared and clinically evaluated in dry eye patients. The ophthalmic emulsion consisted of 10% of arachis oil and 90% of the hydrogel of Carbopol 940. To evaluate retinoic acid emulsion clinically, a placebo-controlled, open-labeled, randomized study was performed with 22 dry-eye patients. Symptoms were recorded before and after the treatments. The Schirmer I test, measurement of tear film break-up time (BUT), rose Bengal and fluorescein staining of cornea and conjunctiva, and mucus fern test were done. Retinoic acid did not improve the dryness, photophobia and foreign body sensation more than placebo. Schirmer test and BUT were significantly improved by retinoic acid treatment. Corneal and conjunctival epithelium maintained their characteristics during the use of retinoic acid, as indicated by rose Bengal and fluorescein staining. Ophthalmic emulsion of retinoic acid can be suggested as a promising approach for the treatment of dry eye.

Bulbar conjunctival fluorescein staining in hydrogel contact lens wearers.

PubMed

Lakkis, C; Brennan, N A

1996-07-01

To evaluate the clinical usefulness of bulbar conjunctival staining in assessing hydrogel contact lens patients. Overall staining, as well as staining at five separate sites (limbal, nasal band, temporal band, superior, and inferior) was graded on an analog scale in 48 contact lens wearing subjects and 50 controls. The degree to which subjects experienced sensations of dryness, wateriness, itchiness, grittiness, and comfort was also assessed using analog scales. Some measure of conjunctival staining was noted in 98% of subjects, with contact lens wearers showing statistically significant greater staining than controls. Only 12% of controls showed staining of greater than grade 1 (equivalent), whereas 62% of contact lens wearers were above this level. Regression analysis showed that overall staining was a function of whether contact lenses were worn, the degree of dryness, and the amount of itchiness. Conjunctival fluorescein staining appears to serve some clinical usefulness as a composite indicator for certain factors and symptoms and, in addition, provides information which is unique for each individual.

The Impact and Implication of Peripheral Vascular Leakage on Ultra-Widefield Fluorescein Angiography in Uveitis.

PubMed

Thomas, Akshay S; Redd, Travis; Campbell, John P; Palejwala, Neal V; Baynham, Justin T; Suhler, Eric B; Rosenbaum, James T; Lin, Phoebe

2017-10-16

To study if peripheral vascular leakage (PVL) on ultra-widefield fluorescein angiography (UWFFA) prognosticates complications of uveitis or necessitates treatment augmentation. Retrospective cohort study of uveitis patients imaged with UWFFA and ≥1 yr of follow-up. We included 73 eyes of 42 patients with uveitis. There was no difference in baseline, intermediate, final visual acuity (p = 0.47-0.95) or rates of cystoid macular edema (CME) (p = 0.37-0.87) in eyes with PVL vs. those without. Eyes with PVL receiving baseline treatment augmentation were more likely to have baseline CME but were not more likely to have impaired visual acuity at final follow-up. PVL was independently associated with treatment augmentation on generalized estimating equation analysis with multivariable linear regression (OR: 4.39, p = 0.015). PVL did not confer an increased risk of impaired VA or CME at ≥1 yr follow-up but was possibly an independent driver of treatment augmentation.

Fluorescein isothiocyanate-labeled human plasma fibronectin in extracellular matrix remodeling.

PubMed

Hoffmann, Celine; Leroy-Dudal, Johanne; Patel, Salima; Gallet, Olivier; Pauthe, Emmanuel

2008-01-01

Fluorescein isothiocyanate (FITC) is a well-known probe for labeling biologically relevant proteins. However, the impact of the labeling procedure on protein structure and biological activities remains unclear. In this work, FITC-labeled human plasma fibronectin (Fn) was developed to gain insight into the dynamic relationship between cells and Fn. The similarities and differences concerning the structure and function between Fn-FITC and standard Fn were evaluated using biochemical as well as cellular approaches. By varying the FITC/Fn ratio, we demonstrated that overlabeling (>10 FITC molecules/Fn molecule) induces probe fluorescence quenching, protein aggregation, and cell growth modifications. A correct balance between reliable fluorescence for detection and no significant modifications to structure and biological function compared with standard Fn was obtained with a final ratio of 3 FITC molecules per Fn molecule (Fn-FITC3). Fn-FITC3, similar to standard Fn, is correctly recruited into the cell matrix network. Also, Fn-FITC3 is proposed to be a powerful molecular tool to investigate Fn organization and cellular behavior concomitantly.

Optical Coherence Tomography Angiography Features of Diabetic Retinopathy

PubMed Central

Hwang, Thomas S.; Jia, Yali; Gao, Simon S.; Bailey, Steven T.; Lauer, Andreas K.; Flaxel, Christina J.; Wilson, David J.; Huang, David

2015-01-01

Purpose To describe the optical coherence tomography (OCT) angiography features of diabetic retinopathy Methods Using a 70kHz OCT and the split-spectrum amplitude decorrelation angiography (SSADA) algorithm, 6 × 6 mm 3-dimensional angiograms of the macula of 4 patients with diabetic retinopathy were obtained and compared with fluorescein angiography (FA) for features catalogued by the Early Treatment of Diabetic Retinopathy Study. Results OCT angiography detected enlargement and distortion of the foveal avascular zone, retinal capillary dropout, and pruning of arteriolar branches. Areas of capillary loss obscured by fluorescein leakage on FA were more clearly defined on OCT angiography. Some areas of focal leakage on FA that were thought to be microaneurysms were found to be small tufts of neovascularization that extended above the inner limiting membrane. Conclusion OCT angiography does not show leakage, but can better delineate areas of capillary dropout and detect early retinal neovascularization. This new noninvasive angiography technology may be useful for routine surveillance of proliferative and ischemic changes in diabetic retinopathy. PMID:26308529

Immunoregulatory effects of covalent antigen-antibody complexes. III. Enhancement or suppression depending on the time of administration of complex relative to a T-independent antigen.

PubMed Central

Tite, J P; Morrison, C A; Taylor, R B

1981-01-01

The photosensitive affinity label NAP (4-azido-2-nitrophenyl) was used to make a stable covalent-bonded monomeric immune complex (Ag2Ab) between rabbit anti-NAP antibody and a bihaptenic compound containing NAP linked to fluorescein (NAP-aminocaproyl-lysyl-Fl). This complex injected into mice had marked effects on their subsequent response to fluorescein coupled to a thymus-independent carrier (Fl-ficoll). Depending on the time at which the complex was administered relative to challenge, it was possible to obtain either enhancing or suppressive effects. The enhancing but not the suppressive effect of complex was dependent on immune recognition of the rabbit IgG carrier. While the suppressive effect probably results from complex-mediated inactivation of T-independent B cells, it is suggested that the enhancing effect results from priming of the T-dependent B cells by Fl-Ficoll followed by their triggering into antibody production by rabbit IgG-specific helper cells. PMID:7007223

Synthesis and evaluation of fluorogenic triglycerides as lipase assay substrates.

PubMed

Andersen, Rokhsana J; Brask, Jesper

2016-06-01

Three racemic fluorogenic triglycerides are synthesized and evaluated as lipase assay substrates. The presented synthesis route goes through a key triglyceride intermediate which can be chemoselectively functionalized with a wide range of different probes. Hence the substrate can be tailor-made for a specific assay, or focus can be on low cost in larger scale for applications in high-throughput screening (HTS) assays. In the specific examples, TG-ED, TG-FD and TG-F2 are assembled with the Edans-Dabcyl or the fluorescein-Dabcyl FRET pair, or relying on fluorescein self-quenching, respectively. Proof-of-concept assays allowed determination of 1st order kinetic parameters (kcat/KM) of 460s(-1)M(-1), 59s(-1)M(-1) and 346s(-1)M(-1), respectively, for the three substrates. Commercially available EnzChek lipase substrate provided 204s(-1)M(-1). Substrate concentration was identified as a critical parameter, with measured reaction rates decreasing at higher concentrations when intermolecular quenching becomes significant. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Indocyanine green angiographic evidence of choroiditis in scleroderma.

PubMed

Abdellatief, Amro; Balasubramaniam, Saranya C; Grube, Thomas J; Gonzalez Santiago, Tania M; Osborn, Thomas G; Pulido, Jose S

2015-01-01

The purpose of this case report was to demonstrate evidence of indocyanine green angiography leakage consistent with choroiditis in a patient with scleroderma. In this case report, the patient underwent a variety of tests and examinations, including systemic evaluation, full ocular examination, skin biopsy, indocyanine green, and fluorescein angiography testing. A 52-year-old man had blurred vision centrally in both eyes. Vision was 20/25 and 20/20. Posterior examination showed cotton-wool spots in both eyes. The patient met European League against Rheumatism (EULAR) criteria for scleroderma. Fluorescein angiography confirmed the presence of leakage from the retinal vessels. More importantly, indocyanine green angiography revealed choroidal vessel leakage in both eyes. This provided evidence of choroiditis before vessel obliteration. Previous studies have shown evidence of choriocapillaris obliteration. Choroidal vessel leakage, however, has not been reported in patients with scleroderma. The results of this case demonstrate the usefulness of indocyanine green angiography in detecting the presence of choroiditis in scleroderma.

Ultrawide-field Fluorescein Angiography for Evaluation of Diabetic Retinopathy

PubMed Central

Kong, Mingui; Lee, Mee Yon

2012-01-01

Purpose To investigate the advantages of ultrawide-field fluorescein angiography (FA) over the standard fundus examination in the evaluation of diabetic retinopathy (DR). Methods Ultrawide-field FAs were obtained in 118 eyes of 59 diabetic patients; 11 eyes with no DR, 71 eyes with nonproliferative diabetic retinopathy (NPDR), and 36 eyes with proliferative diabetic retinopathy (PDR), diagnosed by the standard method. The presence of peripheral abnormal lesions beyond the standard seven fields was examined. Results Ultrawide-field FA images demonstrated peripheral microaneurysms in six (54.5%) of 11 eyes with no DR and all eyes with moderate to severe NPDR and PDR. Peripheral retinal neovascularizations were detected in three (4.2%) of 71 eyes with NPDR and in 13 (36.1%) of 36 eyes with PDR. Peripheral vascular nonperfusion and vascular leakage were found in two-thirds of eyes with severe NPDR and PDR. Conclusions Ultrawide-field FA demonstrates peripheral lesions beyond standard fields, which can allow early detection and a close evaluation of DR. PMID:23204797

Intermolecular G-quadruplex structure-based fluorescent DNA detection system.

PubMed

Zhou, Hui; Wu, Zai-Sheng; Shen, Guo-Li; Yu, Ru-Qin

2013-03-15

Adopting multi-donors to pair with one acceptor could improve the performance of fluorogenic detection probes. However, common dyes (e.g., fluorescein) in close proximity to each other would self-quench the fluorescence, and the fluorescence is difficult to restore. In this contribution, we constructed a novel "multi-donors-to-one acceptor" fluorescent DNA detection system by means of the intermolecular G-quadruplex (IGQ) structure-based fluorescence signal enhancement combined with the hairpin oligonucleotide. The novel IGQ-hairpin system was characterized using the p53 gene as the model target DNA. The proposed system showed an improved assay performance due to the introduction of IGQ-structure into fluorescent signaling probes, which could inhibit the background fluorescence and increase fluorescence restoration amplitude of fluoresceins upon target DNA hybridization. The proof-of-concept scheme is expected to provide new insight into the potential of G-quadruplex structure and promote the application of fluorescent oligonucleotide probes in fundamental research, diagnosis, and treatment of genetic diseases. Copyright © 2012 Elsevier B.V. All rights reserved.

N-bromosuccinimide-fluorescein based sensitive flow-injection chemiluminescence determination of phenformin.

PubMed

Wang, Zhouping; Zhang, Zhujun; Fu, Zhifeng; Fang, Luqiu; Zhang, Xiao

2004-02-01

A novel and highly sensitive method for the determination of phenformin over the range of 6 x 10(-9) - 1 x 10(-5) g ml(-1) in pharmaceutical formulations with flow-injection chemiluminescence (CL) detection is proposed. The method is based on the CL produced during the oxidation of N-bromosuccinimide (NBS) in an alkaline medium in the presence of fluorescein as an effective energy transfer agent. The use of cetyltrimethylammonium bromide (CTAB) as a sensitizer enhances the signal magnitude by about 100 times. The detection limit is 2 x 10(-9) g ml(-1) (3sigma) with a relative standard deviation of 2.3% (n = 11) at 1 x 10(-7) g ml(-1) phenformin. Ninety samples can be determined per hour. The method was evaluated by carrying out a recovery study and by the analysis of commercial formulations. The obtained results compared well with those by an official method, and demonstrated good accuracy and precision. The possible CL mechanism of the proposed system was also briefly analyzed.

Fluorescent Staining of Tea Pathogenic Fungi in Tea Leaves Using Fluorescein-labeled Lectin

NASA Astrophysics Data System (ADS)

Yamada, Kengo; Yoshida, Katsuyuki; Sonoda, Ryoichi

Fluorochrome-labeled lectin, fluorescein conjugated wheat germ agglutinin (F-WGA) was applied to stain tea pathogenic fungi in tea leaf tissue. Infected leaves were fixed and decolorized with a mixture of ethanol and acetic acid, and cleared with 10% KOH for whole mount before staining with F-WGA. Hyphae of Pestalotiopsis longiseta, Pseudocercospora ocellata, Botrytis cinerea and Colletotrichum theae-sinensis fluoresced brightly in whole mount and sectioned samples of infected leaf tissue. In browned tissue, hyphae did not fluoresce frequently in whole mount sample. Autofluorescence of leaf tissue was strong in browned tissue of sections, it was removed by 10% KOH treatment before staining. Penetration hyphae of C. theae-sinensis in cell wall of trichome and hyphae in basal part of trichome did not fluoresced frequently. In whole mount samples of tea leaf infected with Exobasidium vexans and E. reticulatum, hymenia appeared on leaf surface fluoresced, but hyphae in leaf tissue did not fluoresce. In sectioned samples, hyphae fluoresced brightly when sections were treated with 10% KOH before staining.

Identification of Mycobacterial Surface Proteins Released into Subcellular Compartments of Infected Macrophages

PubMed Central

Beatty, Wandy L.; Russell, David G.

2000-01-01

Considerable effort has focused on the identification of proteins secreted from Mycobacterium spp. that contribute to the development of protective immunity. Little is known, however, about the release of mycobacterial proteins from the bacterial phagosome and the potential role of these molecules in chronically infected macrophages. In the present study, the release of mycobacterial surface proteins from the bacterial phagosome into subcellular compartments of infected macrophages was analyzed. Mycobacterium bovis BCG was surface labeled with fluorescein-tagged succinimidyl ester, an amine-reactive probe. The fluorescein tag was then used as a marker for the release of bacterial proteins in infected macrophages. Fractionation studies revealed bacterial proteins within subcellular compartments distinct from mycobacteria and mycobacterial phagosomes. To identify these proteins, subcellular fractions free of bacteria were probed with mycobacterium-specific antibodies. The fibronectin attachment protein and proteins of the antigen 85-kDa complex were identified among the mycobacterial proteins released from the bacterial phagosome. PMID:11083824

TRACE ANALYSIS OF FLUORESCEIN-DERIVATIZED PHENOXY ACID HERBICIDES BY MICELLAR ELECTROKINETIC CHROMATOGRAPHY WITH LASER-INDUCTED FLUORESCENCE DETECTION

EPA Science Inventory

Micellar electrokinetic chromatography (MEKC) with laser-induced fluorescence (LIF) detection was used for the trace analysis of phenoxy acid herbicides. Capillary electrophoresis (CE) with LIF detection, which has not previously been used for pesticide analysis, overcomes the po...

Immunofluorescence Staining — EDRN Public Portal

Cancer.gov

Direct immunofluorescence method is used to detect the deposit of immunoglobulins, complement components, fibrinogen, etc. in tissues. This technique is usually performed on frozen sections. The primary antibody is conjugated to fluorescein binds directly with the antigen and can be detected by the fluorescent tag using a fluorescent microscope.

Tunable Artificial Receptor as a Chemical Sensor for V- and G-agents

DTIC Science & Technology

2012-06-01

shows the design concept for a fluorescent tether. The highly electrophilic nature of fluorescein required the used of carefully selected protecting...atoms removed for clarity) and a space-filling model (Key to figures: carbon: grey; oxygen: red; nitrogen: blue; phosphorus: orange; fluorine : yellow

Comparison of ultra-widefield fluorescein angiography with the Heidelberg Spectralis(®) noncontact ultra-widefield module versus the Optos(®) Optomap(®).

PubMed

Witmer, Matthew T; Parlitsis, George; Patel, Sarju; Kiss, Szilárd

2013-01-01

To compare ultra-widefield fluorescein angiography imaging using the Optos(®) Optomap(®) and the Heidelberg Spectralis(®) noncontact ultra-widefield module. Five patients (ten eyes) underwent ultra-widefield fluorescein angiography using the Optos(®) panoramic P200Tx imaging system and the noncontact ultra-widefield module in the Heidelberg Spectralis(®) HRA+OCT system. The images were obtained as a single, nonsteered shot centered on the macula. The area of imaged retina was outlined and quantified using Adobe(®) Photoshop(®) C5 software. The total area and area within each of four visualized quadrants was calculated and compared between the two imaging modalities. Three masked reviewers also evaluated each quadrant per eye (40 total quadrants) to determine which modality imaged the retinal vasculature most peripherally. Optos(®) imaging captured a total retinal area averaging 151,362 pixels, ranging from 116,998 to 205,833 pixels, while the area captured using the Heidelberg Spectralis(®) was 101,786 pixels, ranging from 73,424 to 116,319 (P = 0.0002). The average area per individual quadrant imaged by Optos(®) versus the Heidelberg Spectralis(®) superiorly was 32,373 vs 32,789 pixels, respectively (P = 0.91), inferiorly was 24,665 vs 26,117 pixels, respectively (P = 0.71), temporally was 47,948 vs 20,645 pixels, respectively (P = 0.0001), and nasally was 46,374 vs 22,234 pixels, respectively (P = 0.0001). The Heidelberg Spectralis(®) was able to image the superior and inferior retinal vasculature to a more distal point than was the Optos(®), in nine of ten eyes (18 of 20 quadrants). The Optos(®) was able to image the nasal and temporal retinal vasculature to a more distal point than was the Heidelberg Spectralis(®), in ten of ten eyes (20 of 20 quadrants). The ultra-widefield fluorescein angiography obtained with the Optos(®) and Heidelberg Spectralis(®) ultra-widefield imaging systems are both excellent modalities that provide views of the peripheral retina. On a single nonsteered image, the Optos(®) Optomap(®) covered a significantly larger total retinal surface area, with greater image variability, than did the Heidelberg Spectralis(®) ultra-widefield module. The Optos(®) captured an appreciably wider view of the retina temporally and nasally, albeit with peripheral distortion, while the ultra-widefield Heidelberg Spectralis(®) module was able to image the superior and inferior retinal vasculature more peripherally. The clinical significance of these findings as well as the area imaged on steered montaged images remains to be determined.

Evaluation of a Perforated Drug Delivery System in Mice for Prolonged and Constant Release of a Hydrophilic Drug

DTIC Science & Technology

2012-02-17

A drug delivery system (DDS) consisting of a perforated microtube ( polyimide , inside diameter= 1.8 mm, tube length= 20 mm, hole size= 0.15 mm) was... biocompatible and capable of long-term constant release of hydrophilic drugs such as sodium fluorescein.

Surface binding properties of aged and fresh (recently excreted) Toxoplasma gondii oocysts

USDA-ARS?s Scientific Manuscript database

The surface properties of aged (stored for 10 years) and fresh (recently excreted) oocysts of Toxoplasma gondii were investigated using monoclonal antibody (mAb) and lectin-binding assays. Fresh oocysts bound a wall-specific mAb labeled with fluorescein isothiocyanate while aged oocysts did not. In ...

USE OF METAL- AND FLUORESCEIN-TAGGED MATERIALS TO STUDY SETTLED PARTICLES EXPOSURE PATHWAYS

EPA Science Inventory

Through the use of ten size ranges of tagged materials (Antley et. al., 2000a), inductively coupled plasma- mass spectrometry (ICP-MS) and flourometry are being used to study the movement of settled particles in the indoor environment, exposure pathways, and the collection effi...

EVALUATION OF CRYPTOSPORIDIUM OOCYST RECOVERY IN WATER BY EPA METHOD 1623 WITH A MODIFIED IMS DISSOCIATION PROCEDURE.

EPA Science Inventory

EPA Methods 1622 and 1623 are the benchmarks for detection of Cryptosporidium spp. oocysts in water. 5-7 These methods consist of filtration, elution, purification by immunomagnetic separation (IMS), and microscopic analysis after staining with a fluorescein isothiocyanate conju...

IMPROVED METHODS FOR HEPATITIS A VIRUS AND ROTAVIRUS CONCENTRATION AND DETECTION IN RECREATIONAL, RAW POTABLE, AND FINISHED WATERS

EPA Science Inventory

The report contains procedures for detecting rotaviruses based upon an immunofluorescence test using a monoclonal antibody and fluorescein-isothiocyanate-conjugated antibody staining method to visualize virus-infected cells. Also contained in the report are test methods for detec...

EXPERIMENTAL METHODOLOGIES AND PRELIMINARY TRANSFER FACTOR DATA FOR ESTIMATION OF DERMAL EXPOSURES TO PARTICLES

EPA Science Inventory

Developmental efforts and experimental data are described that focused on quantifying the transfer of particles on a mass basis from indoor surfaces to human skin. Methods were developed that utilized a common fluorescein-tagged Arizona Test Dust (ATD) as a possible surrogate ...

Evaluation of confocal laser endomicroscopy for assessment and monitoring of therapeutic response in patients with inflammatory bowel disease.

PubMed

Karstensen, John Gásdal

2016-11-01

Crohn's disease (CD) and ulcerative colitis (UC) have been associated with altered intestinal barrier function. Moreover, it has been proposed that a defective barrier function is related to risk of relapse in patients with quiescent CD. Fluorescein-aided confocal laser endomicroscopy (CLE) is a novel endoscopic method, which enables real-time in vivo microscopy. Hence, the intestinal barrier function can be assessed as part of endoscopic evaluation of patients with inflammatory bowel disease (IBD) by measuring microerosions and fluorescein leakage into the intestinal lumen. Furthermore, barrier dysfunction can be correlated with biomarkers associated with intestinal barrier impairments. E-cadherin is a key factor for the adherence of epithelial cells and Smad4 is a cofactor in TGF-β signalling, which is compromised in IBD. To correlate ileal and colonic CLE parameters with endoscopy and histopathology in IBD. Further, we wanted to correlate these features with risk of relapse and evaluate whether they were reproducible and reversible after intensified medical treatment. We also wanted to analyse, whether Smad4 and E-cadherin mucosal protein expression levels were associated with impairments of intestinal barrier function. CLE was performed and correlated to histopathology and endoscopic appearance in two prospective studies in CD (n = 39, controls = 11) and UC patients (n = 22, controls = 7), respectively. In the first study, results were correlated to risk of relapse, whereas the latter assessed the reversibility of CLE features in a longitudinal setting. κ-statistics were used in both studies to assess reproducibility of the CLE findings. Furthermore, ileal biopsy specimens from CD patients and controls were stained by immunohistochemistry for Smad4 and E-cadherin and subsequently correlated to the severity of CD and intestinal barrier impairments. We found that fluorescein leakage and microerosions in the terminal ileum were significantly associated with CD com-pared to controls (p = 0.005 and p = 0.006, respectively) and that ileal fluorescein leakage and microerosions could predict relapse (log-rank p = 0.003 and p = 0.017, respectively). In UC patients with clinical relapse, an augmented crypt architecture and colonic fluorescein leakage were significantly correlated to the severity of the disease (p = 0.001 and p < 0.001, respectively). After intensified medical treatment, a correlation was found between histopathological progress and improvement of abnormal colonic crypt architecture (rs = 0.35, p = 0.016), but we did not observe a resolution of the intestinal barrier dysfunction (rs = 0.09, p = 0.56). The inter-observer variability of CLE parameters ranged from fair to substantial, while the intra-observer variability was somewhat higher. Smad4 expression (rs = 0.56, p = 0.002), but not E-cadherin (rs = 0.01, p = 0.95), was correlated with the severity of the disease; however, Smad4 expression did not correlate with a defect barrier function. CLE can visualise crypt alteration and barrier impairments in both CD and UC, which are otherwise undetectable. Further studies are warranted to incorporate CLE in the endoscopic and therapeutic management algorithm for CD and UC possibly refining the definition of mucosal healing. Smad4 expression was correlated with CD as well as disease severity and may serve as a novel treatment target.

21 CFR 74.1322 - D&C Red No. 22.

Code of Federal Regulations, 2010 CFR

2010-04-01

.... The fluorescein is manufactured by the acid condensation of resorcinol and phthalic acid or its... good manufacturing practice: Sum of volatile matter (at 135°C) and halides and sulfates (calculated as... of phthalic acid, not more than 1 percent. Sodium salt of 2-(3,5-Dibromo-2,4-dihydroxybenzoyl)benzoic...

Energy transfer of nucleic acid products

NASA Astrophysics Data System (ADS)

Jung, Paul M.; Hu, Hsiang-Yun; Khalil, Omar S.

1995-04-01

Fluorescence energy transfer was investigated as a homogeneous detection method for the gapped ligase chain reaction (G-LCR). Oligonucleotides of a Chlamydia trachomatic G-LCR probe set were labeled with fluorescein as the donor and Texas Red as the acceptor fluorophore. Amplification and detection of 10 molecules of synthetic target was demonstrated in spiked urine samples.

Preparation and application of new fluorescein-labeled fumonisins B1 in fluorescence polarization analysis technique

USDA-ARS?s Scientific Manuscript database

Objective: To prepare a new fluorescent tracer against common mycotoxins such as fumonisin B1 in order to replace 6-(4,6-Dichlorotriazinyl) aminofluorescein (6-DTAF), an expensive marker, and to develop a technique for quick detection of fumonisin B1 based on the principle of fluorescence polarizati...

Genetic and metabolic variability in autotrophic and heterotrophic bacteria

NASA Technical Reports Server (NTRS)

Decicco, B. T.

1972-01-01

The studies to evaluate an organism's ability to maintain normal physiological activities over a long period of time in a bioregenerative system are presented. Studies reviewed include: heat tolerant mutants of Pseudomonas fluoresceins, virulence factors of the Staphylococci, and the effect of mutations on the virulence for man in common and ubiquitous microorganisms.

COMPARISON OF RAPID METHODS TO EVALUATE CHLORINE INACTIVATION OF THE BIOLOGICAL AGENT E. COLI 0157:H7

EPA Science Inventory

Rapid viability tests of the Catagory B agent Escherichia coli O157:H7 were evaluated after disinfection with chlorine. The metabolic activity dyes ChemChrome V6, a modified fluorescein diacetate (FDA) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) were compared to standard ...

Multiple storm event impacts on epikarst storage and transport of organic soil amendments in South-Central Kentucky.

USDA-ARS?s Scientific Manuscript database

The groundwater in agricultural karst areas is susceptible to contamination from organic soil amendments and pesticides. During major storm events of winter and spring 2011, dye traces were initiated using sulphorhodamine-B, fluorescein and eosine in a known groundwater recharge area where manure wa...

Inhibition of Oncogenic functionality of STAT3 Protein by Membrane Anchoring

NASA Astrophysics Data System (ADS)

Liu, Baoxu; Fletcher, Steven; Gunning, Patrick; Gradinaru, Claudiu

2009-03-01

Signal Transducer and Activator of Transcription 3 (STAT3) protein plays an important role in oncogenic processes. A novel molecular therapeutic approach to inhibit the oncogenic functionality of STAT3 is to design a prenylated small peptide sequence which could sequester STAT3 to the plasma membrane. We have also developed a novel fluorescein derivative label (F-NAc), which is much more photostable compared to the popular fluorescein label FITC. Remarkably, the new dye shows fluorescent properties that are invariant over a wide pH range, which is advantageous for our application. We have shown that F-NAc is suitable for single-molecule measurements and its properties are not affected by ligation to biomolecules. The membrane localization via high-affinity prenylated small-molecule binding agents is studied by encapsulating FNAc-labeled STAT3 and inhibitors within a liposome model cell system. The dynamics of the interaction between the protein and the prenylated ligands is investigated at single molecule level. The efficiency and stability of the STAT3 anchoring in lipid membranes are addressed via quantitative confocal imaging and single-molecule spectroscopy using a custom-built multiparameter fluorescence microscope.

Simple apparatus for polarization sensing of analytes

NASA Astrophysics Data System (ADS)

Gryczynski, Zygmunt; Gryczynski, Ignacy; Lakowicz, Joseph R.

2000-09-01

We describe a simple device for fluorescence sensing based on an unexpansive light source, a dual photocell and a Watson bridge. The emission is detected from two fluorescent samples, one of which changes intensity in response to the analyte. The emission from these two samples is observed through two orthogonally oriented polarizers and an analyzer polarizer. The latter polarizer is rotated to yield equal intensities from both sides of the dual photocell, as determined by a zero voltage from the Watson bridge. Using this device, we are able to measure fluorescein concentration to an accuracy near 2% at 1 (mu) M fluorescein, and pH values accurate to +/- 0.02 pH units. We also use this approach with a UV hand lamp and a glucose-sensitive protein to measure glucose concentrations near 2 (mu) M to an accuracy of +/- 0.1 (mu) M. This approach requires only simple electronics, which can be battery powered. Additionally, the method is generic, and can be applied with any fluorescent sample that displays a change in intensity. One can imagine this approach being used to develop portable point-of-care clinical devices.

Midrange affinity fluorescent Zn(II) sensors of the Zinpyr family: syntheses, characterization, and biological imaging applications.

PubMed

Nolan, Elizabeth M; Jaworski, Jacek; Racine, Maryann E; Sheng, Morgan; Lippard, Stephen J

2006-11-27

The syntheses and photophysical characterization of ZP9, 2-{2-chloro-6-hydroxy-3-oxo-5-[(2-{[pyridin-2-ylmethyl-(1H-pyrrol-2-ylmethyl)amino]methyl}phenylamino)methyl]-3H-xanthen-9-yl}benzoic acid, and ZP10, 2-{2-chloro-6-hydroxy-5-[(2-{[(1-methyl-1H-pyrrol-2-ylmethyl)pyridin-2-ylmethylamino]methyl}phenylamino)methyl]-3-oxo-3H-xanthen-9-yl}benzoic acid, two asymmetrically derivatized fluorescein-based dyes, are described. These sensors each contain an aniline-based ligand moiety functionalized with a pyridyl-amine-pyrrole group and have dissociation constants for Zn(II) in the sub-micromolar (ZP9) and low-micromolar (ZP10) range, which we define as "midrange". They give approximately 12- (ZP9) and approximately 7-fold (ZP10) fluorescence turn-on immediately following Zn(II) addition at neutral pH and exhibit improved selectivity for Zn(II) compared to the di-(2-picolyl)amine-based Zinpyr (ZP) sensors. Confocal microscopy studies indicate that such asymmetrical fluorescein-based probes are cell permeable and Zn(II) responsive in vivo.

Midrange Affinity Fluorescent Zn(II) Sensors of the Zinpyr Family: Syntheses, Characterization, and Biological Imaging Applications

PubMed Central

Nolan, Elizabeth M.; Jaworski, Jacek; Racine, Maryann E.; Sheng, Morgan; Lippard, Stephen J.

2006-01-01

The syntheses and photophysical characterization of ZP9, 2-{2-chloro-6-hydroxy-3-oxo-5-[(2-{[pyridin-2-ylmethyl-(1H-pyrrol-2-ylmethyl)amino]methyl}phenylamino)methyl]-3H-xanthen-9-yl}benzoic acid, and ZP10, 2-{2-chloro-6-hydroxy-5-[(2-{[(1-methyl-1H-pyrrol-2-ylmethyl)pyridin-2-ylmethylamino]methyl}phenylamino)methyl]-3-oxo-3H-xanthen-9-yl}benzoic acid, two asymmetrically derivatized fluorescein-based dyes, are described. These sensors each contain an aniline-based ligand moiety functionalized with a pyridyl-amine-pyrrole group and have dissociation constants for Zn(II) in the sub-micromolar (ZP9) and low-micromolar (ZP10) range, which we define as “midrange”. They give ~12- (ZP9) and ~7-fold (ZP10) fluorescence turn-on immediately following Zn(II) addition at neutral pH and exhibit improved selectivity for Zn(II) compared to the di-(2-picolyl)amine-based Zinpyr (ZP) sensors. Confocal microscopy studies indicate that such asymmetrical fluorescein-based probes are cell permeable and Zn(II) responsive in vivo. PMID:17112271

Biotin and fluorescent labeling of RNA using T4 RNA ligase.

PubMed Central

Richardson, R W; Gumport, R I

1983-01-01

Biotin, fluorescein, and tetramethylrhodamine derivatives of P1-(6-aminohex-1-yl)-P2-(5'-adenosine) pyrophosphate were synthesized and used as substrates with T4 RNA ligase. In the absence of ATP, the non-adenylyl portion of these substrates is transferred to the 3'-hydroxyl of an RNA acceptor to form a phosphodiester bond and the AMP portion is released. E. coli and D. melanogaster 5S RNA, yeast tRNAPhe, (Ap)3C, and (Ap)3A serve as acceptors with yields of products varying from 50 to 100%. Biotin-labeled oligonucleotides are bound selectively and quantitatively to avidin-agarose and may be eluted with 6 M guanidine hydrochloride, pH 2.5. Fluorescein and tetramethylrhodamine-labeled oligonucleotides are highly fluorescent and show no quenching due to attachment to the acceptor. The diverse structures of the appended groups and of the chain lengths and compositions of the acceptor RNAs show that T4 RNA ligase will be a useful modification reagent for the addition of various functional groups to the 3'-terminus of RNA molecules. Images PMID:6194506

Fluorescein Diacetate Microplate Assay in Cell Viability Detection.

PubMed

Chen, Xi; Yang, Xiu-Ying; Fang, Lian-Hua; DU, Guan-Hua

2016-12-20

Objective To investigate the application of the fluorescein diacetate (FDA) microplate assay in cell viability detection. Methods Cells were seeded in a 96-well culture plate until detection. After incubated with FDA,the plate was detected by fluorescence microplate analyzer. The effects of FDA incubation duration,concentration,and other factors on the assay's accuracy and stability were assessed. We also compared the results of FDA with methyl thiazolyl(MTT) in terms of cell numbers and H 2 O 2 injury. Results Within 0-30 minutes,the fluorescence-cell number coefficient of FDA assay increased with duration and reached 0.99 in 27-30 minutes. The optimum concentration of final FDA in this study was 10-30 μg/ml. On cell viability detection,the result of FDA method was equivalent to MTT method. As to H 2 O 2 injury assay,the sensitivity of FDA method was superior to MTT on the higher concentration H 2 O 2 treatment due to a relative shorter duration for detection. Conclusion As a stable and reliable method,FDA is feasible for cell variability detection under varied conditions.

Allyl Fluorescein Ethers as Promising Fluorescent Probes for Carbon Monoxide Imaging in Living Cells.

PubMed

Feng, Shumin; Liu, Dandan; Feng, Weiyong; Feng, Guoqiang

2017-03-21

Recently, the fluorescent detection of carbon monoxide (CO) in living cells has attracted great attention. However, due to the lack of effective ways to construct fluorescent CO probes, fluorescent detection of CO in living cells is still in its infancy. In this paper, we report for the first time the use of allyl ether as a reaction site for construction of fluorescent CO probes. By this way, two readily available allyl fluorescein ethers were prepared, which were found to be highly selective and sensitive probes for CO in the presence of PdCl 2 . These probes have the merits of good stability, good water-solubility, and rapid and distinct colorimetric and remarkable fluorescent turn-on signal changes. Moreover, a very low dose of these two probes can be used to detect and track CO in living cells, indicating that these two probes could be very promising biological tools for CO detection in living systems. Overall, this work provided not only two new promising fluorescent CO probes but also a new way to devise fluorescent CO probes.

Human Alveolar Macrophages May Not Be Susceptible to Direct Infection by a Human Influenza Virus.

PubMed

Ettensohn, David B; Frampton, Mark W; Nichols, Joan E; Roberts, Norbert J

2016-12-01

The current studies were undertaken to determine the susceptibility of human alveolar macrophages (AMs) to influenza A virus (IAV) infection in comparison with autologous peripheral blood-derived monocytes-macrophages (PBMs). AMs and PBMs were exposed to IAV in vitro and examined for their ability to bind and internalize IAV, and synthesize viral proteins and RNA. PBMs but not AMs demonstrated binding and internalization of the virus, synthesizing viral proteins and RNA. Exposure of AMs in the presence of a sialidase inhibitor or anti-IAV antibody resulted in viral protein synthesis by the cells. Exposure of AMs to fluorescein isothiocyanate-labeled IAV in the presence of anti-fluorescein isothiocyanate antibody also resulted in viral protein synthesis. Thus, human AMs are apparently not susceptible to direct infection by a human IAV but are likely to be infected indirectly in the setting of exposure in the presence of antibody that binds the challenging strain of IAV. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Estimating retinal vascular permeability using the adiabatic approximation to the tissue homogeneity model with fluorescein videoangiography

NASA Astrophysics Data System (ADS)

Tichauer, Kenneth M.; Osswald, Christian R.; Dosmar, Emily; Guthrie, Micah J.; Hones, Logan; Sinha, Lagnojita; Xu, Xiaochun; Mieler, William F.; St. Lawrence, Keith; Kang-Mieler, Jennifer J.

2015-06-01

Clinical symptoms of diabetic retinopathy are not detectable until damage to the retina reaches an irreversible stage, at least by today's treatment standards. As a result, there is a push to develop new, "sub-clinical" methods of predicting the onset of diabetic retinopathy before the onset of irreversible damage. With diabetic retinopathy being associated with the accumulation of long-term mild damage to the retinal vasculature, retinal blood vessel permeability has been proposed as a key parameter for detecting preclinical stages of retinopathy. In this study, a kinetic modeling approach used to quantify vascular permeability in dynamic contrast-enhanced medical imaging was evaluated in noise simulations and then applied to retinal videoangiography data in a diabetic rat for the first time to determine the potential for this approach to be employed clinically as an early indicator of diabetic retinopathy. Experimental levels of noise were found to introduce errors of less than 15% in estimates of blood flow and extraction fraction (a marker of vascular permeability), and fitting of rat retinal fluorescein angiography data provided stable maps of both parameters.

Extraction of Capillary Non-perfusion from Fundus Fluorescein Angiogram

NASA Astrophysics Data System (ADS)

Sivaswamy, Jayanthi; Agarwal, Amit; Chawla, Mayank; Rani, Alka; Das, Taraprasad

Capillary Non-Perfusion (CNP) is a condition in diabetic retinopathy where blood ceases to flow to certain parts of the retina, potentially leading to blindness. This paper presents a solution for automatically detecting and segmenting CNP regions from fundus fluorescein angiograms (FFAs). CNPs are modelled as valleys, and a novel technique based on extrema pyramid is presented for trough-based valley detection. The obtained valley points are used to segment the desired CNP regions by employing a variance-based region growing scheme. The proposed algorithm has been tested on 40 images and validated against expert-marked ground truth. In this paper, we present results of testing and validation of our algorithm against ground truth and compare the segmentation performance against two others methods.The performance of the proposed algorithm is presented as a receiver operating characteristic (ROC) curve. The area under this curve is 0.842 and the distance of ROC from the ideal point (0,1) is 0.31. The proposed method for CNP segmentation was found to outperform the watershed [1] and heat-flow [2] based methods.

Lysozyme as a recognition element for monitoring of bacterial population.

PubMed

Zheng, Laibao; Wan, Yi; Yu, Liangmin; Zhang, Dun

2016-01-01

Bacterial infections remain a significant challenge in biomedicine and environment safety. Increasing worldwide demand for point-of-care techniques and increasing concern on their safe development and use, require a simple and sensitive bioanalysis for pathogen detection. However, this goal is not yet achieved. A design for fluorescein isothiocyanate-labeled lysozyme (FITC-LYZ), which provides quantitative binding information for gram-positive bacteria, Micrococcus luteus, and detects pathogen concentration, is presented. The functional lysozyme is used not only as the pathogenic detection platform, but also as a tracking reagent for microbial population in antibacterial tests. A nonlinear relationship between the system response and the logarithm of the bacterial concentration was observed in the range of 1.2×10(2)-1.2×10(5) cfu mL(-1). The system has a potential for further applications and provides a facile and simple method for detection of pathogenic bacteria. Meanwhile, the fluorescein isothiocyanate -labeled lysozyme is also employed as the tracking agent for antibacterial dynamic assay, which show a similar dynamic curve compared with UV-vis test. Copyright © 2015 Elsevier B.V. All rights reserved.

The Effects of Heteroatoms Si and S on Tuning the Optical Properties of Rhodamine- and Fluorescein-Based Fluorescence Probes: A Theoretical Analysis.

PubMed

Zhou, Panwang; Ning, Cai; Alsaedi, Ahmed; Han, Keli

2016-10-05

The effects of the incorporated heteroatoms Si and S on tuning the optical properties of rhodamine- and fluorescein-based fluorescence probes is investigated using DFT and time-dependent DFT with four different functionals. As previously proposed, the large redshift (90 nm) produced by a Si atom in both the absorption and emission spectra can be attributed to the σ*-π* conjugation between the σ* orbital of the Si atom and the π* orbital of the adjacent carbon atoms. However, the presence of a Si atom does not alter the fluorescence quenching mechanism of the nonfluorescent forms of the investigated compounds. For the first time, these theoretical results indicate that the n orbital of the S atom plays an important role in determining the optical properties of the nonfluorescent form of rhodamine-based fluorescence probes. It alters the fluorescence quenching mechanism by lowering the energy of the dark nπ* state, which is due to breakage of the C10-S52 bond upon photoexcitation. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparison of adaptive optics scanning light ophthalmoscopic fluorescein angiography and offset pinhole imaging

PubMed Central

Chui, Toco Y. P.; Dubow, Michael; Pinhas, Alexander; Shah, Nishit; Gan, Alexander; Weitz, Rishard; Sulai, Yusufu N.; Dubra, Alfredo; Rosen, Richard B.

2014-01-01

Recent advances to the adaptive optics scanning light ophthalmoscope (AOSLO) have enabled finer in vivo assessment of the human retinal microvasculature. AOSLO confocal reflectance imaging has been coupled with oral fluorescein angiography (FA), enabling simultaneous acquisition of structural and perfusion images. AOSLO offset pinhole (OP) imaging combined with motion contrast post-processing techniques, are able to create a similar set of structural and perfusion images without the use of exogenous contrast agent. In this study, we evaluate the similarities and differences of the structural and perfusion images obtained by either method, in healthy control subjects and in patients with retinal vasculopathy including hypertensive retinopathy, diabetic retinopathy, and retinal vein occlusion. Our results show that AOSLO OP motion contrast provides perfusion maps comparable to those obtained with AOSLO FA, while AOSLO OP reflectance images provide additional information such as vessel wall fine structure not as readily visible in AOSLO confocal reflectance images. AOSLO OP offers a non-invasive alternative to AOSLO FA without the need for any exogenous contrast agent. PMID:24761299

Ultra wide field fluorescein angiography can detect macular pathology in central retinal vein occlusion.

PubMed

Tsui, Irena; Franco-Cardenas, Valentina; Hubschman, Jean-Pierre; Yu, Fei; Schwartz, Steven D

2012-01-01

The purpose of this study was to evaluate whether ultra wide field fluorescein angiography (UWFFA), a tool established for the detection of peripheral non-perfusion, can also detect macular pathology. A retrospective imaging review was performed on patients with central retinal vein occlusion. UWFFA was graded for angiographic leakage (petalloid and/or diffuse leakage) and presence of abnormalities in the foveal avascular zone and was then correlated to spectral-domain optical coherence tomography (SD-OCT). Sixty-six eyes met inclusion criteria. Intergrader agreement was highly reliable for grading macular leakage on UWFFA (kappa = 0.75) and moderately reliable for the evaluation of an abnormal foveal avascular zone (kappa = 0.43). Angiographic leakage on UWFFA correlated to macular edema on SD-OCT (P > .0001), and abnormalities in the foveal avascular zone on UWFFA correlated to ganglion cell layer atrophy on SD-OCT (P = .0002). Intergrader reliability in grading UWFFA was better when assessing macular leakage than when assessing macular ischemia. UWFFA findings correlated to macular edema and signs of macular ischemia on SD-OCT. Copyright 2012, SLACK Incorporated.

Blood-Brain Barrier Disruption, Sodium Fluorescein, And Fluorescence-Guided Surgery Of Gliomas.

PubMed

Xiang, Yan; Zhu, Xiao-Peng; Zhao, Jian-Nong; Huang, Guo-Hao; Tang, Jun-Hai; Chen, Huan-Ran; Du, Lei; Zhang, Dong; Tang, Xue-Feng; Yang, Hui; Lv, Sheng-Qing

2018-01-22

Sodium fluorescein (SF) is an ideal dye for intraoperative guided-resection of high-grade gliomas (HGGs). However, it is not well understood whether the SF-guided technique is suitable for different grades of gliomas, and the correlation between fluorescence and pathology is also not yet clear. In this study, we investigated 28 patients, including 23 patients with HGG and 5 patients with low-grade glioma (LGG). All patients were treated using the SF-guided technique on a Pentero 900 microscope (Carl Zeiss, Oberkochen, Germany). Claudin-5 immunohistochemical (IHC) staining for the tumours and peritumour tissues was analyzed. Intraoperative yellow fluorescence was noted in all the HGGs but not in the LGGs. Claudin-5 expression in the blood brain barrier endothelial cells was downregulated and disconnected in the HGGs (p < 0.05), but had no difference or slightly decreased in the LGGs (p > 0.05). The SF-guided technique is suitable for HGG surgery but not for LGG surgery. Downregulation of claudin-5 expression may contribute to the presence of yellow fluorescence in the glioma in SF-guided surgery.

Differentiation of Cariogenic Streptococci by Fluorescent Antibody1

PubMed Central

Jablon, James M.; Zinner, Doran D.

1966-01-01

Jablon, J. M. (University of Miami, Miami, Fla.), and D. D. Zinner. Differentiation of cariogenic streptococci by fluorescent antibody. J. Bacteriol. 92:1590–1596. 1966.—Eight strains of streptococci were isolated from human carious lesions by the fluorescent-antibody (FA) technique. Seven of these strains produced experimental caries in hamsters or rats maintained on a high sucrose diet. The eighth strain was noncariogenic in animals but possessed some antigenic components in common with the cariogenic strains. On the basis of antigen-antibody reactions by microprecipitin and agar-gel diffusion patterns, the strains were divided into four groups; these groups differed with regard to their cariogenic activity in hamsters. Fluorescein-conjugated antisera, prepared against the human strains, showed some cross-reactions which interfered with the efficacy of the FA technique in differentiating between the related streptococcal groups. To eliminate these cross-reactions, a small amount of related-strain antisera was added to the fluorescein-conjugated antisera to the cariogenic strains. This technique is effective in blocking cross-reactions and should be tried wherever cross-reactions are encountered in the FA technique. Images PMID:5334765

ORAC-fluorescein assay to determine the oxygen radical absorbance capacity of resveratrol complexed in cyclodextrins.

PubMed

Lucas-Abellán, C; Mercader-Ros, M T; Zafrilla, M P; Fortea, M I; Gabaldón, J A; Núñez-Delicado, E

2008-03-26

The effect of the complexation of resveratrol with hydroxypropyl-beta-cyclodextrins (HP-beta-CDs) on the antioxidant capacity of the polyphenol is studied for the first time by means of the oxygen radical absorbance capacity (ORAC) method, using fluorescein (FL) as the fluorescent probe. The method is validated through its linearity, precision, and accuracy for measuring the ORAC of resveratrol in the absence or presence of cyclodextrins (CDs). The complexation of resveratrol in CDs increased the net area under the FL decay curve (net AUC) of resveratrol up to its saturation level, at which the polyphenol showed almost double the antioxidant activity it shows in the absence of CDs. The complexation constant ( K c) between resveratrol and HP-beta-CDs was calculated by linear regression of the phase solubility diagram ( K c = 18048 M (-1)). The antioxidant activity of resveratrol was dependent on the complexed resveratrol because CDs acts as a controlled dosage reservoir that protects resveratrol against rapid oxidation by free radicals. In this way, its antioxidant activity is prolonged and only reaches its maximum when all the resveratrol is complexed.

Effectiveness and relevant factors of 2% rebamipide ophthalmic suspension treatment in dry eye.

PubMed

Ueda, Kaori; Matsumiya, Wataru; Otsuka, Keiko; Maeda, Yoshifumi; Nagai, Takayuki; Nakamura, Makoto

2015-06-06

Rebamipide with mucin secretagogue activity was recently approved for the treatment of dry eye. The efficacy and safety in the treatment of rebamipide were shown in two pivotal clinical trials. It was the aim of this study to evaluate the effect of 2% rebamipide ophthalmic suspension in patients with dry eye and analyze relevant factors for favorable effects of rebamipide in clinical practice. This was a retrospective cohort study of 48 eyes from 24 patients with dry eye treated with 2% rebamipide ophthalmic suspension. Dry eye-related symptom score, tear film break-up time (TBUT), fluorescein ocular surface staining score (FOS) and the Schirmer test were used to collect the data from patients at baseline, and at 2, 4, 8, and 12 week visits. To determine the relevant factors, multiple regression analyses were then performed. Mean dry eye-related symptom score showed a significant improvement from the baseline (14.5 points) at 2, 4, 8 and 12 weeks (9.80, 7.04, 7.04 and 7.83 points, corrected P value < 0.001, respectively). Median FOS showed a significant improvement from the baseline (3.0 points) at 2, 4, 8 and 12 weeks (2.0, 2.0, 1.0 and 1.0 points, corrected P value < 0.001, respectively). TBUT and Schirmer test values were not significantly improved after the treatment. For ocular symptoms, three parameters (foreign body sensation, dry eye sensation and ocular discomfort) showed significant improvements at all visits. The multiple regression analyses showed that the fluorescein conjunctiva staining score was significantly correlated with the changes of dry eye-related symptom score at 12 weeks (P value = 0.017) and dry eye-related symptom score was significantly correlated with independent variables for the changes of FOS at 12 weeks (P value = 0.0097). Two percent rebamipide ophthalmic suspension was an effective therapy for dry eye patients. Moreover the fluorescein conjunctiva staining score and dry eye-related symptom score might be good relevant factors for favorable effects of rebamipide.

Fluorometric graphene oxide-based detection of Salmonella enteritis using a truncated DNA aptamer.

PubMed

Chinnappan, Raja; AlAmer, Saleh; Eissa, Shimaa; Rahamn, Anas Abdel; Abu Salah, Khalid M; Zourob, Mohammed

2017-12-18

The work describes a fluorescence-based study for mapping the highest affinity truncated aptamer from the full length sequence and its integration in a graphene oxide platform for the detection of Salmonella enteriditis. To identify the best truncated sequence, molecular beacons and a displacement assay design are applied. In the fluorescence displacement assay, the truncated aptamer was hybridized with fluorescein and quencher-labeled complementary sequences to form a fluorescence/quencher pair. In the presence of S. enteritidis, the aptamer dissociates from the complementary labeled oligonucleotides and thus, the fluorescein/quencher pair becomes physically separated. This leads to an increase in fluorescence intensity. One of the truncated aptamers identified has a 2-fold lower dissociation constant (3.2 nM) compared to its full length aptamer (6.3 nM). The truncated aptamer selected in this process was used to develop a fluorometric graphene oxide (GO) based assay. If fluorescein-labeled aptamer is adsorbed on GO via π stacking interaction, fluorescence is quenched. However, in the presence of target (S. enteriditis), the labeled aptamers is released from surface to form a stable complex with the bacteria and fluorescence is restored, depending on the quantity of bacteria being present. The resulting assay has an unsurpassed detection limit of 25 cfu·mL -1 in the best case. The cross reactivity to Salmonella typhimurium, Staphylococcus aureus and Escherichia coli is negligible. The assay was applied to analyze doped milk samples for and gave good recovery. Thus, we believe that the truncated aptamer/graphene oxide platform is a potential tool for the detection of S. Enteritidis. Graphical abstract Fluorescently labelled aptamer against Salmonella enteritidis was adsorbed on the surface of graphene oxide by π-stacking interaction. This results in quenching of the fluorescence of the label. Addition of Salmonella enteritidis restores fluorescence, and this effect is used for quantification of this food-borne pathogen.

Mechanisms of superficial micropunctate corneal staining with sodium fluorescein: the contribution of pooling.

PubMed

Bandamwar, Kalika L; Garrett, Qian; Papas, Eric B

2012-04-01

To establish if sodium fluorescein (SFL) dye accumulation within intercellular spaces on the ocular surface contributes to the appearance of superficial punctate corneal staining. Thirteen subjects bilaterally wore PureVision™ lenses that had been pre-soaked in ReNu MultiPlus® multipurpose solution. After 1h of lens wear, corneal staining with SFL was assessed using a standard slit-lamp technique. Participants who presented with bilateral, corneal staining were selected for further evaluation. A randomly selected eye was rinsed with saline three times. Fellow eyes (control) received no rinsing. After each rinse, the appearance of SFL staining was recorded without any further instillation of the dye. To eliminate any confounding effects of staining due to residual fluorescein in the tear menisci, corneal staining was induced in freshly excised, isolated, rabbit eyes by topical administration of 0.001% PHMB and staining, rinsing and grading were performed as above. Nine out of 13 subjects presented with bilateral diffuse corneal staining (mean grade±SD: 2.4±0.7). The mean staining grades in test and control eyes respectively after each of the three rinses were (1) 2.41±0.41, 2.25±0.69 (p=0.9); (2) 2.34±0.79, 2.1±0.83 (p=0.8); and (3) 1.71±0.65, 1.60±0.79 (p=0.6) there was no significant reduction in staining with rinsing (p>0.05) and no difference was observed between test and control eyes at any sampling-point. Similar observations made in ex vivo rabbit eyes replicated these results. Pooling or accumulation of SFL solution within intercellular spaces does not appear to contribute to the appearance of superficial micropunctate corneal staining. Copyright © 2011 British Contact Lens Association. Published by Elsevier Ltd. All rights reserved.

Femtosecond laser ablation of porcine intestinal mucosa: potential autologous transplant for segmental cystectomy

NASA Astrophysics Data System (ADS)

Higbee, Russell G.; Irwin, Bryan S.; Nguyen, Michael N.; Zhang, Yuanyuan; Warren, William L.

2005-04-01

Nearly 80% of patients with newly diagnosed bladder cancer present with superficial bladder tumors (confined to the bladder lining such as transitional cell carcinoma [90%], squamous cell carcinoma [6-8%], and adenocarcinoma[2%]) in stages Ta, Tis, or T1. Segmental cystectomy is one surgical treatment for patients who have a low-grade invasive tumor. Transposition of small intestine is a viable surgical treatment option. Success of the transplantation is also dependent upon removal of the entire SI mucosal layer. A Clark Spitfire Ti:Sapphire laser operating at 775 nm and 1 kHz repetition rate, was used to investigate the damage induced to fresh cadaveric porcine small intestinal mucosal epithelium. The laser was held constant at a focal spot diameter of 100 μm using a 200 mm focal point lens, with a power output maximum of 257 mW. A high resolution motorized X-Y-Z stage translated the SI tissue through the beam at 500 μm/sec with a line spacing of 50 μm. This produced a 50% overlap in the laser etching for each pass over a 1 cm x 1.5 cm grid. To determine if the mucosal lining of the SI was adequately removed, the targeted area was covered with 1% fluorescein solution for 30 seconds and then rinsed with phosphate buffered saline. Fluorescein staining was examined under UV illumination, to determine the initial degree of mucosal removal. Tissues were fixed and processed for light and scanning electron microscopy by standard protocols. Brightfield light microscopy of hematoxylin and eosin stained 4 μm thick cross sections, scanning electron microscopy were examined to determine the degree of mucosal tissue removal. Clear delineation of the submucosal layer by fluorescein staining was also observed. The Ti:Sapphire laser demonstrated precise, efficient removal of the mucosal epithelium with minimal submucosal damage.

Novel Fluorescein Angiography-Based Computer-Aided Algorithm for Assessment of Retinal Vessel Permeability

PubMed Central

Chassidim, Yoash; Parmet, Yisrael; Tomkins, Oren; Knyazer, Boris; Friedman, Alon; Levy, Jaime

2013-01-01

Purpose To present a novel method for quantitative assessment of retinal vessel permeability using a fluorescein angiography-based computer algorithm. Methods Twenty-one subjects (13 with diabetic retinopathy, 8 healthy volunteers) underwent fluorescein angiography (FA). Image pre-processing included removal of non-retinal and noisy images and registration to achieve spatial and temporal pixel-based analysis. Permeability was assessed for each pixel by computing intensity kinetics normalized to arterial values. A linear curve was fitted and the slope value was assigned, color-coded and displayed. The initial FA studies and the computed permeability maps were interpreted in a masked and randomized manner by three experienced ophthalmologists for statistical validation of diagnosis accuracy and efficacy. Results Permeability maps were successfully generated for all subjects. For healthy volunteers permeability values showed a normal distribution with a comparable range between subjects. Based on the mean cumulative histogram for the healthy population a threshold (99.5%) for pathological permeability was determined. Clear differences were found between patients and healthy subjects in the number and spatial distribution of pixels with pathological vascular leakage. The computed maps improved the discrimination between patients and healthy subjects, achieved sensitivity and specificity of 0.974 and 0.833 respectively, and significantly improved the consensus among raters for the localization of pathological regions. Conclusion The new algorithm allows quantification of retinal vessel permeability and provides objective, more sensitive and accurate evaluation than the present subjective clinical diagnosis. Future studies with a larger patients’ cohort and different retinal pathologies are awaited to further validate this new approach and its role in diagnosis and treatment follow-up. Successful evaluation of vasculature permeability may be used for the early diagnosis of brain microvascular pathology and potentially predict associated neurological sequelae. Finally, the algorithm could be implemented for intraoperative evaluation of micovascular integrity in other organs or during animal experiments. PMID:23626701

Influence on ocular surface after cataract surgery and effect of topical diquafosol on postoperative dry eye: a multicenter prospective randomized study.

PubMed

Miyake, Kensaku; Yokoi, Norihiko

2017-01-01

To investigate influences of 3% diquafosol sodium ophthalmic solution (DQS) on ocular surface after cataract surgery and effects on postoperative dry eye. This study had two consecutive prospective study phases. The former was an observational study from before cataract surgery to 4 weeks after surgery and the latter was a randomized open-label study from 4 to 8 weeks after surgery. Subjects were 433 eyes of 433 patients undergoing cataract surgery with intraocular lens implantation. Dry eye examination of tear breakup time (BUT), corneal and conjunctival fluorescein staining scores, total subjective symptom score (12 symptoms), and Schirmer I test were conducted before surgery and 4 weeks after surgery. Patient demographics and these examination results were used to analyze risk factors to predict postoperative dry eye. In a randomized study, 154 eyes diagnosed with dry eye postoperatively were applied either DQS or artificial tears (AT) six times daily for 4 weeks. The data of the examinations were compared. At 4 weeks after surgery, BUT was shortened significantly ( P =0.036), fluorescein staining score increased significantly ( P =0.012), but total subjective symptom score was significantly improved ( P <0.001). The majority of postoperative dry eye was shortened BUT type (53.1%). The dry eye prevalence after surgery decreased (55.7%) compared with before surgery (69.7%). Females and the patient with dry eye symptoms before surgery had significant risk factors for postoperative dry eye. In a randomized study, BUT was significantly prolonged in the DQS group ( P =0.015), but not in the AT group. Fluorescein staining score was significantly improved in both groups ( P <0.001). Total subjective symptom score was significantly decreased in the AT group ( P <0.001), but not in the DQS group. Our study suggests that cataract surgery has harmful effects on tear film stability and ocular surface, and DQS has a capability to improve them.

Patterns of peripheral retinal and central macula ischemia in diabetic retinopathy as evaluated by ultra-widefield fluorescein angiography.

PubMed

Sim, Dawn A; Keane, Pearse A; Rajendram, Ranjan; Karampelas, Michael; Selvam, Senthil; Powner, Michael B; Fruttiger, Marcus; Tufail, Adnan; Egan, Catherine A

2014-07-01

To investigate the association between peripheral and central ischemia in diabetic retinopathy. Retrospective, cross-sectional. Consecutive ultra-widefield fluorescein angiography images were collected from patients with diabetes over a 12-month period. Parameters quantified include the foveal avascular zone (FAZ) area, peripheral ischemic index, peripheral leakage index, and central retinal thickness measurements, as well as visual acuity. The peripheral ischemia or leakage index was calculated as the area of capillary nonperfusion or leakage, expressed as a percentage of the total retinal area. Forty-seven eyes of 47 patients were included. A moderate correlation was observed between the peripheral ischemia index and FAZ area (r = 0.49, P = .0001). A moderate correlation was also observed between the peripheral leakage index and FAZ area, but only in eyes that were laser naïve (r = 0.44, P = .02). A thinner retina was observed in eyes with macular ischemia (217 ± 81.8 μm vs 272 ± 36.0 μm) (P = .02), but not peripheral ischemia (258 ± 76.3 μm vs 276 ± 68.0 μm) (P = .24). The relationships between different patterns of peripheral and central macular pathology and visual acuity were evaluated in a step-wise multivariable regression model, and the variables that remained independently associated were age (r = 0.33, P = .03), FAZ area (r = 0.45, P = .02), and central retinal thickness (r = 0.38, P = .01), (R(2)-adjusted = 0.36). Ultra-widefield fluorescein angiography provides an insight into the relationships between diabetic vascular complications in the retinal periphery and central macula. Although we observed relationships between ischemia and vascular leakage in the macula and periphery, it was only macular ischemia and retinal thinning that was independently associated with a reduced visual function. Copyright © 2014 Elsevier Inc. All rights reserved.

NEW ULTRA-WIDE-FIELD ANGIOGRAPHIC GRADING SCHEME FOR RADIATION RETINOPATHY AFTER IODINE-125 BRACHYTHERAPY FOR UVEAL MELANOMA.

PubMed

McCannel, Tara A; Kim, EunAh; Kamrava, Mitchell; Lamb, James; Caprioli, Joseph; Yang, Dong; McCannel, Colin A

2017-10-06

Radiation retinopathy remains incompletely characterized and may cause severe vision loss. Ultra-wide-field fluorescein angiography provides a pan-fundus view of vascular alterations caused by radiation treatment and may predict visual and ocular outcomes. We have developed a grading scheme to describe pan-fundus severity and to predict the progression of radiation retinopathy in patients treated for uveal melanoma with iodine-125 brachytherapy. A retrospective review of patients treated with standard iodine-125 brachytherapy for uveal melanoma at the Ophthalmic Oncology Center at the University of California, Los Angeles, who had undergone both baseline and postbrachytherapy ultra-wide-field fluorescein angiography. A grading scheme was devised based on observations of vascular leakage, retinal perfusion status, and retinal proliferation. The correlation of grade severity with patient characteristics, tumor features, visual acuity, optical coherence tomography findings, and neovascular glaucoma was measured with chi-square and one-way analysis of variance analyses. Sixty-seven patients were identified for review. Consistent wide-field angiographic patterns after brachytherapy were observed and graded as follows: Grade 0: normal; Grade 1: late foveal leakage; Grade 2: late peripheral leakage; Grade 3: presence of nonperfusion; and Grade 4: retinal neovascularization. Six eyes (8.9%) were Grade 0; 16 (23.8%) were Grade 1; 25 (37.3%) were Grade 2; 16 (23.4%) were Grade 3; and 4 (6.0%) were Grade 4. Higher grade radiation severity correlated significantly with duration of follow-up (P < 0.02); younger age (P = 0.035); worse visual acuity (P = 0.001); cystoid macular edema or atrophy on optical coherence tomography (P < 0.0001); and neovascular glaucoma (P = 0.003). Wide-field fluorescein angiography revealed distinct fundus-wide patterns of vascular damage, which were progressive in nature in eyes treated with iodine-125 brachytherapy for uveal melanoma and correlated with signs of progressive vascular injury. This grading scheme may have prognostic value to predict the progression of radiation retinopathy and to prognosticate visual outcomes in patients undergoing brachytherapy.

Influence on ocular surface after cataract surgery and effect of topical diquafosol on postoperative dry eye: a multicenter prospective randomized study

PubMed Central

Miyake, Kensaku; Yokoi, Norihiko

2017-01-01

Purpose To investigate influences of 3% diquafosol sodium ophthalmic solution (DQS) on ocular surface after cataract surgery and effects on postoperative dry eye. Design This study had two consecutive prospective study phases. The former was an observational study from before cataract surgery to 4 weeks after surgery and the latter was a randomized open-label study from 4 to 8 weeks after surgery. Methods Subjects were 433 eyes of 433 patients undergoing cataract surgery with intraocular lens implantation. Dry eye examination of tear breakup time (BUT), corneal and conjunctival fluorescein staining scores, total subjective symptom score (12 symptoms), and Schirmer I test were conducted before surgery and 4 weeks after surgery. Patient demographics and these examination results were used to analyze risk factors to predict postoperative dry eye. In a randomized study, 154 eyes diagnosed with dry eye postoperatively were applied either DQS or artificial tears (AT) six times daily for 4 weeks. The data of the examinations were compared. Results At 4 weeks after surgery, BUT was shortened significantly (P=0.036), fluorescein staining score increased significantly (P=0.012), but total subjective symptom score was significantly improved (P<0.001). The majority of postoperative dry eye was shortened BUT type (53.1%). The dry eye prevalence after surgery decreased (55.7%) compared with before surgery (69.7%). Females and the patient with dry eye symptoms before surgery had significant risk factors for postoperative dry eye. In a randomized study, BUT was significantly prolonged in the DQS group (P=0.015), but not in the AT group. Fluorescein staining score was significantly improved in both groups (P<0.001). Total subjective symptom score was significantly decreased in the AT group (P<0.001), but not in the DQS group. Conclusion Our study suggests that cataract surgery has harmful effects on tear film stability and ocular surface, and DQS has a capability to improve them. PMID:28360509

Additive Effect of preservative-free sodium hyaluronate 0.1% in treatment of dry eye syndrome with diquafosol 3% eye drops.

PubMed

Hwang, Ho Sik; Sung, Yoon-Mi; Lee, Weon Sun; Kim, Eun Chul

2014-09-01

The aim of this study was to evaluate the treatment effect of diquafosol 3% with preservative-free sodium hyaluronate 0.1% eye drops in dry eye syndrome. In total, 150 patients with dry eye syndrome were divided randomly into 3 groups. Group 1 (50 patients) was treated 4 times daily with preserved sodium hyaluronate 0.1%, group 2 (50 patients) was treated 4 times daily with diquafosol 3%, and group 3 (50 patients) was treated 4 times daily with diquafosol 3% and preservative-free sodium hyaluronate 0.1% eye drops for 3 months. Ocular surface disease index (OSDI) score, tear film break-up time, Schirmer I test, corneal fluorescein staining, and impression cytology were evaluated. There were significant improvements in the OSDI score, tear film break-up time, Schirmer I score, fluorescein and Rose Bengal staining, goblet cell density, and impression cytological findings in groups 2 and 3 compared with those for group 1 in patients with dry eye syndrome at 1, 2, and 3 months (P < 0.05). There were statistically significant improvements in the OSDI score (-8.48 ± 0.97, -5.69 ± 0.78; P = 0.02), fluorescein (-1.43 ± 0.21, -1.02 ± 0.18; P = 0.03), and Rose Bengal staining (-1.12 ± 0.26, -0.75 ± 0.12; P = 0.03), goblet cell density (89.65 ± 14.39, 70.36 ± 16.75; P = 0.03), and impression cytological findings (-0.53 ± 0.12, -0.34 ± 0.90; P = 0.01) in group 3 compared with those in group 2 at 3 months. Treatment with diquafosol 3% with preservative-free sodium hyaluronate 0.1% was more effective than diquafosol 3% monotherapy or treatment with preserved sodium hyaluronate 0.1% in dry eye syndrome. Preservative-free sodium hyaluronate 0.1% eye drops can increase the effect of diquafosol 3% in dry eye syndrome.

In vitro eye corrosion study of agrochemicals on isolated chicken eye.

PubMed

Buda, I; Budai, P; Szabó, R; Lehel, J

2013-01-01

Agrochemicals must undergo numberless toxicological tests before marketing. The eye irritation test is part of this test packet. Nowadays, OECD 405 can be used to classify the irritation potential of substances, the base of the OECD 405 guideline is the Draize test, which is one of the most criticized in vivo methods because of the injuries of the test animals and subjective nature of the test in recording the results. Therefore, several in vitro tests have been developed to replace totally or partly the in vivo eye irritation testing. The isolated chicken eye test method (OECD 438), which was used, is one of these alternative methods. Five different agrochemicals were examined in the following way: All test compounds were applied in a single dose onto the cornea of isolated chicken eyes in order to potentially classify the test compounds as ocular corrosive and/or severe irritant. The damages caused by the test substances were assessed by the determination of corneal swelling, opacity, fluorescein retention and morphological effects. These parameters were evaluated pre-treatment and starting at approximately 30, 75, 120, 180, and 240 minutes after the post-treatment rinse. The endpoints evaluated were corneal opacity, swelling, fluorescein retention and morphological effects. All of the endpoints, with the exception of fluorescein retention (which was determined only at pre-treatment and 30 minutes after test substance exposure) were determined at each of the above time points. Positive and negative controls were used and they showed the expected results in each study. In these in vitro eye corrosives and severe irritants studies, using the Isolated Chicken Eye model with five different products, no ocular corrosion or severe irritation potential were observed. These results correspond to the available information about the tested agrochemicals, so these studies with isolated chicken eye are considered to be successful.

Evaluation of Common Angling-Induced Sources of Epithelial Damage for Popular Freshwater Sport Fish using Fluorescein

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

Colotelo, Alison HA; Cooke, Steven J.

Angling is a popular recreational activity across the globe and a large proportion of fish captured by anglers are released due to voluntary or mandatory catch-and-release practices. The handling associated with hook removal and return of the fish to their environment can cause physical damage to the epidermal layer of the fish which may affect the condition and survival of released fish. This study investigated possible sources of epithelial damage associated with several different handling methods (i.e. landing net types, interactions with different boat floor surfaces, tournament procedures) commonly used in recreational angling for two popular freshwater sport fish species,more » largemouth bass (Micropterus salmoides) and northern pike (Esox lucius). Epithelial damage was examined using fluorescein, a non-toxic dye, which has been shown to detect latent epithelial damage. Northern pike exhibited extensive epithelial damage after exposure to several of the induced treatments (i.e., interaction with a carpeted surface, knotted nylon net, and line rolling) but relatively little epithelial damage when exposed to others (i.e., knotless rubber nets, smooth boat surfaces, or lip gripping devices). Largemouth bass did not show significant epithelial damage for any of the treatments, with the exception of fish caught in a semi-professional live release tournament. The detection of latent injuries using fluorescein can be an important management tool as it provides visual examples of potential damage that can be caused by different handling methods. Such visualizations can be used to encourage fish friendly angler behaviour and enhance the survival and welfare of released fish. It can also be used to test new products that are intended to or claim to reduce injury to fish that are to be released. Future research should evaluate the relationship between different levels of epithelial damage and mortality across a range of environmental conditions.« less

The effects of 3% diquafosol sodium application on the tear functions and ocular surface of the Cu,Zn-superoxide dismutase-1 (Sod1)-knockout mice.

PubMed

Kojima, Takashi; Dogru, Murat; Ibrahim, Osama M; Nagata, Taeko; Higa, Kazunari; Shimizu, Takahiko; Shirasawa, Takuji; Satake, Yoshiyuki; Shimazaki, Seika; Shimazaki, Jun; Tsubota, Kazuo

2014-01-01

To investigate the role of a water and mucin secretagogue (3% diquafosol sodium eye drops) on the tear function and conjunctival ocular surface changes in Sod1(-/-) in comparison to the wild-type (WT) mice. Fourteen eyes of 7 Sod1(-/-) male mice with C57BL/background and 14 eyes of 7 C57BL6 strain wild-type male mice were examined at 40 weeks in this study. All mice had application of 3% diquafosol ophthalmic solution six times a day for 2 weeks. Tear film stability and corneal epithelial damage was evaluated by fluorescein and Rose Bengal stainings. Anterior segment photography was performed before and after eye drop instillations. Aqueous tear quantity was measured with phenol red-impregnated cotton threads without anesthesia. Animals were sacrificed at 42 weeks after diquafosol treatment and the whole globe specimens were subjected to periodic acid Schiff staining. Goblet cell density was quantified by J Image software. Quantitative real-time PCR for conjunctival muc 5AC messenger RNA expression was also performed. Sod1(-/-) mice had significantly higher fluorescein staining scores compared to the WT mice before eye drop instillation. The mean tear film breakup time, Rose Bengal staining scores, and muc5 messenger RNA expression improved significantly with diquafosol treatment in both the WT and the knockout mice. The mean fluorescein staining score and aqueous tear quantity significantly improved in the Sod1(-/-) mice with treatment. A notable and consistent increase in goblet cells and decrease in inflammatory cell infiltrates could be confirmed in all specimens after 2 weeks of diquafosol eye drop application. Three percent diquafosol ophthalmic solution appears to be effective in the treatment of ocular surface disease in this age-related dry eye disease mouse model.

Transmural Intestinal Wall Permeability in Severe Ischemia after Enteral Protease Inhibition

PubMed Central

Altshuler, Angelina E.; Lamadrid, Itze; Li, Diana; Ma, Stephanie R.; Kurre, Leena; Schmid-Schönbein, Geert W.; Penn, Alexander H.

2014-01-01

In intestinal ischemia, inflammatory mediators in the small intestine's lumen such as food byproducts, bacteria, and digestive enzymes leak into the peritoneal space, lymph, and circulation, but the mechanisms by which the intestinal wall permeability initially increases are not well defined. We hypothesize that wall protease activity (independent of luminal proteases) and apoptosis contribute to the increased transmural permeability of the intestine's wall in an acutely ischemic small intestine. To model intestinal ischemia, the proximal jejunum to the distal ileum in the rat was excised, the lumen was rapidly flushed with saline to remove luminal contents, sectioned into equal length segments, and filled with a tracer (fluorescein) in saline, glucose, or protease inhibitors. The transmural fluorescein transport was determined over 2 hours. Villi structure and epithelial junctional proteins were analyzed. After ischemia, there was increased transmural permeability, loss of villi structure, and destruction of epithelial proteins. Supplementation with luminal glucose preserved the epithelium and significantly attenuated permeability and villi damage. Matrix metalloproteinase (MMP) inhibitors (doxycycline, GM 6001), and serine protease inhibitor (tranexamic acid) in the lumen, significantly reduced the fluorescein transport compared to saline for 90 min of ischemia. Based on these results, we tested in an in-vivo model of hemorrhagic shock (90 min 30 mmHg, 3 hours observation) for intestinal lesion formation. Single enteral interventions (saline, glucose, tranexamic acid) did not prevent intestinal lesions, while the combination of enteral glucose and tranexamic acid prevented lesion formation after hemorrhagic shock. The results suggest that apoptotic and protease mediated breakdown cause increased permeability and damage to the intestinal wall. Metabolic support in the lumen of an ischemic intestine with glucose reduces the transport from the lumen across the wall and enteral proteolytic inhibition attenuates tissue breakdown. These combined interventions ameliorate lesion formation in the small intestine after hemorrhagic shock. PMID:24805256

Preparation of fluorescently labeled silica nanoparticles using an amino acid-catalyzed seeds regrowth technique: Application to latent fingerprints detection and hemocompatibility studies.

PubMed

Abdelwahab, Walid M; Phillips, Edjohnier; Patonay, Gabor

2018-02-15

The efficiency of an amino acid catalyzed seed regrowth technique (ACSRT) in synthesizing twelve fluorescently labeled core-shell silica nanoparticles (FLSNPs) with tunable sizes, tailored hydrophobicity, low polydispersity as well as high labeling efficiency and minimized dye leakage using different combinations of organosilicate monomers and fluorophores have been systematically investigated in this report. The utilization of some of these FLSNPs in some applications that are facilitated by hydrophobicity such as developing and visualizing latent fingerprints (LFPs) on different surfaces was also investigated. The non-specific binding affinity of the developed nanoparticles to human serum albumin (HSA) and immunoglobulin G (IgG) has also been studied. Fluorescein, fluorescein isothiocyanate and its more hydrophilic butenamine derivative (WA6) have been used in this study. Also, the alkoxysilane precursor, tetraethoxyorthosilicate (TEOS) and its binary mixture with phenyltriethoxysilane (PTEOS) or 3-aminopropyl triethoxysilane (APTES) have been used in preparing the FLSNPs with tailored compositions for the core and shell of the nanoparticles. The mean diameters of the PTEOS-coated FLSNPs were between 33.4±5.9 and 42.2±10.8 nm as shown by the SEM measurements. The obtained results highlight the advantages of having a hydrophobic surface along with proper selection of the monomers forming the core to match the properties of the fluorescent reporters for clear detection of LFPs even using dyes of low hydrophobicity such as fluorescein and WA6. Furthermore, some of the developed FLSNPs were compared with bare silica nanoparticles in terms of nonspecific protein adsorption and hemolysis. The obtained results proved that the selected FLSNPs had a superior hemocompatibility in comparison with bare silica nanoparticles. These FLSNPs could also be used in some bio-related and diagnostic applications such as immunoassays and cell imaging purposes. Copyright © 2017 Elsevier Inc. All rights reserved.

Comparison of ultra-widefield fluorescein angiography with the Heidelberg Spectralis® noncontact ultra-widefield module versus the Optos® Optomap®

PubMed Central

Witmer, Matthew T; Parlitsis, George; Patel, Sarju; Kiss, Szilárd

2013-01-01

Purpose To compare ultra-widefield fluorescein angiography imaging using the Optos® Optomap® and the Heidelberg Spectralis® noncontact ultra-widefield module. Methods Five patients (ten eyes) underwent ultra-widefield fluorescein angiography using the Optos® panoramic P200Tx imaging system and the noncontact ultra-widefield module in the Heidelberg Spectralis® HRA+OCT system. The images were obtained as a single, nonsteered shot centered on the macula. The area of imaged retina was outlined and quantified using Adobe® Photoshop® C5 software. The total area and area within each of four visualized quadrants was calculated and compared between the two imaging modalities. Three masked reviewers also evaluated each quadrant per eye (40 total quadrants) to determine which modality imaged the retinal vasculature most peripherally. Results Optos® imaging captured a total retinal area averaging 151,362 pixels, ranging from 116,998 to 205,833 pixels, while the area captured using the Heidelberg Spectralis® was 101,786 pixels, ranging from 73,424 to 116,319 (P = 0.0002). The average area per individual quadrant imaged by Optos® versus the Heidelberg Spectralis® superiorly was 32,373 vs 32,789 pixels, respectively (P = 0.91), inferiorly was 24,665 vs 26,117 pixels, respectively (P = 0.71), temporally was 47,948 vs 20,645 pixels, respectively (P = 0.0001), and nasally was 46,374 vs 22,234 pixels, respectively (P = 0.0001). The Heidelberg Spectralis® was able to image the superior and inferior retinal vasculature to a more distal point than was the Optos®, in nine of ten eyes (18 of 20 quadrants). The Optos® was able to image the nasal and temporal retinal vasculature to a more distal point than was the Heidelberg Spectralis®, in ten of ten eyes (20 of 20 quadrants). Conclusion The ultra-widefield fluorescein angiography obtained with the Optos® and Heidelberg Spectralis® ultra-widefield imaging systems are both excellent modalities that provide views of the peripheral retina. On a single nonsteered image, the Optos® Optomap® covered a significantly larger total retinal surface area, with greater image variability, than did the Heidelberg Spectralis® ultra-widefield module. The Optos® captured an appreciably wider view of the retina temporally and nasally, albeit with peripheral distortion, while the ultra-widefield Heidelberg Spectralis® module was able to image the superior and inferior retinal vasculature more peripherally. The clinical significance of these findings as well as the area imaged on steered montaged images remains to be determined. PMID:23458976

Modification of the tear function index and its use in the diagnosis of Sjögren's syndrome

PubMed Central

Kaye, S.; Sims, G.; Willoughby, C.; Field, A.; Longman, L.; Brown, M.

2001-01-01

BACKGROUND—The tear function index (TFI) has been shown to be of value in the diagnosis of patients suffering from Sjögren's syndrome. It is dependent, however, on introducing into the conjunctival fornix the correct concentration of fluorescein in at least one and a half times the normal tear volume. The stimulus and effect of this added volume on the tear dynamics is likely to vary between individuals. These factors, together with the method of performing the test, limit its general applicability.
AIM—To devise a method of performing the TFI with less variability and more general applicability. To present a theoretical and in vitro assessment of the dynamics of the TFI.
METHOD—The study was divided into three parts. The first part was to compare the results obtained using a prepared strip containing 1.3 µl of 0.5% fluorescein with the introduction of the same amount of fluorescein as a drop. The second part was to compare the results obtained with prepared strips with the standard method of performing the TFI, both with and without topical anaesthetic. The third part was an in vitro study of the rate of flow of graded volumes on a filter paper strip. 42 subjects with a diagnosis of Sjögren's syndrome according to the European criteria and 126 without Sjögren's syndrome were included.
RESULTS—There was no significant difference between the results obtained with a prepared strip and the introduction of 1.3 µl into the eye before performing the Schirmer's test and TFI (0.1

Feasibility of simultaneous sodium fluorescein and indocyanine green injection in neurosurgical procedures.

PubMed

Acerbi, F; Restelli, F; Broggi, M; Schiariti, M; Ferroli, P

2016-07-01

The objective of this study is to assess the feasibility of simultaneous Sodium Fluorescein (SF) and Indocyanine Green (ICG) injection during neurosurgical procedures. Three patients harboring a high-grade glioma (HGG) were retrospectively identified in the surgical database of the Neurosurgical Unit 2 at the Foundation IRCCS Istituto Neurologico C. Besta in Milan, by having received intraoperatively both SF for tumor resection and ICG for vasculature angiographic studies in the same surgical procedure. We identified 2 males and 1 female (age range 25-60). Lesions were located in the left temporo-polar area and hippocampus (1 case), right superior frontal gyrus (1 case), left supplementary motor area (1 case). All the three lesions showed Magnetic Resonance Imaging (MRI) characteristics of HGG and, for this reason, in all patients a fluorescein-guided tumor removal was proposed. In the same surgical procedure ICG videoangiography was considered necessary in order to study arterial and venous vasculature, given by the strict relation of the tumor with an unexpected Posterior Communicating Artery (PComA) aneurysm in one case and with cortical drainage veins complexes in the other two cases. In all cases a microscope equipped with both YELLOW560 and IR800 integrated filters (Pentero 900, Carl Zeiss, Oberkorchen, Germany) was used. Fluorescein was i.v. injected at a dose of 5mg/kg immediately after patient intubation. ICG was i.v. injected in bolus on demand of the operating surgeon at a dose of 12.5mg. No side-effects related to simultaneous injection of SF and ICG were identified. In all three cases, the use of SF allowed to better visualize the tumor areas during surgical removal, thus leading to a radical resection until no macroscopic appearance of residual tumor mass and no fluorescence was visible in the surgical cavity. ICG videoangiography confirmed the patency of branches of internal carotid artery after clipping of an unexpected small PComA aneurysm found intraoperatively during tumor removal in one case, while in patient 2 and 3 it allowed to evaluate patency and study flow pattern in cortical drainage veins that were intimately related to the tumors and the way of the surgical approach. Postoperative MRI showed a Gross Total Resection of the tumors in all cases. This study showed for the first time the feasibility of intravenous SF injection and ICG videoangiography in the same surgical procedure. The presence of different fluorescence filters on the same surgical microscope allows the surgeon to recognize and safely resect the tumor and simultaneously evaluate local brain vascularization. Copyright © 2016 Elsevier B.V. All rights reserved.

Complementary Spectroscopic Assays for Investigating Protein-Ligand Binding Activity: A Project for the Advanced Chemistry Laboratory

ERIC Educational Resources Information Center

Mascotti, David P.; Waner, Mark J.

2010-01-01

A protein-ligand binding, guided-inquiry laboratory project with potential application across the advanced undergraduate curriculum is described. At the heart of the project are fluorescence and spectrophotometric assays utilizing biotin-4-fluorescein and streptavidin. The use of the same stock solutions for an assay that may be examined by two…

Bibliography of Soviet Laser Developments, Number 90, July-August 1987

DTIC Science & Technology

1988-06-27

8, 1987, 1250-1255. 51. Cernai, K.; Novakov , M. 0. Concentration dependence of stimulated emission in 4-methylumbelliferone-fluorescein laser dye...118 PANCHENKO I P 129 NADENENKO A V 29 NOVAK A 9 PANCHENKO V YA 18 NADEZHDINSKIY A 1 115 NOVAKOV 14 7 PANKOV F N 88 NAGORNYKH S N 121 NOVICHENKOV V YE

General Chemistry Laboratory Experiment to Demonstrate Organic Synthesis, Fluorescence, and Chemiluminescence through Production of a Biphasic Glow Stick

ERIC Educational Resources Information Center

Pay, Adam L.; Kovash, Curtiss; Logue, Brian A.

2017-01-01

A laboratory experiment is described for beginning, nonmajor chemistry students, which allows students to examine the phenomena of fluorescence and chemiluminescence, as well as gain experience in basic organic synthesis. Students synthesize fluorescein and bis(2,4,6-trichlorophenyl) oxalate (TCPO) to explore fluorescence and chemiluminescence by…

Bone Sialoproteins and Breast Cancer Detection

DTIC Science & Technology

2004-07-01

used to follow proteolytic activity on more natural macromolecular substrates. These substrates are so highly substituted with fluorescein moieties that...uninformative for breast cancer, but does correlate with bone mineral density, parathyroid hormone and phosphorus . (Summary of Appendix II). Normal MEPE...calcium, phosphorus , vitamin D, as well as novel phosphatonin(s), and the bone and kidney organs. Candidate phosphaturic factors include MEPE; PHEX, a

Department of the Army Supply Bulletin, Army Medical Department Supply Information, SB8-75-S10

DTIC Science & Technology

2001-10-20

12 BTL FLUORESCEIN NA OPTH STRIPS 1 MG 300s 6505-01-159-1493 91C $125.24 1 PG GUAIFENESIN / DEXTRAMETHORAPHAN COUGH SYRUP 4...oz 6505-01-318-1565 91B $1.00 12 BTL GUAIFENESIN EXTENDED RELEASE TABLETS 600MG 100s 6505-01-238-9443 91C $3.89 3 BTL HEMORRHOIDAL

Ocular fundus manifestation of two patients following long-term chloroquine therapy: a case report

PubMed Central

2010-01-01

This report describes the typical manifestations of chloroquine retinopathy with some advanced new technology. A series of examinations were performed on the patients, including the fundus fluorescein angiography, optical coherence tomography, GDxVCC Nerve Fiber Analyzer, full-field electroretinography, multifocal electroretinography and visual field examinations, to provide a better understanding of chloroquine retinopathy. PMID:20346186

Choroidal nevus with subretinal pigment epithelial neovascular membrane and a positive P-32 test

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

Snip, R.C.; Green, W.R.; Jaegers, K.R.

A 62-year-old white female was found to have a small, flat pigmented choroidal tumor. After fluorescein angiography and a positive P-32 test, the eye was enucleated for presumed malignant melanoma. Histologically, the tumor proved to be a choroidal nevus with a break in Bruch's membrane and a subretinal-pigment-epithelial neovascular membrane.

Restoration of the Retinal Structure and Function after Injury

DTIC Science & Technology

2013-10-01

cells in the retina, pan-retinal photocoagulation for proliferative diabetic retinopathy involves the purposeful destruction of a significant fraction of...also causes scotomata and scarring (Morgan and Schatz, 1989; Schatz et al., 1991; Early Treatment Diabetic Retinopathy Study Research Group, 1995... Retinopathy Study Research Group 1995 Focal photocoagulation treatment of diabetic macular edema–relationship of treatment effect to fluorescein

Targeted Deposition of Antibodies on a Multiplex CMOS Microarray and Optimization of a Sensitive Immunoassay Using Electrochemical Detection

DTIC Science & Technology

2010-03-19

multiplex array. The array had capture Abs against ricin, Bacillus globigii spores, M13 phage , a1 acid glycoprotein, and fluorescein. Initially, antigen (Ag...comply with a collection of information if it does not display a currently valid OMB control number. 1. REPORT DATE JAN 2010 2. REPORT TYPE 3

Bacterial Aerosols from a Field Source during Multiple-Sprinkler Irrigation: Deer Creek Lake State Park, Ohio

DTIC Science & Technology

1979-09-01

was funded under Civil Works Project CWIS 31280, Evaluation of Existing Facilities for Waste- water Land Treatment. Robert Emerson of the Atmospheric...20 Estimation of Coliphage .................................. 23 Fluorescein Dye Runs ..................................... 23 Chemicai and...California Water Pollletion Control Board 1957) in which data were obtained on the travel of coliform bacteria from spray operations using

Traumatic central retinal artery occlusion with sickle cell trait.

PubMed

Sorr, E M; Goldberg, R E

1975-10-01

An 8-year-old black boy with sickle cell trait struck his left brow and globe on the edge of a table and had immediate blurring of vision. Six days later visual acuity was light projection, and traumatic iritis with secondary glaucoma and perimacular edema were present. Fluorescein angiography indicated obstructed peripheral and perimacular arterioles and dye leakage from the disk.

Identification and Characterization of a Gene Cluster Mediating Enteroaggregative Escherichia Coli Aggregative Adherence Fimbria I Biogenesis

DTIC Science & Technology

1994-08-01

microscopy were performed by standard methods with fluoresceinated forward and reverse pUC primers (ABI) with a JOEL JEM 1200 EX 11 transr-ission...MyfB 11 enzrocolinca 41 220 uous plasmid regions required for AAF/I expre:sion and AA. PapD E. cobi 34 200 In this paper. we present a detailed analysis

Estrogen Receptor Driven Inhibitor Synthesis

DTIC Science & Technology

2006-09-01

labeling thiols in cellular pro- tein extracts [6,7]. But because these probes are based on nonspecific electrophiles , they are inherently less...selective electrophiles such as maleimide, alkyl halides, and iodoacetamide [5,25,26]. Conclusions DSSA probes composed of fluorescein tethered to rho...Haugland, F. Mao, Fluorinated xanthene derivatives, US patent. 6 (162), (2000) 931. [21] T. Nguyen, M.B. Francis, Practical route to functionalized rhoda

Hoechst tagging: a modular strategy to design synthetic fluorescent probes for live-cell nucleus imaging.

PubMed

Nakamura, Akinobu; Takigawa, Kazumasa; Kurishita, Yasutaka; Kuwata, Keiko; Ishida, Manabu; Shimoda, Yasushi; Hamachi, Itaru; Tsukiji, Shinya

2014-06-11

We report a general strategy to create small-molecule fluorescent probes for the nucleus in living cells. Our strategy is based on the attachment of the DNA-binding Hoechst compound to a fluorophore of interest. Using this approach, simple fluorescein, BODIPY, and rhodamine dyes were readily converted to novel turn-on fluorescent nucleus-imaging probes.

Glucocorticoid Regulation of Rat Renal Sodium Potassium Adenosine Triphosphatase

DTIC Science & Technology

1990-03-29

sequences; restriction enzymes fluorescein isothiocyanate glomerular filtration rate Horseradish Peroxidase immunoglobulin G kllodalton Magnesium...studies were conducted, in this project , to determine whether the observed changes in NaK-ATPase activity occurred after, and possibly as the result of...excitable tissue required for nerve impulse transmission and 6 muscle contraction (Skou, 1957), the functioning of hepatic amino acid and bile acid

Electrokinetically controlled fluid injection into unicellular microalgae.

PubMed

Zhou, Xuewen; Zhang, Xixi; Boualavong, Jonathan; Durney, Andrew R; Wang, Tonghui; Kirschner, Scott; Wentz, Michaela; Mukaibo, Hitomi

2017-10-01

Electrokinetically controlled microinjection is reported as an effective transport mechanism for microinjection into the wild-type strain of the widely studied model microalga Chlamydomonas reinhardtii. A microinjection system using glass capillary pipettes was developed to capture and impale the motile cells. To apply an electric field and induce electrokinetic flow (e.g., electrophoresis and electroosmosis), an electrode was inserted directly into the solution inside the impaling injection pipette and another electrode was inserted into the external cell media. The viability of the impaled cells was confirmed for more than an hour under 0.01 V using the fluorescein diacetate/propidium iodide dual fluorescent dye based assay. The viability was also found to increase almost logarithmically with decreasing voltage and to depend strongly on the solution within the injection pipette. Successful electrokinetic microinjection into cells was confirmed by both an increase in cell volume under an applied voltage and electric field dependent delivery of fluorescent fluorescein molecules into an impaled cell. Our study offers novel opportunities for quantitative delivery of biomolecules into microalgae and advancing the research and development of these organisms as biosynthetic factories. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Real-time monitoring of endogenous cysteine levels in living cells using a CD-based ratiometric fluorescent nanoprobe.

PubMed

Wang, Hong; Zhang, Peisheng; Tian, Yong; Zhang, Yuan; Yang, Heping; Chen, Shu; Zeng, Rongjin; Long, Yunfei; Chen, Jian

2018-04-30

A simple and readily available fluorescent probe is needed for the real-time monitoring of endogenous cysteine (Cys) levels in living cells, as such a probe could be used to study the role of Cys in related diseases. Herein, we report the first fluorescent probe based on carbon dots (CDs-FITA) for the selective and ratiometric imaging of endogenous Cys in live cells. In this ratiometric fluorescent probe, a fluorescein derivative (FITA) that recognizes Cys is covalently linked to the surfaces of carbon dots (CDs); employing CDs greatly improves the water solubility of the probe. Acrylate on FITA is selectively cleaved by Cys in aqueous solution under mild conditions, leading to a dramatic increase in the fluorescence from fluorescein. The probe therefore allows the highly selective ratiometric fluorescent detection of Cys even in the presence of various interferents. The as-prepared CDs-FITA showed excellent performance when applied to detect Cys in blood serum. In addition, due to its negligible cytotoxicity, the CDs-FITA can also be utilized for the real-time monitoring of endogenous cysteine (Cys) levels in living cells. Graphical abstract Illustration of the CD-based probe for Cys imaging in living cells.

Preliminary effects of oral uridine on the ocular surface in dry eye patients.

PubMed

Chang, Ki Cheol; Oh, Joo Youn; In, Youn Seok; Kim, Mee Kum; Shin, Ki Cheul; Wee, Won Ryang; Lee, Jin Hak; Park, Myung Gyu

2009-08-01

We designed a randomized, double blinded, 3-months controlled prospective clinical study to investigate effects of oral uridine on the ocular surface in dry eye patients. Twenty-seven patients who diagnosed as dry eye with lower than 5 mm of wetting in the Schirmer strip, with corneal epithelial erosion and who completely followed-up till 3 months were enrolled. Corneal-conjunctival fluorescein staining, non-anesthetic Schirmer test, impression cytology, and Ocular Surface Disease Index (OSDI) were evaluated in the experimental and placebo groups at the baseline, 1 and 3 months after start of medication in a double blinded manner. Fluorescein stain score of the cornea was markedly decreased in oral uridine group compared to the placebo group at 3 months after medication (P=0.032, Mann-Whitney U test). The Schirmer wetting score for the oral uridine group was significantly increased (P=0.001, Wilcoxon signed rank test) at 3 months and its difference between two groups was statistically significant (P=0.030, Mann-Whitney U test). OSDI scores were significantly decreased at 1 and 3 months in treatment group. Oral uridine is effective in treatment of dry eyes.

Integrated on-chip derivatization and electrophoresis for the rapid analysis of biogenic amines.

PubMed

Beard, Nigel P; Edel, Joshua B; deMello, Andrew J

2004-07-01

We demonstrate the monolithic integration of a chemical reactor with a capillary electrophoresis device for the rapid and sensitive analysis of biogenic amines. Fluorescein isothiocyanate (FITC) is widely employed for the analysis of amino-group containing analytes. However, the slow reaction kinetics hinders the use of this dye for on-chip labeling applications. Other alternatives are available such as o-phthaldehyde (OPA), however, the inferior photophysical properties and the UV lambdamax present difficulties when using common excitation sources leading to a disparity in sensitivity. Consequently, we present for the first time the use of dichlorotriazine fluorescein (DTAF) as a superior in situ derivatizing agent for biogenic amines in microfluidic devices. The developed microdevice employs both hydrodynamic and electroosmotic flow, facilitating the creation of a polymeric microchip to perform both precolumn derivatization and electrophoretic analysis. The favorable photophysical properties of the DTAF and its fast reaction kinetics provide detection limits down to 1 nM and total analysis times (including on-chip mixing and reaction) of <60 s. The detection limits are two orders of magnitude lower than current limits obtained with both FITC and OPA. The optimized microdevice is also employed to probe biogenic amines in real samples.

Highly sensitive microfluidic competitive enzyme immunoassay based on chemiluminescence resonance energy transfer for the detection of neuron-specific enolase.

PubMed

Yang, Tingzhen; Vdovenko, Marina; Jin, Xue; Sakharov, Ivan Yu; Zhao, Shulin

2014-07-01

A microfluidic competitive enzyme immunoassay based on chemiluminescence resonance energy transfer (CRET) was developed for highly sensitive detection of neuron-specific enolase (NSE). The CRET system consisted of horseradish peroxidase (HRP)/luminol as a light donor and fluorescein isothiocyanate as an acceptor. When fluorescein isothiocyanate-labeled antibody binds with HRP-labeled antigen to form immunocomplex, the donor and acceptor are brought close each other and CRET occurs in the immunocomplex. In the MCE, the immunocomplex and excess HRP-NSE were separated, and the chemiluminescense intensity of immunocomplex was used to estimate NSE concentration. The calibration curve showed a linearity in the range of NSE concentrations from 9.0 to 950 pM with a correlation coefficient of 0.9964. Based on a S/N of 3, the detection limit for NSE determination was estimated to be 4.5 pM, which is two-order magnitude lower than that of without CRET detection. This assay was applied for NSE quantification in human serum. The obtained results demonstrated that the proposed immunoassay may serve as an alternative tool for clinical analysis of NSE. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Pinhole shifting lifetime imaging microscopy

PubMed Central

Ramshesh, Venkat K.; Lemasters, John J.

2009-01-01

Lifetime imaging microscopy is a powerful tool to probe biological phenomena independent of luminescence intensity and fluorophore concentration. We describe time-resolved imaging of long-lifetime luminescence with an unmodified commercial laser scanning confocal/multiphoton microscope. The principle of the measurement is displacement of the detection pinhole to collect delayed luminescence from a position lagging the rasting laser beam. As proof of principle, luminescence from microspheres containing europium (Eu3+), a red emitting probe, was compared to that of short-lifetime green-fluorescing microspheres and/or fluorescein and rhodamine in solution. Using 720-nm two-photon excitation and a pinhole diameter of 1 Airy unit, the short-lifetime fluorescence of fluorescein, rhodamine and green microspheres disappeared much more rapidly than the long-lifetime phosphorescence of Eu3+ microspheres as the pinhole was repositioned in the lagging direction. In contrast, repositioning of the pinhole in the leading and orthogonal directions caused equal loss of short- and long-lifetime luminescence. From measurements at different lag pinhole positions, a lifetime of 270 μs was estimated for the Eu3+ microspheres, consistent with independent measurements. This simple adaptation is the basis for quantitative 3-D lifetime imaging microscopy. PMID:19123648

New Ways to Detect Pediatric Sickle Cell Retinopathy: A Comprehensive Review.

PubMed

Pahl, Daniel A; Green, Nancy S; Bhatia, Monica; Chen, Royce W S

2017-11-01

Sickle retinopathy reflects disease-related vascular injury of the eye, which can potentially result in visual loss from vitreous hemorrhage or retinal detachment. Here we review sickle retinopathy among children with sickle cell disease, describe the epidemiology, pediatric risk factors, pathophysiology, ocular findings, and treatment. Newer, more sensitive ophthalmological imaging modalities are available for retinal imaging, including ultra-widefield fluorescein angiography, spectral-domain optical coherence tomography, and optical coherence tomography angiography. Optical coherence tomography angiography provides a noninvasive view of retinal vascular layers that could previously not be imaged and can be quantified for comparative or prospective analyses. Ultra-widefield fluorescein angiography provides a more comprehensive view of the peripheral retina than traditional imaging techniques. Screening for retinopathy by standard fundoscopic imaging modalities detects a prevalence of approximately 10%. In contrast, these more sensitive methods allow for more sensitive examination that includes the retina perimeter where sickle retinopathy is often first detectable. Use of these new imaging modalities may detect a higher prevalence of early sickle pathology among children than has previously been reported. Earlier detection may help in better understanding the pathogenesis of sickle retinopathy and guide future screening and treatment paradigms.

Comparison between optical coherence tomography angiography and fluorescein angiography findings in retinal vasculitis.

PubMed

Abucham-Neto, Julio Zaki; Torricelli, André Augusto Miranda; Lui, Aline Cristina Fioravanti; Guimarães, Sarah Napoli; Nascimento, Heloisa; Regatieri, Caio Vinícius

2018-01-01

To describe optical coherence tomography angiography (OCT-A) findings in patients with retinal vasculitis and to compare them to current fluorescein angiography (FA) findings. This was an observational case series. Nineteen eyes in 10 patients with retinal vasculitis of various etiologies were imaged with FA (TRC-50DX, Topcon) and OCT-A (SD-OCT, Optovue). The images were reviewed and analyzed. The mean age was 36 years (range 24-67 years); there were three males and seven females. The primary vessels involved were veins (89%). Fourteen eyes (74%) had active inflammatory disease during the study period, with signs of vascular sheathing and perivascular leakage on FA. Interestingly, in this group, OCT-A was not able to detect clear signs of active inflammation around the affected vessels. Nevertheless, OCT-A was able to detect secondary lesions in fourteen eyes (74%), including some findings not clearly shown on FA. Most of these were within the macular area. OCT-A was particularly effective in cases of capillary dropout, increased foveal avascular zone, telangiectasias, shunts, and areas of neovascularization. FA remains an essential complementary exam for detection of retinal vasculitis. However, OCT-A extends FA findings and affords better assessment of secondary complications.

Ratiometric, visual, dual-signal fluorescent sensing and imaging of pH/copper ions in real samples based on carbon dots-fluorescein isothiocyanate composites.

PubMed

Zhu, Xinxin; Jin, Hui; Gao, Cuili; Gui, Rijun; Wang, Zonghua

2017-01-01

In this article, a facile aqueous synthesis of carbon dots (CDs) was developed by using natural kelp as a new carbon source. Through hydrothermal carbonization of kelp juice, fluorescent CDs were prepared and the CDs' surface was modified with polyethylenimine (PEI). The PEI-modified CDs were conjugated with fluorescein isothiocyanate (FITC) to fabricate CDs-FITC composites. To exploit broad applications, the CDs-FITC composites were developed as fluorescent sensing or imaging platforms of pH and Cu 2+ . Analytical performances of the composites-based fluorescence (FL) sensors were evaluated, including visual FL imaging of pH in glass bottle, ratiometric FL sensing of pH in yogurt samples, visual FL latent fingerprint and leaf imaging detection of [Cu 2+ ], dual-signal FL sensing of [Cu 2+ ] in yogurt and human serum samples. Experimental results from ratiometric, visual, dual-signal FL sensing and imaging applications confirmed the high feasibility, accuracy, stabilization and simplicity of CDs-FITC composites-based FL sensors for the detection of pH and Cu 2+ ions in real samples. Copyright © 2016 Elsevier B.V. All rights reserved.

Fullerol-fluorescein isothiocyanate-concanavalin agglutinin phosphorescent sensor for the detection of alpha-fetoprotein and forecast of human diseases

NASA Astrophysics Data System (ADS)

Liu, Jia-ming; Lin, Li-ping; Jiang, Shu-Lian; Cui, Ma Lin; Jiao, Li; Zhang, Xiao Yang; Zhang, Li-hong; Zheng, Zhi Yong; Lin, Xuan; Lin, Shao-qin

2013-11-01

Based on the reaction of the active -OH group in fullerol (F) with the dissociated -COOH group in fluorescein isothiocyanate (FITC) to form an F-FITC and the enhanced effect of N, N-dimethylaniline (DMA) on phosphorescence signal of F-FITC, a new phosphorescent labeling reagent (DMA-F-FITC) was developed. What's more, a phosphorescent sensor for the determination of alpha-fetoprotein variant (AFP-V) has been designed via the coupling technique of the high sensitivity for affinity adsorption-solid substrate-room temperature phosphorimetry (AA-SS-RTP) with the strong specificity reaction between DMA-F-FITC-Con A and AFP-V. The DMA-F-FITC increased the number of luminescent molecules in the biological target which improved the sensitivity of phosphorescent sensor. The proposed sensor was responsive, simple, selective and sensitive, and it has been applied to the determination of trace AFP-V in human serum and the forecast of human diseases using phosphorescence emission wavelength of F or FITC, with the results agreed well with those obtained by enzyme-linked immunoassay (ELISA). Meanwhile, the mechanisms for the labeling reaction and the sensing detection of AFP-V were discussed.

Pattern of glomerular diseases in Oman: a study based on light microscopy and immunofluorescence.

PubMed

Alwahaibi, Nasar Yousuf; Alhabsi, Taiseer Ahmed; Alrawahi, Samira Abdullah

2013-03-01

Light microscopy and immunofluorescence play an important part in the final diagnosis of renal biopsy. The aim of this study was to analyze the pattern of various glomerular diseases in Oman. A total of 424 renal biopsies were retrospectively analyzed at the Sultan Qaboos University Hospital between 1999 and 2010. Focal and segmental glomerulosclerosis (FSGS), minimal change disease (MCD), membranous glomerulopathy (MGN) and IgA nephropathy were the most common primary glomerular diseases encountered, accounting for 21.2%, 17%, 12.3% and 8.3%, respectively, of all cases. Lupus nephritis was the most common secondary glomerular disease and was the most prevalent among all biopsies, accounting for 30.4% of all biopsies. Amyloidosis was seen in only two cases. The presence of fluorescein isothiocyanatefibrin in all renal cases was low when compared with IgG, IgA, IgM, C3 and C1q markers. In conclusion, based on the findings of this study, lupus nephritis was the most common of all glomerular diseases and FSGS was the most common primary glomerular disease. The importance of fluorescein isothiocyanate-fibrin in the diagnosis of renal biopsy needs to be further investigated.

Fluorescence probe techniques to monitor protein adsorption-induced conformation changes on biodegradable polymers.

PubMed

Benesch, Johan; Hungerford, Graham; Suhling, Klaus; Tregidgo, Carolyn; Mano, João F; Reis, Rui L

2007-08-15

The study of protein adsorption and any associated conformational changes on interaction with biomaterials is of great importance in the area of implants and tissue constructs. This study aimed to evaluate some fluorescent techniques to probe protein conformation on a selection of biodegradable polymers currently under investigation for biomedical applications. Because of the fluorescence emanating from the polymers, the use of monitoring intrinsic protein fluorescence was precluded. A highly solvatochromic fluorescent dye, Nile red, and a well-known protein label, fluorescein isothiocyanate, were employed to study the adsorption of serum albumin to polycaprolactone and to some extent also to two starch-containing polymer blends (SPCL and SEVA-C). A variety of fluorescence techniques, steady state, time resolved, and imaging were employed. Nile red was found to leach from the protein, while fluorescein isothiocyanate proved useful in elucidating a conformational change in the protein and the observation of protein aggregates adsorbed to the polymer surface. These effects were seen by making use of the phenomenon of energy migration between the fluorescent tags to monitor interprobe distance and the use of fluorescence lifetime imaging to ascertain the surface packing of the protein on polymer.

Retinal reperfusion in diabetic retinopathy following treatment with anti-VEGF intravitreal injections.

PubMed

Levin, Ariana M; Rusu, Irene; Orlin, Anton; Gupta, Mrinali P; Coombs, Peter; D'Amico, Donald J; Kiss, Szilárd

2017-01-01

The aim of this study is to report peripheral reperfusion of ischemic areas of the retina on ultra-widefield fluorescein angiography (UWFA) following anti-vascular endothelial growth factor (VEGF) intravitreal injections in patients treated for diabetic retinopathy. This study is a retrospective review of 16 eyes of 15 patients with diabetic retinopathy, who received anti-VEGF intravitreal injections and underwent pre- and postinjection UWFA. The main outcome measured was the presence of reperfusion in postinjection UWFA images in areas of the retina that demonstrated nonperfusion in preinjection images. Images were analyzed for reperfusion qualitatively and quantitatively by two graders. Twelve of 16 eyes (75%) or 11 of 15 patients (73.3%) demonstrated reperfusion following anti-VEGF injection. On UWFA, reperfusion was detected both within the field of 7-standard field (7SF) fluorescein angiography and in the periphery outside the 7SF. Four of 16 eyes or 4 of 15 patients did not demonstrate reperfusion, one of which had extensive scarring from prior panretinal photocoagulation. In patients with diabetic retinopathy, treatment with anti-VEGF agents can be associated with reperfusion of areas of nonperfusion, as demonstrated by UWFA.

Single-primer fluorescent sequencing

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

Ruth, J.L.; Morgan, C.A.; Middendorf, L.R.

Modified linker arm oligonucleotides complementary to standard M13 priming sites were synthesized, labelled with either one, two, or three fluoresceins, and purified by reverse-phase HPLC. When used as primers in standard dideoxy M13 sequencing with /sup 32/P-dNTPs, normal autoradiographic patterns were obtained. To eliminate the radioactivity, direct on-line fluorescence detection was achieved by the use of a scanning 10 mW Argon laser emitting 488 nm light. Fluorescent bands were detected directly in standard 0.2 or 0.35 mm thick polyacrylamide gels at a distance of 24 cm from the loading wells by a photomultiplier tube filtered at 520 nm. Horizontal andmore » temporal location of each band was displayed by computer as a band in real time, providing visual appearance similar to normal 4-lane autoradiograms. Using a single primer labelled with two fluoresceins, sequences of between 500 and 600 bases have been read in a single loading with better than 98% accuracy; up to 400 bases can be read reproducibly with no errors. More than 50 sequences have been determined by this method. This approach requires only 1-2 ug of cloned template, and produces continuous sequence data at about one band per minute.« less

Synthesis of molecularly imprinted dye-silica nanocomposites with high selectivity and sensitivity: Fluorescent imprinted sensor for rapid and efficient detection of τ-fluvalinate in vodka.

PubMed

Wang, Yunyun; Wang, Jixiang; Cheng, Rujia; Sun, Lin; Dai, Xiaohui; Yan, Yongsheng

2018-04-01

An imprinted fluorescent sensor was fabricated based on SiO 2 nanoparticles encapsulated with a molecularly imprinted polymer containing allyl fluorescein. High fluorine cypermethirin as template molecules, methyl methacrylate as functional monomer, and allyl fluorescein as optical materials synthesized a core-shell fluorescent molecular imprinted sensor, which showed a high and rapid sensitivity and selectivity for the detection of τ-fluvalinate. The sensor presented appreciable sensitivity with a limit of 13.251 nM, rapid detection that reached to equilibrium within 3 min, great linear relationship in the relevant concentration range from 0 to 150 nM, and excellent selectivity over structural analogues. In addition, the fluorescent sensor demonstrated desirable regeneration ability (eight cycling operations). The molecularly imprinted polymers ensured specificity, while the fluorescent dyes provided the stabile sensitivity. Finally, an effective application of the sensor was implemented by the detection of τ-fluvalinate in real samples from vodka. The molecularly imprinted fluorescent sensor showed a promising potential in environmental monitoring and food safety. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Quantification of plant cell coupling with live-cell microscopy.

PubMed

Liesche, Johannes; Schulz, Alexander

2015-01-01

Movement of nutrients and signaling compounds from cell to cell is an essential process for plant growth and development. To understand processes such as carbon allocation, cell communication, and reaction to pathogen attack it is important to know a specific molecule's capacity to pass a specific cell wall interface. Transport through plasmodesmata, the cell wall channels that directly connect plant cells, is regulated not only by a fixed size exclusion limit, but also by physiological and pathological adaptation. The noninvasive approach described here offers the possibility of precisely determining the plasmodesmata-mediated cell wall permeability for small molecules in living cells.The method is based on photoactivation of the fluorescent tracer caged fluorescein. Non-fluorescent caged fluorescein is applied to a target tissue, where it is taken up passively into all cells. Imaged by confocal microscopy, loaded tracer is activated by UV illumination in a target cell and its spread to neighboring cells monitored. When combined with high-speed acquisition by resonant scanning or spinning disc confocal microscopy, the high signal-to-noise ratio of photoactivation allows collection of three-dimensional (3D) time series. These contain all necessary functional and anatomical data to measure cell coupling in complex tissues noninvasively.

Relationship between the structures of flavonoids and oxygen radical absorbance capacity values: a quantum chemical analysis.

PubMed

Zhang, Di; Liu, Yixiang; Chu, Le; Wei, Ying; Wang, Dan; Cai, Shengbao; Zhou, Feng; Ji, Baoping

2013-02-28

Various radical-scavenging activities (RSA) assessment assays are based on discrete mechanisms and on using different radical sources. Few studies have analyzed the structural significance of flavonoids in their peroxyl radical activities in the oxygen radical absorbance capacity (ORAC) assay. In this study, the RSA of 13 flavonoids in two ORAC assays with different probes (fluorescein and pyrogallol red) were investigated. Neither O-H bond dissociation enthalpy nor ionization potential values of flavonoids correlated with ORAC values. The proton affinity (PA) and electron transfer enthalpy (ETE) values, which were obtained via the sequential proton-loss electron-transfer mechanism, were significantly associated with the ORAC(pyrogallol Red) and ORAC(fluorescein) assays, respectively. Thus, PA represented the kinetic aspect of RSA, whereas ETE reflected the RSA extent. The PA values and the most acidic sites of flavonoids were affected by intramolecular electronic interactions, H-bonding, 3-hydroxyl group in the C ring, and conjugation systems. The stability of the deprotonated flavonoid determined the ETE value. Apart from the PA and ETE values in the first oxidation step of flavonoids, the PA and ETE values in the second oxidation step also affected the ORAC values of flavonoids.

A highly sensitive solid substrate room temperature phosphorimetry for carbaryl detection based on its activating effect on NaIO4 oxidizing fluorescein.

PubMed

Liu, Jiaming; Huang, Qitong; Liu, Zhen-bo; Lin, Xiaofeng; Zhang, Li-Hong; Lin, Chang-Qing; Zheng, Zhi-Yong

2014-11-01

Fluorescein (HFin) could emit strong and stable room temperature phosphorescence (RTP) signal on polyamide membrane (PAM) using Pb(2+) as the ion perturber. Carbaryl could activate effect on NaIO4 oxidating HFin, which caused the RTP signal of the system to quench sharply. The phosphorescence intensity (ΔI p) of activating system higher 3.3 times (119.4/36.0) than that of non-activating system, and is directly proportional to the content of carbaryl. Thus, an activating solid substrate room temperature phosphorimetry (SSRTP) for carbaryl detection has been established. This sensitive (the limit of quantification (LOQ) was 2.0 × 10(-13) g mL(-1)), selective, simple and rapid method has been applied to determine trace carbaryl in water samples with the results consisting with those obtained by fluorimetry, showing its high accuracy. The apparent activation energy (E) and rate constant (k) of this activating reaction were 20.77 kJ mol(-1) and 1.85 × 10(-4) s(-1), respectively. Meanwhile, the mechanism of activating SSRTP for carbaryl detection was also discussed using infrared spectra (IR).

Coats-like retinitis pigmentosa: Reports of three cases

PubMed Central

Kan, Emrah; Yilmaz, Turgut; Aydemir, Orhan; Güler, Mete; Kurt, Jülide

2007-01-01

Purpose: Describing the ophthalmic findings of an exudative vasculopathy called as Coats-like retinitis pigmentosa on three patients. The etiology of the Coats-like retinitis pigmentosa is obscure. The principal theories have been discussed in this article. Methods: Three observational case series have been discussed. Complete ophthalmic examinations and color fundus photos, visual field, and fluorescein angiography have been performed. Results: We have identified 3 patients who have some typical clinical features of Coats-like retinitis pigmentosa; peripheral serous retinal detachment, telangiectasia, prominent lipid deposition, pigmentary changes in peripheral retina, and loss of vision. None of the three patients had positive family history. All of the patients have had symptoms of nyctalopia, decreased central vision, and two of them have had constriction of visual field. All of the patients have had cataracts and two of them underwent cataract surgery. Fundus examination and fluorescein angiography of patients revealed typical retinitis pigmentosa with Coats-type changes in bilateral inferiotemporal quadrants. Conclusion: A better understanding of clinical features and genetic etiology of Coats-type retinitis pigmentosa will aid diagnosis and development of new therapies. If sufficient conditions arise, genetic factors that influence the expression of CRB1 mutations in Coats-like retinitis pigmentosa should be detected. PMID:19668510

Coats-like retinitis pigmentosa: Reports of three cases.

PubMed

Kan, Emrah; Yilmaz, Turgut; Aydemir, Orhan; Güler, Mete; Kurt, Jülide

2007-06-01

Describing the ophthalmic findings of an exudative vasculopathy called as Coats-like retinitis pigmentosa on three patients. The etiology of the Coats-like retinitis pigmentosa is obscure. The principal theories have been discussed in this article. Three observational case series have been discussed. Complete ophthalmic examinations and color fundus photos, visual field, and fluorescein angiography have been performed. We have identified 3 patients who have some typical clinical features of Coats-like retinitis pigmentosa; peripheral serous retinal detachment, telangiectasia, prominent lipid deposition, pigmentary changes in peripheral retina, and loss of vision. None of the three patients had positive family history. All of the patients have had symptoms of nyctalopia, decreased central vision, and two of them have had constriction of visual field. All of the patients have had cataracts and two of them underwent cataract surgery. Fundus examination and fluorescein angiography of patients revealed typical retinitis pigmentosa with Coats-type changes in bilateral inferiotemporal quadrants. A better understanding of clinical features and genetic etiology of Coats-type retinitis pigmentosa will aid diagnosis and development of new therapies. If sufficient conditions arise, genetic factors that influence the expression of CRB1 mutations in Coats-like retinitis pigmentosa should be detected.

Fluorescence turn-on detection of iodide, iodate and total iodine using fluorescein-5-isothiocyanate-modified gold nanoparticles.

PubMed

Chen, Yi-Ming; Cheng, Tian-Lu; Tseng, Wei-Lung

2009-10-01

Selective turn-on fluorescence detection of I(-) was accomplished using fluorescein isothiocyanate-decorated gold nanoparticles (FITC-AuNPs). FITC molecules, which fluoresce strongly in an alkaline solution, were severely quenched when they were attached to the surface of AuNPs through their isothiocyanate group. Upon the addition of I(-), FITC molecules were detached because of I(-) adsorption on the surface of AuNPs. As a result, released FITC molecules were restored to their original fluorescence intensity. Because I(-) has a higher binding affinity to the surface of Au than do Br(-), Cl(-), or F(-), the FITC-AuNPs obviously have a higher selectivity toward I(-) than toward these other anions. Meanwhile, after IO(3)(-) was reduced to I(-) with ascorbic acid, the detection of IO(3)(-) was successfully achieved using the FITC-AuNPs. Under an optimum pH and AuNP concentration, the lowest detectable concentrations of I(-) and IO(3)(-) using this probe were 10.0 and 50.0 nM, respectively. The FITC-AuNPs provide a number of advantages, including easy preparation, selectivity, sensitivity, and low cost. This unique probe was applied to an analysis of the total iodine in edible salt and seawater.

Fluorescein diacetate for determination of cell viability in 3D fibroblast-collagen-GAG constructs.

PubMed

Powell, Heather M; Armour, Alexis D; Boyce, Steven T

2011-01-01

Quantification of cell viability and distribution within engineered tissues currently relies on representative histology, phenotypic assays, and destructive assays of viability. To evaluate uniformity of cell density throughout 3D collagen scaffolds prior to in vivo use, a nondestructive, field assessment of cell viability is advantageous. Here, we describe a field measure of cell viability in lyophilized collagen-glycosaminoglycan (C-GAG) scaffolds in vitro using fluorescein diacetate (FdA). Fibroblast-C-GAG constructs are stained 1 day after cellular inoculation using 0.04 mg/ml FdA followed by exposure to 366 nm UV light. Construct fluorescence quantified using Metamorph image analysis is correlated with inoculation density, MTT values, and histology of corresponding biopsies. Construct fluorescence correlates significantly with inoculation density (p  <  0.001) and MTT values (p  <  0.001) of biopsies collected immediately after FdA staining. No toxicity is detected in the constructs, as measured by MTT assay before and after the FdA assay at different time points; normal in vitro histology is demonstrated for the FdA-exposed constructs. In conclusion, measurement of intracellular fluorescence with FdA allows for the early, comprehensive measurement of cellular distributions and viability in engineered tissue.

Magneto-actuated immunoassay for the detection of Mycobacterium fortuitum in hemodialysis water.

PubMed

Brugnera, Michelle Fernanda; Bundalian, Reynaldo; Laube, Tamara; Julián, Esther; Luquin, Marina; Zanoni, Maria Valnice Boldrin; Pividori, Maria Isabel

2016-06-01

This paper addresses a sensitive method for the detection of mycobacteria in hemodialysis water samples based on a magneto-actuated immunoassay with optical readout. In this approach, micro (2.8μm) sized magnetic particles were modified with an antibody against the lipoarabinomannan (LAM) located in the mycobacterial cell wall. The system relies on the immunocapturing of the mycobacteria with the tailored antiLAM magnetic particles to pre-concentrate the bacteria from the hemodialysis samples throughout an immunological reaction. The performance of the immunomagnetic separation on the magnetic carrier was evaluated using confocal microscopy to study the binding pattern, as well as a magneto-actuated immunoassay with optical readout for the rapid detection of the bacteria in spiked hemodialysis samples. In this approach, the antiLAM polyclonal antibody was labeled with fluorescein isothiocyanate. The optical readout was achieved by the incubation with a secondary anti-fluorescein antibody labeled with peroxidase as optical reporter. The magneto-actuated immunoassay was able to detect mycobacteria contamination in hemodialysis water at a limit of detection of 13CFUmL(-1) in a total assay time of 3h without any previous culturing pre-enrichment step. Copyright © 2016 Elsevier B.V. All rights reserved.

DNA methylation detection based on difference of base content

NASA Astrophysics Data System (ADS)

Sato, Shinobu; Ohtsuka, Keiichi; Honda, Satoshi; Sato, Yusuke; Takenaka, Shigeori

2016-04-01

Methylation frequently occurs in cytosines of CpG sites to regulate gene expression. The identification of aberrant methylation of certain genes is important for cancer marker analysis. The aim of this study was to determine the methylation frequency in DNA samples of unknown length and/or concentration. Unmethylated cytosine is known to be converted to thymine following bisulfite treatment and subsequent PCR. For this reason, the AT content in DNA increases with an increasing number of methylation sites. In this study, the fluorescein-carrying bis-acridinyl peptide (FKA) molecule was used for the detection of methylation frequency. FKA contains fluorescein and two acridine moieties, which together allow for the determination of the AT content of double-stranded DNA fragments. Methylated and unmethylated human genomes were subjected to bisulfide treatment and subsequent PCR using primers specific for the CFTR, CDH4, DBC1, and NPY genes. The AT content in the resulting PCR products was estimated by FKA, and AT content estimations were found to be in good agreement with those determined by DNA sequencing. This newly developed method may be useful for determining methylation frequencies of many PCR products by measuring the fluorescence in samples excited at two different wavelengths.

The analytical measurement of fluorescein, quinine and trace metal concentrations in solution using single bubble sonoluminescence

NASA Astrophysics Data System (ADS)

Wallace, P.; McCallum, K.; Barnard, C. L. R.; Clement, C.; Marshall, J.; Carroll, J.

2007-03-01

A single bubble was generated and levitated in a high-intensity sound field within a spherical flask excited in its fundamental mode. Under optimum experimental conditions the bubble was observed to emit light in the form of short flashes. This phenomenon is known as single bubble sonoluminescence (SBSL). Using this process, the emitted light from the bubble was monitored when solutions containing fluorescein, quinine and sodium, potassium and copper salts were placed in the cell. The results obtained indicated that reproducible signals related directly to the concentration of the species present in solution could be achieved using single bubble sonoluminescence. The results for the molecular species were compared with those obtained by fluorescence spectroscopy and, in the case of quinine, parallel determinations of concentration in a test solution were performed with consistent results. SBSL signals were also observed to exhibit a linear correlation with the concentration of several trace metal salts introduced to the solution in the measurement cell. However, it was not possible to demonstrate that the SBSL signals were derived from stimulated atomic emission or fluorescence, and it was concluded that the effect may result from an indirect effect involving the bubble excitation mechanism.

Novel flow cytometric analysis of the progress and route of internalization of a monoclonal anti-carcinoembryonic antigen (CEA) antibody.

PubMed

Ford, C H; Tsaltas, G C; Osborne, P A; Addetia, K

1996-03-01

A flow cytometric method of studying the internalization of a monoclonal antibody (Mab) directed against carcinoembryonic antigen (CEA) has been compared with Western blotting, using three human colonic cancer cell lines which express varying amounts of the target antigen. Cell samples incubated for increasing time intervals with fluoresceinated or unlabelled Mab were analyzed using flow cytometry or polyacrylamide gel electrophoresis and Western blotting. SDS/PAGE analysis of cytosolic and membrane components of solubilized cells from the cell lines provided evidence of non-degraded internalized anti-CEA Mab throughout seven half hour intervals, starting at 5 min. Internalized anti-CEA was detected in the case of high CEA expressing cell lines (LS174T, SKCO1). Very similar results were obtained with an anti-fluorescein flow cytometric assay. Given that these two methods consistently provided comparable results, use of flow cytometry for the detection of internalized antibody is suggested as a rapid alternative to most currently used methods for assessing antibody internalization. The question of the endocytic route followed by CEA-anti-CEA complexes was addressed by using hypertonic medium to block clathrin mediated endocytosis.

Colorful Polyelectrolytes: An Atom Transfer Radical Polymerization Route to Fluorescent Polystyrene Sulfonate.

PubMed

Huberty, Wayne; Tong, Xiaowei; Balamurugan, Sreelatha; Deville, Kyle; Russo, Paul S; Zhang, Donghui

2016-03-01

A labeled green fluorescent polystyrene sulfonate (LNaPSS) has been synthesized using atom transfer radical polymerization of a styrene sulfonate monomer with a fluorescent co-monomer, fluorescein thiocyanate-vinyl aniline. As a result this 100 % sulfonated polymer contains no hydrophobic patches along the chain backbone besides the fluorescent marker itself. The concentration of the fluorescent monomer was kept low to maintain the characteristic properties of the anionic polyelectrolyte, LNaPSS. ATRP conditions facilitated the production of polymers spanning a range of molecular weights from 35,000 to 175,000 in gram-scale batches with polydispersity indices of 1.01-1.24. Molecular weight increased with the monomer to initiator ratio. Gel permeation chromatography results show a unimodal distribution, and the polymer structure was also confirmed by (1)H NMR and FT-IR spectroscopy. Fluorescence spectroscopy confirmed covalent bonding of fluorescein isothiocyanate to the polymer, indicating that the polymer is suitable as a probe in fluorescence microscopy. To demonstrate this ability, the polymer was used to locate structural features in salt crystals formed during drying, as in the evaporation of sea mist. A second application to probe diffusion studies is also demonstrated.

A new tool for the rapid remote detection of leaks from subsea pipelines during remotely operated vehicle inspections

NASA Astrophysics Data System (ADS)

McStay, D.; McIlroy, J.; Forte, A.; Lunney, F.; Greenway, T.; Thabeth, K.; Dean, G.

2005-06-01

A new 2000 m depth rated subsea sensor that can effectively, rapidly and remotely detect leaks of fluorescein dye, leak detection chemicals and hydraulic fluids from underwater structures is reported. The system utilizes ultra-bright LED technology to project a structured beam of light, at a wavelength suitable to excite the fluorescence of the target material, into the water column. The resultant fluorescence is collected and digital signal processing used to extract the intensity. The system is capable of detecting ppm concentrations of fluorescein at a range of 2.5 m in water in real time. The ability to stand-off from subsea structures, while rapidly detecting the chemicals makes the system highly suited to subsea leak inspections with remotely operated vehicles or autonomous underwater vehicles, as it allows the vehicles to be flown quickly and safely over the structure to be inspected. This increases both the speed and effectiveness of the inspection. The remote detection capability is also highly effective for probing complex underwater structures. The system has been successfully used in real subsea survey applications and has been found to be effective, user friendly and to dramatically reduce inspection times and hence costs.

Analysis of ground-water flow in the Madison aquifer using fluorescent dyes injected in Spring Creek and Rapid Creek near Rapid City, South Dakota, 2003-04

USGS Publications Warehouse

Putnam, Larry D.; Long, Andrew J.

2007-01-01

The Madison aquifer, which contains fractures and solution openings in the Madison Limestone, is used extensively for water supplies for the city of Rapid City and other suburban communities in the Rapid City, S. Dak., area. The 48 square-mile study area includes the west-central and southwest parts of Rapid City and the outcrops of the Madison Limestone extending from south of Spring Creek to north of Rapid Creek. Recharge to the Madison Limestone occurs when streams lose flow as they cross the outcrop. The maximum net loss rate for Spring and Rapid Creek loss zones are 21 and 10 cubic feet per second (ft3/s), respectively. During 2003 and 2004, fluorescent dyes were injected in the Spring and Rapid Creek loss zones to estimate approximate locations of preferential flow paths in the Madison aquifer and to measure the response and transit times at wells and springs. Four injections of about 2 kilograms of fluorescein dye were made in the Spring Creek loss zone during 2003 (sites S1, S2, and S3) and 2004 (site S4). Injection at site S1 was made in streamflow just upstream from the loss zone over a 12-hour period when streamflow was about equal to the maximum loss rate. Injections at sites S2, S3, and S4 were made in specific swallow holes located in the Spring Creek loss zone. Injection at site R1 in 2004 of 3.5 kilograms of Rhodamine WT dye was made in streamflow just upstream from the Rapid Creek loss zone over about a 28-hour period. Selected combinations of 27 wells, 6 springs, and 3 stream sites were monitored with discrete samples following the injections. For injections at sites S1-S3, when Spring Creek streamflow was greater than or equal to 20 ft3/s, fluorescein was detected in samples from five wells that were located as much as about 2 miles from the loss zone. Time to first arrival (injection at site S1) ranged from less than 1 to less than 10 days. The maximum fluorescein concentration (injection at site S1) of 120 micrograms per liter (ug/L) at well CO, which is located adjacent to the loss zone, was similar to the concentration in the stream. Fluorescein arrived at well NON (injection at site S1), which is located about 2 miles northeast of the loss zone, within about 1.6 days, and the maximum concentration was 44 ug/L. For injection at site S4, when streamflow was about 12 ft3/s, fluorescein was detected in samples from six wells and time to first arrival ranged from 0.2 to 16 days. Following injection at site S4 in 2004, the length of time that dye remained in the capture zone of well NON, which is located approximately 2 miles from the loss zone, was almost an order of magnitude greater than in 2003. For injection at site R1, Rhodamine WT was detected at well DRU and spring TI-SP with time to first arrival of about 0.5 and 1.1 days and maximum concentrations of 6.2 and 0.91 ug/L, respectively. Well DRU and spring TI-SP are located near the center of the Rapid Creek loss zone where the creek has a large meander. Measurable concentrations were observed for spring TI-SP as many as 109 days after the dye injection. The direction of a conduit flow path in the Spring Creek area was to the northeast with ground-water velocities that ranged from 770 to 6,500 feet per day. In the Rapid Creek loss zone, a conduit flow path east of the loss zone was not evident from the dye injection.

Correlation between Octopus perimetry and fluorescein angiography after strontium-90 plaque brachytherapy for subfoveal exudative age related macular degeneration.

PubMed

Jaakkola, A; Vesti, E; Immonen, I

1998-07-01

To evaluate the correlation between the central visual field and changes in fluorescein angiography and fundus photography in patients treated with strontium plaque radiotherapy for subfoveal exudative age related macular degeneration (AMD). Octopus program 34 automated static perimetry, fluorescein angiography, and colour fundus photography were performed on 19 patients at baseline and at 12 months after strontium-90 plaque therapy. A schematic picture outlining the areas of hyperfluorescent neovascular membranes and subretinal blood was drawn of a projected 30 degrees fundus fluorescein angiogram. This drawing was superimposed on the size adjusted Octopus visual field. The changes in retinal sensitivity were calculated and related to angiographic changes. Three of the 19 patients had a reliability factor (RF) > 15% and were excluded from further analysis. In the remaining 16 patients the mean defect (MD) and loss variance (LV) values remained unchanged in patients showing regression of the choroidal neovascular membrane (CNVM) to irradiation at 12 months. MD was 7.7 (SD 1.7) at baseline and 7.6 (1.9) at 12 months (p = 0.86), and LV was 32.6 (13.9) at baseline and 32.4 (15.7) at 12 months (p = 0.94). However, in patients with progression of the CNVM at 12 months, both the MD and LV increased significantly during the 12 month follow up (MD from 7.3 (2.9) to 13.1 (3.6) (p = 0.05) and LV from 31.0 (22.9) to 71.8 (24.1) (p = 0.017)). When comparing the mean retinal sensitivity in the area of the primary CNVM (including classic, occult, and haemorrhagic components), the results were analogous: in patients with a regression of the CNVM after irradiation the mean sensitivity remained almost unchanged. It was 10.3 (6.4) dB at baseline and 9.4 (7.3) dB at 12 months (p = 0.58). In five out of 11 patients (45%) with regression of the CNVM, the mean retinal sensitivity even improved by 2.0-5.0 dB in the area of the original lesion during follow up. Instead, in patients showing progression of the CNVM at 12 months, there was a significant loss in mean retinal sensitivity--from 9.9 (4.6) dB at baseline to 1.0 (1.1) dB at 12 months (p = 0.019). The mean retinal sensitivity in the area of the irradiated but clinically normal retina during follow up was not significantly altered (21.5 dB at baseline, 19.7 dB at 12 months (p = 0.10)). Regression of subfoveal choroidal membranes in AMD after focal strontium irradiation is connected with stabilisation or even improvement of retinal sensitivity in central visual field measured by automated perimetry. Strontium plaque irradiation does not change the sensitivity in clinically normal paramacular retina during a 12 month follow up.

Effects of Microwave Radiation on Humans. Monkeys Exposed to 1.25 GHZ pulsed Microwaves

DTIC Science & Technology

1992-03-13

matter. The entire retina was examined by indirect ophthalmoscopy. Photographic documentation included photomicrographs of the corneal endothelial cells...segment and retina appear normal. Photos of corneal endothelium pre and post irradiation demonstrated no change. The retinal photos also appeared normal...and photographic documentation. However, baseline fluorescein angiograms of the retina and iris were also included. It was determined that baseline

FITC-modified PPy nanotubes embedded in nanoporous AAO membrane can detect trace PCB20 via fluorescence ratiometric measurement.

PubMed

Wang, Meiling; Meng, Guowen; Huang, Qing; Xu, Qiaoling; Chu, Zhaoqin; Zhu, Chuhong

2011-04-07

A highly sensitive and selective fluorescence ratiometric sensor membrane for 2,3,3'-trichlorobiphenyl has been achieved, via depositing polypyrrole nanotubes (PPyNTs, the fluorescence indicator) in nano-porous anodic aluminium oxide (NPAAO) template and subsequently immobilizing fluorescein isothiocyanate (as an internal reference) onto the inner walls of the PPyNTs embedded in the NPAAO.

Evaluation of changes in serum chemistry in association with feed withdrawal or high dose oral gavage with Dextran Sodium Sulfate (DSS) induced gut leakage in broiler chickens

USDA-ARS?s Scientific Manuscript database

Dextran sodium sulfate (DSS) has been shown to be effective at inducing enteric inflammation in broiler chickens, resulting in increased leakage of orally administered fluorescein isothiocyanate dextran to circulation. In a previous study, two doses of DSS (0.45g/dose) administered as oral gavage re...

Synthesis and Characterization of Functional Mesostructures Using Colloidal Crystal Templating

DTIC Science & Technology

2004-01-01

fluorescent probes in aqueous polymer solutions . Khoury and co-workers measured the diffusion coefficient of several fluorescein-labeled proteins in...diffraction naq refractive index of the aqueous solution phase xvii ni refractive index of component i ngel refractive index of the hydrogel...phase Tg glass transition temperature α angle of diffraction φaq volume fraction of the aqueous solution phase φi volume fraction of

Imidazole catalyzes chlorination by unreactive primary chloramines

PubMed Central

Roemeling, Margo D.; Williams, Jared; Beckman, Joseph S.; Hurst, James K.

2015-01-01

Hypochlorous acid and simple chloramines (RNHCl) are stable biologically-derived chlorinating agents. In general, the chlorination potential of HOCl is much greater than that of RNHCl, allowing it to oxidize or chlorinate a much wider variety of reaction partners. However, in this study we demonstrate by kinetic analysis that the reactivity of RNHCl can be dramatically promoted by imidazole and histidyl model compounds via intermediary formation of the corresponding imidazole chloramines. Two biologically relevant reactions were investigated—loss of imidazole-catalyzed chlorinating capacity and phenolic ring chlorination using fluorescein and the tyrosine analog, 4-hydroxyphenylacetic acid (HPA). HOCl reacted stoichiometrically with imidazole, N-acetylhistidine (NAH), or imidazoleacetic acid to generate the corresponding imidazole chloramines which subsequently decomposed. Chloramine (NH2Cl) also underwent a markedly accelerated loss in chlorinating capacity when NAH was present, although in this case NAHCl did not accumulate, indicating that the catalytic intermediate must be highly reactive. Mixing HOCl with 1-methylimidazole (MeIm) led to very rapid loss in chlorinating capacity via formation of a highly reactive chlorinium ion (MeImCl+) intermediate; this behavior suggests that the reactive forms of the analogous imidazole chloramines are their conjugate acids, e.g., the imidazolechlorinium ion (HImCl+). HOCl-generated imidazole chloramine (ImCl) reacted rapidly with fluorescein in a specific acid-catalyzed second order reaction to give 3′-monochloro and 3′,5′-dichloro products. Equilibrium constants for the transchlorination reactions: HOCl + HIm = H2O + ImCl and NH2Cl + HIm = NH3 + ImCl were estimated from the dependence of the rate constants upon [HIm]/[HOCl] and literature data. Acid catalysis again suggests that the actual chlorinating agent is HImCl+; consistent with this interpretation, MeIm markedly catalyzed fluorescein chlorination by HOCl. Time-dependent imidazole-catalyzed HPA chlorination by NH2Cl was also demonstrated by product analyses. Quantitative assessment of the data suggests that physiological levels of histidyl groups will react with primary chloramines to generate a flux of imidazole chloramine sufficient to catalyze biological chlorination via HImCl+, particularly in environments that generate high concentrations of HOCl such as the neutrophil phagosome. PMID:25660996

MO-AB-BRA-04: Radiation Measurements with a DNA Double-Strand-Break Dosimeter

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

Obeidat, M; Cline, K; Stathakis, S

Purpose: Many types of dosimeters are used to measure radiation, but none of them directly measures the biological effect of this dose. The purpose here is to create a dosimeter that can measure the probability of double-strand breaks (DSB) for DNA, which is directly related to the biological effect of radiation. Methods: The dosimeter has DNA strands, which are labeled on one end with biotin and on the other with fluorescein. The biotin attaches these strands to magnetic beads. We suspended the DNA dosimeter in phosphate-buffered saline (PBS) as it matches the internal environment of the body. We placed smallmore » volumes (50µL) of the DNA dosimeter into tubes and irradiated these samples in a water-equivalent plastic phantom with several doses (three samples per dose). After irradiating the samples, a magnet was placed against the tubes. The fluorescein attached to broken DNA strands was extracted (called the supernatant) and placed into a different tube. The fluorescein on the unbroken strands remained attached to the beads in the tube and was re-suspended with 50µL of PBS. A fluorescence reader was used to measure the fluorescence for both the re-suspended beads and supernatant. To prove that we are measuring DSB, we tested dosimeter response with two different lengths of attached DNA strands (1 and 4 kilo-base pair). Results: The probability of DSB at the dose levels of 5, 10, 25, and 50 Gy were 0.05, 0.08, 0.12, and 0.19, respectively, while the coefficients of variation were 0.14, 0.07, 0.02, and 0.01, respectively. The 4 kilo-base-pair dosimeter produced 5.3 times the response of the 1 kilo-base-pair dosimeter. Conclusion: The DNA dosimeter yields a measurable response to dose that scales with the DNA strand length. The goal now is to refine the dosimeter fabrication to reproducibly create a low coefficient of variation for the lower doses. This work was supported in part by Yarmouk University (Irbid, Jordan) and CPRIT (RP140105)« less

A Quantitative and Standardized Method for the Evaluation of Choroidal Neovascularization Using MICRON III Fluorescein Angiograms in Rats

PubMed Central

Wigg, Jonathan P.; Zhang, Hong; Yang, Dong

2015-01-01

Introduction In-vivo imaging of choroidal neovascularization (CNV) has been increasingly recognized as a valuable tool in the investigation of age-related macular degeneration (AMD) in both clinical and basic research applications. Arguably the most widely utilised model replicating AMD is laser generated CNV by rupture of Bruch’s membrane in rodents. Heretofore CNV evaluation via in-vivo imaging techniques has been hamstrung by a lack of appropriate rodent fundus camera and a non-standardised analysis method. The aim of this study was to establish a simple, quantifiable method of fluorescein fundus angiogram (FFA) image analysis for CNV lesions. Methods Laser was applied to 32 Brown Norway Rats; FFA images were taken using a rodent specific fundus camera (Micron III, Phoenix Laboratories) over 3 weeks and compared to conventional ex-vivo CNV assessment. FFA images acquired with fluorescein administered by intraperitoneal injection and intravenous injection were compared and shown to greatly influence lesion properties. Utilising commonly used software packages, FFA images were assessed for CNV and chorioretinal burns lesion area by manually outlining the maximum border of each lesion and normalising against the optic nerve head. Net fluorescence above background and derived value of area corrected lesion intensity were calculated. Results CNV lesions of rats treated with anti-VEGF antibody were significantly smaller in normalised lesion area (p<0.001) and fluorescent intensity (p<0.001) than the PBS treated control two weeks post laser. The calculated area corrected lesion intensity was significantly smaller (p<0.001) in anti-VEGF treated animals at 2 and 3 weeks post laser. The results obtained using FFA correlated with, and were confirmed by conventional lesion area measurements from isolectin stained choroidal flatmounts, where lesions of anti-VEGF treated rats were significantly smaller at 2 weeks (p = 0.049) and 3 weeks (p<0.001) post laser. Conclusion The presented method of in-vivo FFA quantification of CNV, including acquisition variable corrections, using the Micron III system and common use software establishes a reliable method for detecting and quantifying CNV enabling longitudinal studies and represents an important alternative to conventional CNV quantification methods. PMID:26024231

Expression of SIRT1 and oxidative stress in diabetic dry eye.

PubMed

Liu, Hao; Sheng, Minjie; Liu, Yu; Wang, Peng; Chen, Yihui; Chen, Li; Wang, Weifang; Li, Bing

2015-01-01

To explore the expression of SIRT1 with oxidative stress and observe physiological and pathological changes in the corneas as well as the association between SIRT1 and oxidative stress of diabetic dry eyes in mice. Forty-eight C57BL/6Jdb/db mice at eight weeks of age were divided randomly into two groups: the diabetic dry eye group and the diabetic group. An additional forty-eight C57BL/6J mice at eight weeks of age were divided randomly into two groups: the dry eye group and the control group. Every mouse in the dry eye groups (diabetic and normal) was injected with scopolamine hydrobromide three times daily, combined with low humidity to establish a dry eye model. After the intervention, phenol red cotton string tests and corneal fluorescein staining were performed. In addition, HE staining and immunofluorescence were done. Expression of SIRT1 in the cornea was examined by real-time PCR and Western Blot and expression of FOXO3 and MnSOD proteins was detected by Western Blot. At one, four, and eight weeks post intervention, all of the groups except the controls showed significant decreases in tear production and increases in the corneal fluorescein stain (P<0.05 vs control). Between the experimental groups, the diabetic dry eye group had the least tear production and the highest corneal fluorescein stain score (P<0.05). As the disease progressed, all of the experimental groups showed obviously pathological changes in HE staining, particularly the diabetic dry eye group. In the 1(st) and 4(th) week, the expression of SIRT1, FOXO3, and MnSOD were significantly higher in the diabetic DE and DM groups but lower in the DE group compared to the controls (P<0.05). In the 8(th) week, the expression of SIRT1, FOXO3, and MnSOD was significantly down-regulated in the diabetic DE group and the DM group (P<0.05). Immunofluorescence showed similar results. In the condition of diabetic dry eye, tear production declined markedly coupled with seriously wounded corneal epithelium. Oxidative stress in the cornea was enhanced significantly and the expression of SIRT1 was decreased.

Influence of Fluorescein Angiography on the Diagnosis and Management of Retinopathy of Prematurity

PubMed Central

Klufas, Michael A.; Patel, Samir N.; Ryan, Michael C.; Gupta, Mrinali Patel; Jonas, Karyn E.; Ostmo, Susan; Martinez-Castellanos, Maria Ana; Berrocal, Audina M.; Chiang, Michael F.; Chan, R.V. Paul

2016-01-01

Purpose To examine the influence of fluorescein angiography (FA) on the diagnosis and management of retinopathy of prematurity (ROP). Design Prospective cohort study. Participants Nine recognized ROP experts (3 pediatric ophthalmologists; 6 retina specialists) interpreted 32 sets (16 color fundus photographs; 16 color fundus photographs paired with the corresponding FAs) of wide-angle retinal images from infants with ROP. Methods All experts independently reviewed the 32 image sets on a secure web site and provided a diagnosis and management plan for the case presented, first based on color fundus photographs alone, and then by color fundus photographs and corresponding FA. Main Outcome Measures Sensitivity and specificity of the ROP diagnosis (zone, stage, plus disease, and category – i.e. no ROP, mild ROP, type-2 ROP, and treatment-requiring ROP) was calculated using a consensus reference standard diagnosis, determined from the diagnosis of the color fundus photographs by three experienced readers in combination with the clinical diagnosis based on ophthalmoscopic examination. The kappa statistic was used to analyze the average intergrader agreement among experts for the diagnosis of zone, stage, plus disease, and category. Results Addition of FA to color fundus photographs resulted in a significant improvement in sensitivity for diagnosis of stage 3 or worse disease (39.8% vs. 74.1%, P = 0.008), type-2 or worse ROP (69.4% vs. 86.8%, P = 0.013), and pre-plus or worse disease (50.5 vs. 62.6%, P = 0.031). There was a nonsignificant trend towards improved sensitivity for diagnosis of treatment-requiring ROP (22.2% vs. 40.3%, P = 0.063). Using the kappa statistic, addition of FA to color fundus photographs significantly improved intergrader agreement for diagnosis of treatment-requiring ROP. Addition of FA to color fundus photographs did not significantly affect intergrader agreement for the diagnosis of stage, zone, or plus disease. Conclusions Compared to color fundus photographs alone, fluorescein angiography may improve the sensitivity of diagnosis of ROP by experts, particularly for stage 3 disease. In addition, intergrader agreement for diagnosis of treatment-requiring ROP may improve with FA interpretation. PMID:26028345

Severe blood-brain barrier disruption and surrounding tissue injury.

PubMed

Chen, Bo; Friedman, Beth; Cheng, Qun; Tsai, Phil; Schim, Erica; Kleinfeld, David; Lyden, Patrick D

2009-12-01

Blood-brain barrier opening during ischemia follows a biphasic time course, may be partially reversible, and allows plasma constituents to enter brain and possibly damage cells. In contrast, severe vascular disruption after ischemia is unlikely to be reversible and allows even further extravasation of potentially harmful plasma constituents. We sought to use simple fluorescent tracers to allow wide-scale visualization of severely damaged vessels and determine whether such vascular disruption colocalized with regions of severe parenchymal injury. Severe vascular disruption and ischemic injury was produced in adult Sprague Dawley rats by transient occlusion of the middle cerebral artery for 1, 2, 4, or 8 hours, followed by 30 minutes of reperfusion. Fluorescein isothiocyanate-dextran (2 MDa) was injected intravenously before occlusion. After perfusion-fixation, brain sections were processed for ultrastructure or fluorescence imaging. We identified early evidence of tissue damage with Fluoro-Jade staining of dying cells. With increasing ischemia duration, greater quantities of high molecular weight dextran-fluorescein isothiocyanate invaded and marked ischemic regions in a characteristic pattern, appearing first in the medial striatum, spreading to the lateral striatum, and finally involving cortex; maximal injury was seen in the mid-parietal areas, consistent with the known ischemic zone in this model. The regional distribution of the severe vascular disruption correlated with the distribution of 24-hour 2,3,5-triphenyltetrazolium chloride pallor (r=0.75; P<0.05) and the cell death marker Fluoro-Jade (r=0.86; P<0.05). Ultrastructural examination showed significantly increased areas of swollen astrocytic foot process and swollen mitochondria in regions of high compared to low leakage, and compared to contralateral homologous regions (ANOVA P<0.01). Dextran extravasation into the basement membrane and surrounding tissue increased significantly from 2 to 8 hours of occlusion duration (Independent samples t test, P<0.05). Severe vascular disruption, as labeled with high-molecular-weight dextran-fluorescein isothiocyanate leakage, is associated with severe tissue injury. This marker of severe vascular disruption may be useful in further studies of the pathoanatomic mechanisms of vascular disruption-mediated tissue injury.

Diffusion coefficients of organic molecules in sucrose-water solutions and comparison with Stokes-Einstein predictions

NASA Astrophysics Data System (ADS)

Chenyakin, Yuri; Ullmann, Dagny A.; Evoy, Erin; Renbaum-Wolff, Lindsay; Kamal, Saeid; Bertram, Allan K.

2017-02-01

The diffusion coefficients of organic species in secondary organic aerosol (SOA) particles are needed to predict the growth and reactivity of these particles in the atmosphere. Previously, viscosity measurements, along with the Stokes-Einstein relation, have been used to estimate the diffusion rates of organics within SOA particles or proxies of SOA particles. To test the Stokes-Einstein relation, we have measured the diffusion coefficients of three fluorescent organic dyes (fluorescein, rhodamine 6G and calcein) within sucrose-water solutions with varying water activity. Sucrose-water solutions were used as a proxy for SOA material found in the atmosphere. Diffusion coefficients were measured using fluorescence recovery after photobleaching. For the three dyes studied, the diffusion coefficients vary by 4-5 orders of magnitude as the water activity varied from 0.38 to 0.80, illustrating the sensitivity of the diffusion coefficients to the water content in the matrix. At the lowest water activity studied (0.38), the average diffusion coefficients were 1.9 × 10-13, 1.5 × 10-14 and 7.7 × 10-14 cm2 s-1 for fluorescein, rhodamine 6G and calcein, respectively. The measured diffusion coefficients were compared with predictions made using literature viscosities and the Stokes-Einstein relation. We found that at water activity ≥ 0.6 (which corresponds to a viscosity of ≤ 360 Pa s and Tg/T ≤ 0.81), predicted diffusion rates agreed with measured diffusion rates within the experimental uncertainty (Tg represents the glass transition temperature and T is the temperature of the measurements). When the water activity was 0.38 (which corresponds to a viscosity of 3.3 × 106 Pa s and a Tg/T of 0.94), the Stokes-Einstein relation underpredicted the diffusion coefficients of fluorescein, rhodamine 6G and calcein by a factor of 118 (minimum of 10 and maximum of 977), a factor of 17 (minimum of 3 and maximum of 104) and a factor of 70 (minimum of 8 and maximum of 494), respectively. This disagreement is significantly smaller than the disagreement observed when comparing measured and predicted diffusion coefficients of water in sucrose-water mixtures.

CdS/TiO2-fluorescein isothiocyanate nanoparticles as fluorescence resonance energy transfer probe for the determination of trace alkaline phosphatase based on affinity adsorption assay.

PubMed

Liu, Jia-Ming; Lin, Li-ping; Jiao, Li; Cui, Ma-Lin; Wang, Xin-Xing; Zhang, Li-Hong; Zheng, Zhi-Yong

2012-08-30

The CdS/TiO(2)-fluorescein isothiocyanate (FITC) luminescent nanoparticles (CdS/TiO(2)-FITC) with the particle size of 20 nm have been synthesized by sol-gel method. CdS/TiO(2)-FITC could emit the fluorescence of both FITC and CdS/TiO(2). The fluorescence resonance energy transfer (FRET) occurred between the donor CdS/TiO(2) and the acceptor FITC in the CdS/TiO(2)-FITC. Taking advantages of the excellent characteristics of FRET, a new CdS/TiO(2)-FITC FRET labeling reagent and a CdS/TiO(2)-FITC-wheat germ agglutinin (CdS/TiO(2)-FITC-WGA) fluorescent probe have been developed. The FRET occurring between the donor CdS/TiO(2) and the acceptor FITC in the labelled product CdS/TiO(2)-FITC-WGA-AP, formed in the affinity adsorption reaction between the WGA in this CdS/TiO(2)-FITC-WGA fluorescent probe and alkaline phosphatase (AP), sharply enhanced the fluorescence signal of FITC and quench the fluorescence signal of CdS/TiO(2). Moreover, the ΔF (the change of the fluorescence signal) of FITC and CdS/TiO(2) were proportional to the content of AP, respectively. Thus, a new method that CdS/TiO(2)-fluorescein isothiocyanate nanoparticles for the determination of trace AP based on FRET-affinity adsorption assay has been established. The limit of quantification (LOQ) of the method was 1.3×10(-17) g AP mL(-1) for CdS/TiO(2) and 1.1×10(-17) g AP mL(-1) for FITC, respectively. This sensitive, rapid, high selective and precise method has been applied to the determination of AP in human serum and the prediction of human disease with the results agreed well with enzyme-linked immunosorbent assay (ELISA) in Zhangzhou Municipal Hospital of Fujian Province. Simultaneously, the reaction mechanism for the determination of AP was also discussed. Copyright © 2012 Elsevier B.V. All rights reserved.

Ultrasound-triggered drug delivery using acoustic droplet vaporization

NASA Astrophysics Data System (ADS)

Fabiilli, Mario Leonardo

The goal of targeted drug delivery is the spatial and temporal localization of a therapeutic agent and its associated bioeffects. One method of drug localization is acoustic droplet vaporization (ADV), whereby drug-laden perfluorocarbon (PFC) emulsions are vaporized into gas bubbles using ultrasound, thereby releasing drug locally. Transpulmonary droplets are converted into bubbles that occlude capillaries, sequestering the released drug within an organ or tumor. This research investigates the relationship between the ADV and inertial cavitation (IC) thresholds---relevant for drug delivery due to the bioffects generated by IC---and explores the delivery of lipophilic and hydrophilic compounds using PFC double emulsions. IC can positively and negatively affect ultrasound mediated drug delivery. The ADV and IC thresholds were determined for various bulk fluid, droplet, and acoustic parameters. At 3.5 MHz, the ADV threshold occurred at a lower rarefactional pressure than the IC threshold. The results suggest that ADV is a distinct phenomenon from IC, the ADV nucleus is internal to the droplet, and the IC nucleus is the bubble generated by ADV. The ADV triggered release of a lipophilic chemotherapeutic agent, chlorambucil (CHL), from a PFC-in-oil-in-water emulsion was explored using plated cells. Cells exposed to a CHL-loaded emulsion, without ADV, displayed 44% less growth inhibition than cells exposed to an equal concentration of CHL in solution. Upon ADV of the CHL-loaded emulsion, the growth inhibition increased to the same level as cells exposed to CHL in solution. A triblock copolymer was synthesized which enabled the formulation of stable water-in-PFC-in-water (W1/PFC/W2) emulsions. The encapsulation of fluorescein in the W1 phase significantly decreased the mass flux of fluorescein; ADV was shown to completely release the fluorescein from the emulsions. ADV was also shown to release thrombin, dissolved in the W1 phase, which could be used in vivo to extend synergistically the duration of ADV-generated, microbubble-based embolizations. Overall, the results suggest that PFC double emulsions can be used as an ultrasound-triggered drug delivery system. Compared to traditional drug delivery systems, ADV could be used to increase the therapeutic efficacy and decrease the systemic toxicity of drug therapy.

Multifunctional Polymer Microbubbles for Advanced Sentinel Lymph Node Imaging and Mapping

DTIC Science & Technology

2012-06-01

of thiolated poly(acrylic acid) with fluorescein attached. (b) Bright field image of large bubbles stabilized by polymer and phospholipid...Page 1 of 6 AD_________________ Award Number: W81XWH-11-1-0215 TITLE:   Multifunctional Polymer Microbubbles for Advanced... Polymer Microbubbles for Advanced Sentinel Lymph Node Imaging and Mapping 5b. GRANT NUMBER W81XWH-11-1-0215   5c. PROGRAM ELEMENT NUMBER 6

[Central aleolar choroidal dystrophy in sibilings coexisting with alopecia].

PubMed

Brydak-Godowska, Joanna; Dróbecka-Brydak, Ewa; Paćkowska, Maria; Kecik, Dariusz

2007-01-01

Central areolar choroidal dystrophy is localized in macular region and is characterized by atrophy of pigment epithelium, photoreceptors and choriocapillaris. This paper presents the history of two sibilings at the age of 23 and 30, with central aleolar choroidal dystrophy coexisting with alopecia. The results of erg, eog and fluorescein angiography are presented. The results of therapy for glaucoma associated with the Sturge-Weber syndrome are often disappointing.

The operation was a success, but the patient cannot see.

PubMed

Semmer, Anne E; Lee, Michael S; Taban, Mehran; Smith, Scott; Kosmorsky, Gregory

2009-01-01

A 71-year-old African American woman presented with severe vision loss in her left eye one day following trabeculectomy with mitomycin C and retrobulbar anesthesia. She had a new left relative afferent pupillary defect and macular whitening. The optic disc appeared normal. Intraocular pressure and fluorescein angiography were normal. Westergren erythrocyte sedimentation rate and C-reactive protein were elevated. Temporal artery biopsy was positive for giant cell arteritis.

The Effect of Autologous Platelet Lysate Eye Drops: An In Vivo Confocal Microscopy Study

PubMed Central

Fea, Antonio M.; Testa, Valeria; Machetta, Federica; Parisi, Simone; D'Antico, Sergio; Spinetta, Roberta; Fusaro, Enrico; Grignolo, Federico M.

2016-01-01

Purpose. To determine the effectiveness of autologous platelet lysate (APL) eye drops in patients with primary Sjögren syndrome (SS) dry eye, refractory to standard therapy, in comparison with patients treated with artificial tears. We focused on the effect of APL on cornea morphology with the in vivo confocal microscopy (IVCM). Methods. Patients were assigned to two groups: group A used autologous platelet lysate QID, and group B used preservative-free artificial tears QID, for 90 days. Ophthalmological assessments included ocular surface disease index (OSDI), best corrected visual acuity (BCVA), Schirmer test, fluorescein score, and breakup time (BUT). A subgroup of patients in group A underwent IVCM: corneal basal epithelium, subbasal nerves, Langerhans cells, anterior stroma activated keratocytes, and reflectivity were evaluated. Results. 60 eyes of 30 patients were enrolled; in group A (n = 20 patients) mean OSDI, fluorescein score, and BUT showed significant improvement compared with group B (n = 10 patients). The IVCM showed a significant increase in basal epithelium cells density and subbasal nerve plexus density and number and a decrease in Langerhans cells density (p < 0.05). Conclusion. APL was found effective in the treatment of SS dry eye. IVCM seems to be a useful tool to visualize cornea morphologic modifications. PMID:27200376

Determination of Trace Deoxyribonucleic Acid by Using Fluorescein Isothiocyanate-Phenosafranine as a Double-Luminescent Phosphorescence Probe

PubMed Central

Huang, Xiao-Mei; Liu, Zhen-Bo; Li, Fei-Ming; Lin, Li-Ping; Wang, Xin-Xing; Lin, Chang-Qing; Huang, Ya-Hong; Li, Zhi-Ming; Lin, Shao-Qin

2010-01-01

Using Pb2+ as ion perturber, phenosafranine (PF) and fluorescein isothiocyanate (FITC) could emit strong and stable room temperature phosphorescence (RTP) signal on the filter paper, respectively. When they were mixed, the phenomenon that the RTP signal of PF and FITC enhanced significantly was found. And 1.12 ag DNA spot−1 (sample volume was 0.40 μL, corresponding concentration was 2.8 × 10–15 g mL–1) could cause the RTP signal of both PF and FITC to enhance sharply. The content of DNA was proportional to the ΔIp of PF and FITC in the system at 634 and 659 nm. Thus, a new solid substrate room temperature phosphorimetry (SSRTP) for the determination of trace DNA was established by using FITC-PF as double-luminescent phosphorescence probe. The detection limit (LD) of this method calculated by 3Sb/k was 14 zg DNA spot–1 for PF and 18 zg DNA spot–1 for FITC, respectively, showing high sensitivity. It has been applied to the determination of trace DNA in practical samples and the analysis results were in accordance with those of fluorescence probe. The reaction mechanism of SSRTP for the determination of trace DNA was also discussed. PMID:20665096

Antigenic specificity and morphologic characteristics of Chlamydia trachomatis, strain SFPD, isolated from hamsters with proliferative ileitis.

PubMed

Fox, J G; Stills, H F; Paster, B J; Dewhirst, F E; Yan, L; Palley, L; Prostak, K

1993-10-01

Profound diarrhea associated with proliferating intestinal cells containing intraepithelial campylobacter-like organisms (ICLO) occurs in a variety of mammalian hosts, particularly swine and hamsters. Recently, intracellular bacteria were isolated from proliferative intestinal tissue of hamsters and propagated in intestine cell line 407. Oral inoculation of hamsters with cell culture lysates containing these organisms reproduced the disease in susceptible hamsters. In the present study, an intracellular bacterium from the INT 407 cell line was shown by a variety of techniques to be a member of the genus Chlamydia and has been designated Chlamydia sp. strain SFPD. McCoy cells infected with Chlamydia sp. strain SFPD demonstrated bright fluorescent-stained intracytoplasmic inclusions when examined with fluorescein-labeled species-specific C. trachomatis monoclonal antibodies. The organism also reacted to fluorescein-labeled polyclonal but not monoclonal ICLO "omega" antisera. Ultrastructural examination of the Chlamydia sp. strain SFPD from McCoy cells revealed electrondense elementary bodies and a less electron-dense reticulate-like body that was circular; both features are consistent in morphology to developmental forms of Chlamydia and do not conform to ICLO morphology. Molecular studies, 16S ribosomal sequence analysis, and sequencing of the outer membrane protein confirmed that the isolate is a C. trachomatis closely related to the mouse pneumonitis strain of C. trachomatis.

The Effect of Autologous Platelet Lysate Eye Drops: An In Vivo Confocal Microscopy Study.

PubMed

Fea, Antonio M; Aragno, Vittoria; Testa, Valeria; Machetta, Federica; Parisi, Simone; D'Antico, Sergio; Spinetta, Roberta; Fusaro, Enrico; Grignolo, Federico M

2016-01-01

Purpose. To determine the effectiveness of autologous platelet lysate (APL) eye drops in patients with primary Sjögren syndrome (SS) dry eye, refractory to standard therapy, in comparison with patients treated with artificial tears. We focused on the effect of APL on cornea morphology with the in vivo confocal microscopy (IVCM). Methods. Patients were assigned to two groups: group A used autologous platelet lysate QID, and group B used preservative-free artificial tears QID, for 90 days. Ophthalmological assessments included ocular surface disease index (OSDI), best corrected visual acuity (BCVA), Schirmer test, fluorescein score, and breakup time (BUT). A subgroup of patients in group A underwent IVCM: corneal basal epithelium, subbasal nerves, Langerhans cells, anterior stroma activated keratocytes, and reflectivity were evaluated. Results. 60 eyes of 30 patients were enrolled; in group A (n = 20 patients) mean OSDI, fluorescein score, and BUT showed significant improvement compared with group B (n = 10 patients). The IVCM showed a significant increase in basal epithelium cells density and subbasal nerve plexus density and number and a decrease in Langerhans cells density (p < 0.05). Conclusion. APL was found effective in the treatment of SS dry eye. IVCM seems to be a useful tool to visualize cornea morphologic modifications.

Binding of fluoresceinated epidermal growth factor to A431 cell sub-populations studied using a model-independent analysis of flow cytometric fluorescence data.

PubMed Central

Chatelier, R C; Ashcroft, R G; Lloyd, C J; Nice, E C; Whitehead, R H; Sawyer, W H; Burgess, A W

1986-01-01

A method is developed for determining ligand-cell association parameters from a model-free analysis of data obtained with a flow cytometer. The method requires measurement of the average fluorescence per cell as a function of ligand and cell concentration. The analysis is applied to data obtained for the binding of fluoresceinated epidermal growth factor to a human epidermoid carcinoma cell line, A431. The results indicate that the growth factor binds to two classes of sites on A431 cells: 4 X 10(4) sites with a dissociation constant (KD) of less than or equal to 20 pM, and 1.5 X 10(6) sites with a KD of 3.7 nM. A derived plot of the average fluorescence per cell versus the average number of bound ligands per cell is used to construct binding isotherms for four sub-populations of A431 cells fractionated on the basis of low-angle light scatter. The four sub-populations bind the ligand with equal affinity but differ substantially in terms of the number of binding sites per cell. We also use this new analysis to critically evaluate the use of 'Fluorotrol' as a calibration standard in flow cytometry. PMID:3015587

STRUCTURAL AND FUNCTIONAL CHARACTERIZATION OF BENIGN FLECK RETINA USING MULTIMODAL IMAGING.

PubMed

Neriyanuri, Srividya; Rao, Chetan; Raman, Rajiv

2017-01-01

To report structural and functional features in a case series of benign fleck retina using multimodal imaging. Four cases with benign fleck retina underwent complete ophthalmic examination that included detailed history, visual acuity, and refractive error testing, FM-100 hue test, dilated fundus evaluation, full field electroretinogram, fundus photography with autofluorescence, fundus fluorescein angiography, and swept-source optical coherence tomography. Age group of the cases ranged from 19 years to 35 years (3 males and 1 female). Parental consanguinity was reported in two cases. All of them were visually asymptomatic with best-corrected visual acuity of 20/20 (moderate astigmatism) in both the eyes. Low color discrimination was seen in two cases. Fundus photography showed pisciform flecks which were compactly placed on posterior pole and were discrete, diverging towards periphery. Lesions were seen as smaller dots within 1500 microns from fovea and were hyperfluorescent on autofluorescence. Palisading retinal pigment epithelium defects were seen in posterior pole on fundus fluorescein angiography imaging; irregular hyper fluorescence was also noted. One case had reduced cone responses on full field electroretinogram; the other three cases had normal electroretinogram. On optical coherence tomography, level of lesions varied from retinal pigment epithelium, inner segment to outer segment extending till external limiting membrane. Functional and structural deficits in benign fleck retina were picked up using multimodal imaging.

Long-term effects of short-term retinal bleb detachments in rabbits.

PubMed

Ivert, Lena; Kjeldbye, Hild; Gouras, Peter

2002-03-01

To examine the effects of saline-induced bleb detachments in rabbit retina. Retinal bleb detachments were produced by the injection of 50 microl of balanced salt solution (BSS) into the subretinal space of one eye of each of six rabbits using a glass pipette with a flat tip, 50 microm in diameter. The retina was examined by biomicroscopy, scanning laser ophthalmoscopy (SLO), auto-fluorescence and simultaneous fluorescein and indocyanine green (ICG) angiography. Histological examination was carried out at 1, 2, 3 and 4 months after surgery. All rabbits showed leakage of fluorescein for at least a day after detachment, but within 1 month the leakage ceased. ICG staining developed gradually at the level of the RPE or Bruch's membrane near sites of previous staining. Lipofuscin fluorescence also developed gradually around areas of staining. Histology revealed the source of the excessive lipofuscin to be in the RPE layer, especially in cells migrating away from Bruch's membrane. Short-term bleb detachments cause a transient breakdown in the blood-retinal barrier, long-term ICG staining at or deep to the RPE layer, hyperlipofuscinosis and migration of the RPE. The abnormal lipofuscin accumulation is apparent on fluorescence ophthalmoscopy and can be confused with markers such as green fluorescent protein.

Adenosine receptor agonist NECA increases cerebral extravasation of fluorescein and low molecular weight dextran independent of blood-brain barrier modulation

PubMed Central

Cheng, Chih-Chung; Yang, Ya Lan; Liao, Kate Hsiurong; Lai, Ted Weita

2016-01-01

Conventional methods for therapeutic blood-brain barrier (BBB) disruption facilitate drug delivery but are cumbersome to perform. A previous study demonstrated that adenosine receptor (AR) stimulation by 5′-N-ethylcarboxamide adenosine (NECA) increased the extravasation of intravascular tracers into the brain and proposed that AR agonism may be an effective method for therapeutic BBB disruption. We attempted to confirm the extravasation of tracers into the brain and also investigated tracer extravasation into peripheral organs and tracer retention in the blood. We found that NECA not only increased the extravasation of intravascular fluorescein and low molecular weight dextran into the brain of mice but also increased the concentrations of these tracers in the blood. In fact, the brain:blood ratio-normalized BBB permeability for either tracer is actually decreased by NECA administration. Elevated blood urea nitrogen levels in mice following NECA treatment suggested that renal function impairment was a probable cause of tracer retention. Therefore, NECA has almost no effect on the extravasation of intravascular Evans blue dye (EBD), an albumin-binding tracer with little renal clearance. Rather than inducing BBB disruption, our study demonstrated that NECA increased tracer extravasation into the brain by increasing the concentration gradient of the tracer across the BBB. PMID:27025761

Nanosecond to submillisecond dynamics in dye-labeled single-stranded DNA, as revealed by ensemble measurements and photon statistics at single-molecule level.

PubMed

Kaji, Takahiro; Ito, Syoji; Iwai, Shigenori; Miyasaka, Hiroshi

2009-10-22

Single-molecule and ensemble time-resolved fluorescence measurements were applied for the investigation of the conformational dynamics of single-stranded DNA, ssDNA, connected with a fluorescein dye by a C6 linker, where the motions both of DNA and the C6 linker affect the geometry of the system. From the ensemble measurement of the fluorescence quenching via photoinduced electron transfer with a guanine base in the DNA sequence, three main conformations were found in aqueous solution: a conformation unaffected by the guanine base in the excited state lifetime of fluorescein, a conformation in which the fluorescence is dynamically quenched in the excited-state lifetime, and a conformation leading to rapid quenching via nonfluorescent complex. The analysis by using the parameters acquired from the ensemble measurements for interphoton time distribution histograms and FCS autocorrelations by the single-molecule measurement revealed that interconversion in these three conformations took place with two characteristic time constants of several hundreds of nanoseconds and tens of microseconds. The advantage of the combination use of the ensemble measurements with the single-molecule detections for rather complex dynamic motions is discussed by integrating the experimental results with those obtained by molecular dynamics simulation.

Reliability of fluorescein dye disappearance test in assessment of adults with nasolacrimal duct obstruction.

PubMed

Kashkouli, Mohsen Bahmani; Mirzajani, Hoora; Jamshidian-Tehrani, Mansooreh; Pakdel, Farzad; Nojomi, Marzieh; Aghaei, Gholam-Hossein

2013-01-01

To evaluate the role of fluorescein dye disappearance test (FDDT) in assessment of adults with primary acquired nasolacrimal duct obstruction (PANDO) and its correlation with age and severity of epiphora. In a prospective, cross-sectional comparative study, FDDT at 2 (2-FDDT), 5 (5-FDDT), and 10 (10-FDDT) minutes were performed in 58 eyes of 58 patients with PANDO (case) and 58 eyes of 58 subjects without epiphora (control) between 2008 and 2010. Exclusion criteria were any disease, trauma, or medications that might have an effect on tear drainage system. Patients with functional or incomplete obstruction and upper lacrimal drainage system obstruction were excluded in the case group. FDDT and severity of epiphora were graded. Case and control groups were matched. 2-FDDT showed a sensitivity of 82.8%, specificity of 91.4%, positive predictive value of 90.6%, and negative predictive value of 84.1%. Sensitivity and negative predictive value decreased to 71.1% and 78.6% in 5-FDDT. 5-FDDT specificity and positive predictive value, however, increased to 94.8% and 93.5%, respectively. There was no statistically significant correlation between severity of epiphora and age with FDDT. FDDT is a simple, reliable, and highly specific test in assessment of adults with PANDO.

Optimization and validation of an existing, surgical and robust dry eye rat model for the evaluation of therapeutic compounds.

PubMed

Joossen, Cedric; Lanckacker, Ellen; Zakaria, Nadia; Koppen, Carina; Joossens, Jurgen; Cools, Nathalie; De Meester, Ingrid; Lambeir, Anne-Marie; Delputte, Peter; Maes, Louis; Cos, Paul

2016-05-01

The aim of this research was to optimize and validate an animal model for dry eye, adopting clinically relevant evaluation parameters. Dry eye was induced in female Wistar rats by surgical removal of the exorbital lacrimal gland. The clinical manifestations of dry eye were evaluated by tear volume measurements, corneal fluorescein staining, cytokine measurements in tear fluid, MMP-9 mRNA expression and CD3(+) cell infiltration in the conjunctiva. The animal model was validated by treatment with Restasis(®) (4 weeks) and commercial dexamethasone eye drops (2 weeks). Removal of the exorbital lacrimal gland resulted in 50% decrease in tear volume and a gradual increase in corneal fluorescein staining. Elevated levels of TNF-α and IL-1α have been registered in tear fluid together with an increase in CD3(+) cells in the palpebral conjunctiva when compared to control animals. Additionally, an increase in MMP-9 mRNA expression was recorded in conjunctival tissue. Reference treatment with Restasis(®) and dexamethasone eye drops had a positive effect on all evaluation parameters, except on tear volume. This rat dry eye model was validated extensively and judged appropriate for the evaluation of novel compounds and therapeutic preparations for dry eye disease. Copyright © 2016 Elsevier Ltd. All rights reserved.

Fluorometric prediction of successful amputation level in the ischemic limb.

PubMed

Silverman, D G; Rubin, S M; Reilly, C A; Brousseau, D A; Norton, K J; Wolf, G L

1985-01-01

The present study was undertaken to compare fluorometric documentation of fluorescein dye delivery with the standard means of determining the level at which an amputation should be performed in the dysvascular extremity. Thirty-nine patients underwent lower-extremity amputation at the level determined by the surgeon based upon physical examination, angiography, segmental pressure indices, and/or pulse volume recordings. In addition, fiberoptic fluorometry was performed preoperatively. After intravenous administration of sodium fluorescein (4-8 mg/kg), fluorometric readings were obtained by placing the fluorometer's light guide on 126 reading sites. Fluorometric findings were evaluated retrospectively, and therefore did not influence the surgeon's decision. Of the 39 amputations performed overall, only 26 healed. The accuracy of the standard criteria was lowest for the 20 below-ankle amputations, where only 12 cases healed. Alternatively, fluorometric indices separated healing from nonhealing sites in 36 of the 39 cases and in 18 of the 20 below-ankle amputations. Overall, healing sites averaged 94 percent of the fluorescence of the healthy reference area, while nonhealing sites averaged only 29 percent. We conclude that fluorometry should prove to be a valuable adjunct in the assessment of the dysvascular extremity. It uses a low dose of dye, is easy to perform, and is readily repeatable.

Isoflurane anesthesia results in reversible ultrastructure and occludin tight junction protein expression changes in hippocampal blood-brain barrier in aged rats.

PubMed

Cao, Yiyun; Ni, Cheng; Li, Zhengqian; Li, Lunxu; Liu, Yajie; Wang, Chunyi; Zhong, Yanfeng; Cui, Dehua; Guo, Xiangyang

2015-02-05

The underlying mechanism of isoflurane-induced cognitive dysfunction in older individuals is unknown. In this study, the effects of isoflurane exposure on the hippocampal blood-brain barrier (BBB) in aged rats were investigated because it was previously shown that BBB disruption involves in cognitive dysfunction. Twenty-month-old rats randomly received 1.5% isoflurane or vehicle gas as control. Hippocampal BBB ultrastructure was analyzed by transmission electron microscopy and expression of tight junction proteins was measured by western blot analysis. BBB permeability was detected with sodium fluorescein extravasation and further confirmed by immunoglobulin G immunohistochemistry. Spatial learning and memory were assessed by the Morris water maze test. Isoflurane anesthesia resulted in reversible time-dependent BBB ultrastructure morphological damage and significant decreases in expression of the tight junction proteins occludin, which contributed to sodium fluorescein and IgG leakage. Rats with isoflurane exposure also showed significant cognitive deficits in the Morris water maze test. This in vivo data indicate that occludin down-regulation may be one of the mediators of isoflurane-induced hippocampus BBB disruption, and may contribute to hippocampus-dependent cognitive impairment after isoflurane exposure in aged rats. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Hydrophobic interactions in donor-disulphide-acceptor (DSSA) probes looking beyond fluorescence resonance energy transfer theory.

PubMed

Sanjeeva, Shilpa Kammaradi; Korrapati, Swathi; Nair, Chandrasekhar B; Rao, P V Subba; Pullela, Phani Kumar; Vijayalakshmi, U; Siva, Ramamoorthy

2014-07-01

Donor-linker-acceptor (DSSA) is a concept in fluorescence chemistry with acceptor being a fluorescent compound (FRET) or quencher. The DSSA probes used to measure thiol levels in vitro and in vivo. The reduction potential of these dyes are in the range of -0.60 V, much lower than the best thiol reductant reported in literature, the DTT (-0.33 V). DSSA disulphide having an unusually low reduction potential compared to the typical thiol reductants is a puzzle. Secondly, DSSA probes have a cyclized rhodamine ring as acceptor which does not have any spectral overlap with fluorescein, but quenches its absorbance and fluorescence. To understand the structural features of DSSA probes, we have synthesized DSSANa and DSSAOr. The calculated reduction potential of these dyes suggest that DSSA probes have an alternate mechanism from the FRET based quenching, namely hydrophobic interaction or dye to dye quenching. The standard reduction potential change with increasing complexity and steric hindrance of the molecule is small, suggesting that ultra- low Eo' has no contribution from the disulphide linker and is based on structural interactions between fluorescein and cyclized rhodamine. Our results help to understand the DSSA probe quenching mechanism and provide ways to design fluorescent probes.

Fluorescence endoscopic imaging study of anastomotic recurrence of Crohn's disease after right ileocolonic resection

NASA Astrophysics Data System (ADS)

Mordon, Serge R.; Maunoury, Vincent; Klein, Olivier; Colombel, Jean-Frederic

1995-12-01

Crohn's disease is an inflammatory bowel disease of unknown etiology. Vasculitis is hypothesized but it was never demonstrated in vivo. This study aimed to evaluate the vascular mucosa perfusion using fluorescence imaging in 13 patients who had previously undergone eileocolonic resection and who agreed to participate in a prospective endoscopic study of anastomotic recurrence. This anastomotic recurrence rate is known to be high (73% after 1 year follow-up) and is characterized by ulcerations. The fluorescence study was started with an I.V. bolus injection of sodium fluorescein. The pre-anastomotic mucosa was endoscopically examined with blue light that stimulates fluorescein fluorescence. Fluorescence emission was recorded with an ultra-high-sensitivity camera connected to the endoscope via an interference filter (520 - 560 nm). A uniform fluorescence was observed a few seconds after the injection and lasted for 15 min in healthy subjects. In case of recurrence, the centers of the ulcerations displayed a very low fluorescence indicating localized ischemia. In contrast, the rims of the ulcers revealed brighter fluorescent images than those of normal mucosa. The anastomotic ulcerations of Crohn's disease recurrence exhibit a high fluorescence intensity at their margins indicating an increased mucosal blood flow and/or enhanced transcapillary diffusion. These findings support the hypothesis of a primary vasculitis in Crohn's disease.

Adenosine receptor agonist NECA increases cerebral extravasation of fluorescein and low molecular weight dextran independent of blood-brain barrier modulation.

PubMed

Cheng, Chih-Chung; Yang, Ya Lan; Liao, Kate Hsiurong; Lai, Ted Weita

2016-03-30

Conventional methods for therapeutic blood-brain barrier (BBB) disruption facilitate drug delivery but are cumbersome to perform. A previous study demonstrated that adenosine receptor (AR) stimulation by 5'-N-ethylcarboxamide adenosine (NECA) increased the extravasation of intravascular tracers into the brain and proposed that AR agonism may be an effective method for therapeutic BBB disruption. We attempted to confirm the extravasation of tracers into the brain and also investigated tracer extravasation into peripheral organs and tracer retention in the blood. We found that NECA not only increased the extravasation of intravascular fluorescein and low molecular weight dextran into the brain of mice but also increased the concentrations of these tracers in the blood. In fact, the brain:blood ratio-normalized BBB permeability for either tracer is actually decreased by NECA administration. Elevated blood urea nitrogen levels in mice following NECA treatment suggested that renal function impairment was a probable cause of tracer retention. Therefore, NECA has almost no effect on the extravasation of intravascular Evans blue dye (EBD), an albumin-binding tracer with little renal clearance. Rather than inducing BBB disruption, our study demonstrated that NECA increased tracer extravasation into the brain by increasing the concentration gradient of the tracer across the BBB.

Analysis of Fundus Photography and Fluorescein Angiography in Nonarteritic Anterior Ischemic Optic Neuropathy and Optic Neuritis.

PubMed

Kim, Min Kyung; Kim, Ungsoo Samuel

2016-08-01

We evaluated fundus and fluorescein angiography (FAG) findings and characteristics that can help distinguish nonarteritic anterior ischemic optic neuropathy (NAION) from optic neuritis (ON). Twenty-three NAION patients and 17 ON with disc swelling patients were enrolled in this study. We performed fundus photography and FAG. The disc-swelling pattern, hyperemia grade, presence of splinter hemorrhages, cotton-wool spots, artery/vein ratio and degree of focal telangiectasia were investigated. The FAG findings for each patient were compared with respect to the following features: the pattern of disc leakage in the early phase, arteriovenous (artery/vein) transit time (second), and the presence and pattern of the filling delay. Cotton-wool spots, focal telangiectasia, and venous congestion were more common in the affected eyes of NAION patients. Upon FAG, 76.5% of the patients in the ON group exhibited normal choroidal circulation. However, 56.5% of patients in the NAION group demonstrated abnormal filling defects, such as peripapillary, generalized, or watershed zone filling delays. Fundus findings, including cotton-wool spots, focal telangiectasia, and venous congestion in the affected eye, may be clues that can be used to diagnose NAION. In addition, choroidal insufficiencies on FAG could be also helpful in differentiating NAION from ON.

A fluorescein-based chemosensor for relay fluorescence recognition of Cu(ii) ions and biothiols in water and its applications to a molecular logic gate and living cell imaging.

PubMed

Fu, Zhen-Hai; Yan, Lu-Bin; Zhang, Xiaolong; Zhu, Fan-Fan; Han, Xin-Long; Fang, Jianguo; Wang, Ya-Wen; Peng, Yu

2017-05-16

Relay recognition of copper(ii) ions and biothiols via a fluorescence "on-off-on" cascade was designed and realized as a new sequential combination of cations and small molecules. Probe 1 bearing a fluorescein skeleton was thus synthesized, which performed well in 100% HEPES buffer (pH = 7.0) solution, as a highly sensitive, selective fluorescence sensor for Cu 2+ . The limit of detection (LOD, 0.017 ppm) was obtained, and this value is much lower than 1.3 ppm, allowed by US EPA. The 1 : 1 complex generated from fast sensing of Cu 2+ when excited at 491 nm, showed good relay recognition for biothiols (i.e., Cys, Hcy and GSH with low detection limits of 0.12 μM, 0.036 μM and 0.024 μM, respectively) via remarkable fluorescence enhancement. The origin of this relay process was disclosed through ESI-MS and corresponding density functional theory (DFT) computations. Notably, probe 1 can be utilized for the construction of a molecular logic gate with the IMPLICATION function by using the above fluorescence changes. Moreover, this relay recognition was also applied to HepG2 cell imaging successfully.

Genotyping of single nucleotide polymorphism by probe-gated silica nanoparticles.

PubMed

Ercan, Meltem; Ozalp, Veli C; Tuna, Bilge G

2017-11-15

The development of simple, reliable, and rapid approaches for molecular detection of common mutations is important for prevention and early diagnosis of genetic diseases, including Thalessemia. Oligonucleotide-gated mesoporous nanoparticles-based analysis is a new platform for mutation detection that has the advantages of sensitivity, rapidity, accuracy, and convenience. A specific mutation in β-thalassemia, one of the most prevalent inherited diseases in several countries, was used as model disease in this study. An assay for detection of IVS110 point mutation (A > G reversion) was developed by designing probe-gated mesoporous silica nanoparticles (MCM-41) loaded with reporter fluorescein molecules. The silica nanoparticles were characterized by AFM, TEM and BET analysis for having 180 nm diameter and 2.83 nm pore size regular hexagonal shape. Amine group functionalized nanoparticles were analysed with FTIR technique. Mutated and normal sequence probe oligonucleotides)about 12.7 nmol per mg nanoparticles) were used to entrap reporter fluorescein molecules inside the pores and hybridization with single stranded DNA targets amplified by PCR gave different fluorescent signals for mutated targets. Samples from IVS110 mutated and normal patients resulted in statistically significant differences when the assay procedure were applied. Copyright © 2017 Elsevier Inc. All rights reserved.

Growth promotion of genetically modified hematopoietic progenitors using an antibody/c-Mpl chimera.

PubMed

Kawahara, Masahiro; Chen, Jianhong; Sogo, Takahiro; Teng, Jinying; Otsu, Makoto; Onodera, Masafumi; Nakauchi, Hiromitsu; Ueda, Hiroshi; Nagamune, Teruyuki

2011-09-01

Thrombopoietin is a potent cytokine that exerts proliferation of hematopoietic stem cells (HSCs) through its cognate receptor, c-Mpl. Therefore, mimicry of c-Mpl signaling by a receptor recognizing an artificial ligand would be attractive to attain specific expansion of genetically modified HSCs. Here we propose a system enabling selective expansion of genetically modified cells using an antibody/receptor chimera that can be activated by a specific antigen. We constructed an antibody/c-Mpl chimera, in which single-chain Fv (ScFv) of an anti-fluorescein antibody was tethered to the extracellular D2 domain of the erythropoietin receptor and transmembrane/cytoplasmic domains of c-Mpl. When the chimera was expressed in interleukin (IL)-3-dependent pro-B cell line Ba/F3, genetically modified cells were selectively expanded in the presence of fluorescein-conjugated BSA (BSA-FL) as a specific antigen. Furthermore, highly purified mouse HSCs transduced with the retrovirus carrying antibody/c-Mpl chimera gene proliferated in vitro in response to BSA-FL, and the cells retained in vivo long-term repopulating abilities. These results demonstrate that the antibody/c-Mpl chimera is capable of signal transduction that mimics wild-type c-Mpl signaling. Copyright © 2011 Elsevier Ltd. All rights reserved.

COMPLEMENT FIXATION IN DISEASED TISSUES

PubMed Central

Burkholder, Peter M.

1961-01-01

An immunohistologic complement fixation test has been used in an effort to detect immune complexes in sections of kidney from rats injected with rabbit anti-rat kidney serum and in sections of biopsied kidneys from four humans with membranous glomerulonephritis. Sections of the rat and human kidneys were treated with fluorescein-conjugated anti-rabbit globulin or antihuman globulin respectively. Adjacent sections in each case were incubated first with fresh guinea pig serum and then in a second step were treated with fluorescein-conjugated antibodies against fixed guinea pig complement to detect sites of fixation of the complement. It was demonstrated that the sites of rabbit globulin in glomerular capillary walls of the rat kidneys and the sites of localized human globulin in thickened glomerular capillary walls and swollen glomerular endothelial cells of the human kidneys were the same sites in which guinea pig complement was fixed in vitro. It was concluded from these studies that rabbit nephrotoxic antibodies localize in rat glomeruli in complement-fixing antigen-antibody complexes. Furthermore, it was concluded that the deposits of human globulin in the glomeruli of the human kidneys behaved like antibody globulin in complement-fixing antigen-antibody complexes. The significance of demonstrating complement-fixing immune complexes in certain diseased tissues is discussed in regard to determination of the causative role of allergic reactions in disease. PMID:19867205

Reverse micelle-loaded lipid nano-emulsions: new technology for nano-encapsulation of hydrophilic materials.

PubMed

Anton, Nicolas; Mojzisova, Halina; Porcher, Emilien; Benoit, Jean-Pierre; Saulnier, Patrick

2010-10-15

This study presents novel, recently patented technology for encapsulating hydrophilic species in lipid nano-emulsions. The method is based on the phase-inversion temperature method (the so-called PIT method), which follows a low-energy and solvent-free process. The nano-emulsions formed are stable for months, and exhibit droplet sizes ranging from 10 to 200 nm. Hydrophilic model molecules of fluorescein sodium salt are encapsulated in the oily core of these nano-emulsion droplets through their solubilisation in the reverse micellar system. As a result, original, multi-scaled nano-objects are generated with a 'hydrophilic molecule in a reverse-micelles-in-oil-in-water' structure. Once fluorescein has been encapsulated it remains stable, for thermodynamic reasons, and the encapsulation yields can reach 90%. The reason why such complex objects can be formed is due to the soft method used (PIT method) which allows the conservation of the structure of the reverse micelles throughout the formulation process, up to their entrapment in the nano-emulsion droplets. In this study, we focus the investigation on the process itself, revealing its potential and limits. Since the formulation of nanocarriers for the encapsulation of hydrophilic substances still remains a challenge, this study may constitute a significant advance in this field. Copyright 2010 Elsevier B.V. All rights reserved.

Thermally activated delayed fluorescence of fluorescein derivative for time-resolved and confocal fluorescence imaging.

PubMed

Xiong, Xiaoqing; Song, Fengling; Wang, Jingyun; Zhang, Yukang; Xue, Yingying; Sun, Liangliang; Jiang, Na; Gao, Pan; Tian, Lu; Peng, Xiaojun

2014-07-09

Compared with fluorescence imaging utilizing fluorophores whose lifetimes are in the order of nanoseconds, time-resolved fluorescence microscopy has more advantages in monitoring target fluorescence. In this work, compound DCF-MPYM, which is based on a fluorescein derivative, showed long-lived luminescence (22.11 μs in deaerated ethanol) and was used in time-resolved fluorescence imaging in living cells. Both nanosecond time-resolved transient difference absorption spectra and time-correlated single-photon counting (TCSPC) were employed to explain the long lifetime of the compound, which is rare in pure organic fluorophores without rare earth metals and heavy atoms. A mechanism of thermally activated delayed fluorescence (TADF) that considers the long wavelength fluorescence, large Stokes shift, and long-lived triplet state of DCF-MPYM was proposed. The energy gap (ΔEST) of DCF-MPYM between the singlet and triplet state was determined to be 28.36 meV by the decay rate of DF as a function of temperature. The ΔE(ST) was small enough to allow efficient intersystem crossing (ISC) and reverse ISC, leading to efficient TADF at room temperature. The straightforward synthesis of DCF-MPYM and wide availability of its starting materials contribute to the excellent potential of the compound to replace luminescent lanthanide complexes in future time-resolved imaging technologies.

OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY SHOWS INNER CHOROIDAL ISCHEMIA IN ACUTE POSTERIOR MULTIFOCAL PLACOID PIGMENT EPITHELIOPATHY.

PubMed

Dolz-Marco, Rosa; Sarraf, David; Giovinazzo, Vincent; Freund, K Bailey

2017-01-01

To describe multimodal imaging findings of an evolving case of acute posterior multifocal placoid pigment epitheliopathy occurring in a young healthy male. Case report of a patient with acute posterior multifocal placoid pigment epitheliopathy including comprehensive systemic and ocular examinations. Ultra-widefield autofluorescence, fluorescein angiography, indocyanine green angiography, and serial optical coherence tomography angiography were performed. A 34-year-old male presented with acute vision loss in his left eye for 2 weeks. His best-corrected visual acuity was 20/20 in his right eye and 20/200 in his left eye. Dilated funduscopic examination revealed multiple creamy white deep retinal lesions showing macular involvement of the left eye with a diffuse area of pigmentary changes. The presence of multiple areas of hypoperfusion of the inner choroid were demonstrated with fluorescein and indocyanine green angiography. Serial optical coherence tomography angiography showed multiple evolving areas of decreased flow at the level of the inner choroid. Although the pathogenesis of acute posterior multifocal placoid pigment epitheliopathy remains unknown, there is growing evidence of a primary choroidal involvement with secondary damage to the overlying retinal pigment epithelium and the outer retinal layers. Optical coherence tomography angiography may provide valuable information for the diagnosis and follow-up of this condition avoiding invasive angiographic procedures.

Comparison between wide-angle OCT angiography and ultra-wide field fluorescein angiography for detecting non-perfusion areas and retinal neovascularization in eyes with diabetic retinopathy.

PubMed

Sawada, Osamu; Ichiyama, Yusuke; Obata, Syunpei; Ito, Yuka; Kakinoki, Masashi; Sawada, Tomoko; Saishin, Yoshitsugu; Ohji, Masahito

2018-04-30

To compare the ability of wide-angle optical coherence tomography angiography (OCTA) with that of ultra-wide field fluorescein angiography (UWFFA) to detect non-perfusion areas (NPAs) or retinal neovascularization (NV) in eyes with diabetic retinopathy (DR). Patients with DR underwent UWFFA using the Optos® panoramic 200Tx imaging system and wide-angle OCTA with 12 × 12 mm fields of five visual fixations using the PLEX Elite 9000®. We compared the abilities of UWFFA and OCTA to detect NPAs and NV. Fifty-eight eyes of 33 patients (mean age, 60.0 years old; female/male, 16/17) with DR were evaluated. NPAs were detected in 47 out of 58 eyes using UWFFA and in 48 eyes using OCTA. NVs were detected in 25 out of the 58 eyes using UWFFA and in 26 eyes using OCTA. The sensitivity for detection of NPA using OCTA was 0.98, and the specificity was 0.82. The sensitivity for detection of NV was 1.0, and the specificity was 0.97. The wide-angle OCTA seems to be clinically useful for the detection of NPAs or NV.

GAMMA RADIATION OF EYE MEDICAMENTS

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

Ogg, A.J.

1963-07-13

The use of x rays to sterilize ophthalmological pharmaceutical preparations is described, particularly penicillin and fluorescein drops. The method used with penicillin is to place a tablet (1500 units) in a small vial which is closed with a cork and attached with Sellotape to a 5-ml eye-drop bottle containing 3 ml of normal saline. The bottles were sealed in two polythene envelopes and irradiated at the Wantage Research Lab (AERE) at a dose of 2.5 Mrad. The bags are then stored in a refrigerator at 4 deg C. When required for operation, the outer bag is opened with sterile scissors,more » and the contents tipped onto the surface of the sterile instrument tray. The dropper bulb is filled with saline and introduced into the vial, thereby dissolving the tablet of penicillin. All irradiated preparations were found to be absolutely sterile and fully potent. In the case of fluorescein, 1-in. lengths of gauge 2-intraarterial polythene tubes are filled with the dye from a syringe, and the ends of the tube sealed in a flame. Several hundreds of tubes are irradiated in a single container, and when required, one end of the tube is cut off and a drop squeezed out into the conjunctival sac. (BBB)« less

Discrepancy between fluorescein angiography and optical coherence tomography in detection of macular disease.

PubMed

Kozak, Igor; Morrison, Victoria L; Clark, Thomas M; Bartsch, Dirk-Uwe; Lee, Byung Ro; Falkenstein, Iryna; Tammewar, Ajay M; Mojana, Francesca; Freeman, William R

2008-04-01

To compare high-resolution optical coherence tomography (OCT) and fluorescein angiography (FA) in detection of macular edema (ME) of various etiologies. In a retrospective study over a 12-month period at one retina center, data for consecutive eyes that had undergone simultaneous conventional FA (HRA; Heidelberg Engineering, Vista, CA) and StratusOCT (Carl Zeiss Meditec, Dublin, CA) to rule out ME were reviewed. A subset of patients underwent additional examination with extremely high-resolution (6-microm)/ultrahigh-speed spectral OCT/scanning laser ophthalmoscopy (OTI, Inc., Toronto, Ontario, Canada). Of 1,272 eyes, 1,208 (94.97%) had the finding of ME or subretinal fluid confirmed by both techniques. There were 49 eyes (3.86%) for which FA showed dye leakage in the macular area and OCT showed normal foveal contour. Of 10 eyes in this group that underwent imaging with ultrahigh-speed spectral OCT/scanning laser ophthalmoscopy, 8 had subtle diffuse lucencies in the retina. For 15 eyes (1.17%), OCT showed intraretinal and subretinal fluid, which was missed by FA. Both FA and high-resolution OCT are highly sensitive techniques and correlate well in detection of ME. However, there is a small chance that when performed alone they might miss existing subtle ME.

Growth control of genetically modified cells using an antibody/c-Kit chimera.

PubMed

Kaneko, Etsuji; Kawahara, Masahiro; Ueda, Hiroshi; Nagamune, Teruyuki

2012-05-01

Gene therapy has been regarded as an innovative potential treatment against serious congenital diseases. However, applications of gene therapy remain limited, partly because its clinical success depends on therapeutic gene-transduced cells acquiring a proliferative advantage. To address this problem, we have developed the antigen-mediated genetically modified cell amplification (AMEGA) system, which uses chimeric receptors to enable the selective proliferation of gene-transduced cells. In this report, we describe mimicry of c-Kit signaling and its application to the AMEGA system. We created an antibody/c-Kit chimera in which the extracellular domain of c-Kit is replaced with an anti-fluorescein single-chain Fv antibody fragment and the extracellular D2 domain of the erythropoietin receptor. A genetically modified mouse pro-B cell line carrying this chimera showed selective expansion in the presence of fluorescein-conjugated BSA (BSA-FL) as a growth inducer. By further engineering the transmembrane domain of the chimera to reduce interchain interaction we attained stricter ligand-dependency. Since c-Kit is an important molecule in the expansion of hematopoietic stem cells (HSCs), this antibody/c-Kit chimera could be a promising tool for gene therapy targeting HSCs. Copyright © 2011 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Real-world assessment of diquafosol in dry eye patients with risk factors such as contact lens, meibomian gland dysfunction, and conjunctivochalasis: subgroup analysis from a prospective observational study

PubMed Central

Yamaguchi, Masahiko; Nishijima, Takeshi; Shimazaki, Jun; Takamura, Etsuko; Yokoi, Norihiko; Watanabe, Hitoshi; Ohashi, Yuichi

2015-01-01

Purpose To evaluate the efficacy and safety of diquafosol (DQS) ophthalmic solution in dry eye (DE) patients wearing contact lenses (CLs) or with concomitant meibomian gland dysfunction (MGD) or conjunctivochalasis in a real-world setting. Patients and methods From a cohort of patients enrolled in a prospective observational study, DE patients who met the Japanese diagnostic criteria and who received DQS as a monotherapy were extracted and stratified according to the presence or absence of CL use, MGD, and conjunctivochalasis. Corneal and conjunctival fluorescein staining score, tear film break-up time, total symptom score (12 DE-related subjective symptoms), patient-reported outcomes, and adverse reactions were investigated. Results DQS treatment resulted in significant improvement in total symptom score, corneal and conjunctival fluorescein staining score, and tear film break-up time without significant differences between patient subgroups with versus without CL use, MGD, or conjunctivochalasis. Comparable proportions of patients perceived symptomatic improvements in all subgroups. There were no adverse reactions specifically associated with the CL use or any comorbidity of MGD or conjunctivochalasis. Conclusion DQS can be used effectively and safely as a monotherapy for the treatment of DE patients wearing CLs or with concomitant MGD or conjunctivochalasis. PMID:26664039

Skull Base Meningiomas and Cranial Nerves Contrast Using Sodium Fluorescein: A New Application of an Old Tool

PubMed Central

da Silva, Carlos Eduardo; da Silva, Vinicius Duval; da Silva, Jefferson Luis Braga

2014-01-01

Objective The identification of cranial nerves is one of the most challenging goals in the dissection of skull base meningiomas. The authors present an application of sodium fluorescein (SF) in skull base meningiomas with the purpose of improving the identification of cranial nerves. Design A prospective study within-subjects design. Setting Hospital Ernesto Dornelles, Porto Alegre, Brazil. Participants Patients with skull base meningiomas. Main Outcomes Measures Cranial nerve identification. Results The group of nine meningiomas was composed of one cavernous sinus, three petroclival, one tuberculum sellae, two sphenoid wing, one olfactory groove, and one temporal floor meningioma. The SF enhancement in all tumors was strong, and the contrast with cranial nerves clearly evident. There were one definite olfactory nerve deficit, one transient abducens deficit, and one definite hemiparesis. All lesions were resected (Simpson grades 1 and 2). The analysis of the difference of the delta SF wavelength between the meningiomas and cranial nerve contrast was performed by the Wilcoxon signed rank test and showed p = 0.011. Conclusions The contrast between the enhanced meningiomas and cranial nerves was evident and assisted in the visualization and microsurgical dissection of these structures. The anatomical preservation of these structures was improved using the contrast. PMID:27054056

Skull Base Meningiomas and Cranial Nerves Contrast Using Sodium Fluorescein: A New Application of an Old Tool.

PubMed

da Silva, Carlos Eduardo; da Silva, Vinicius Duval; da Silva, Jefferson Luis Braga

2014-08-01

Objective The identification of cranial nerves is one of the most challenging goals in the dissection of skull base meningiomas. The authors present an application of sodium fluorescein (SF) in skull base meningiomas with the purpose of improving the identification of cranial nerves. Design A prospective study within-subjects design. Setting Hospital Ernesto Dornelles, Porto Alegre, Brazil. Participants Patients with skull base meningiomas. Main Outcomes Measures Cranial nerve identification. Results The group of nine meningiomas was composed of one cavernous sinus, three petroclival, one tuberculum sellae, two sphenoid wing, one olfactory groove, and one temporal floor meningioma. The SF enhancement in all tumors was strong, and the contrast with cranial nerves clearly evident. There were one definite olfactory nerve deficit, one transient abducens deficit, and one definite hemiparesis. All lesions were resected (Simpson grades 1 and 2). The analysis of the difference of the delta SF wavelength between the meningiomas and cranial nerve contrast was performed by the Wilcoxon signed rank test and showed p = 0.011. Conclusions The contrast between the enhanced meningiomas and cranial nerves was evident and assisted in the visualization and microsurgical dissection of these structures. The anatomical preservation of these structures was improved using the contrast.

Fluorescence-Doped Particles for Simultaneous Temperature and Velocity Imaging

NASA Technical Reports Server (NTRS)

Danehy, Paul M.; Tiemsin, Pacita I.; Wohl, Chrostopher J.; Verkamp, Max; Lowe, T.; Maisto, P.; Byun, G.; Simpson, R.

2012-01-01

Polystyrene latex microspheres (PSLs) have been used for particle image velocimetry (PIV) and laser Doppler velocimetry (LDV) measurements for several decades. With advances in laser technologies, instrumentation, and data processing, the capability to collect more information about fluid flow beyond velocity is possible using new seed materials. To provide additional measurement capability, PSLs were synthesized with temperature-sensitive fluorescent dyes incorporated within the particle. These multifunctional PSLs would have the greatest impact if they could be used in large scale facilities with minimal modification to the facilities or the existing instrumentation. Consequently, several potential dyes were identified that were amenable to existing laser systems currently utilized in wind tunnels at NASA Langley Research Center as well as other wind and fluid (water) tunnels. PSLs incorporated with Rhodamine B, dichlorofluorescein (DCF, also known as fluorescein 548 or fluorescein 27) and other dyes were synthesized and characterized for morphology and spectral properties. The resulting particles were demonstrated to exhibit fluorescent emission, which would enable determination of both fluid velocity and temperature. They also would allow near-wall velocity measurements whereas laser scatter from surfaces currently prevents near-wall measurements using undoped seed materials. Preliminary results in a wind tunnel facility located at Virginia Polytechnic Institute and State University (Virginia Tech) have verified fluorescent signal detection and temperature sensitivity of fluorophore-doped PSLs.

Altered expression of zonula occludens-2 precedes increased blood-brain barrier permeability in a murine model of fulminant hepatic failure.

PubMed

Shimojima, Naoki; Eckman, Christopher B; McKinney, Michael; Sevlever, Daniel; Yamamoto, Satoshi; Lin, Wenlang; Dickson, Dennis W; Nguyen, Justin H

2008-01-01

Brain edema secondary to increased blood-brain barrier (BBB) permeability is a lethal complication in fulminant hepatic failure (FHF). Intact tight junctions (TJ) between brain capillary endothelial cells are critical for normal BBB function. However, the role of TJ in FHF has not been explored. We hypothesized that alterations in the composition of TJ proteins would result in increased BBB permeability in FHF. In this study, FHF was induced in C57BL/6J mice by using azoxymethane. BBB permeability was assessed with sodium fluorescein. Expression of TJ proteins was determined by Western blot, and their cellular distribution was examined using immunofluorescent microscopy. Comatose FHF mice had significant cerebral sodium fluorescein extravasation compared with control and precoma FHF mice, indicating increased BBB permeability. Western blot analysis showed a significant decrease in zonula occludens (ZO)-2 expression starting in the precoma stage. Immunofluorescent microscopy showed a significantly altered distribution pattern of ZO-2 in isolated microvessels from precoma FHF mice. These changes were more prominent in comatose FHF animals. Significant alterations in ZO-2 expression and distribution in the tight junctions preceded the increased BBB permeability in FHF mice. These results suggest that ZO-2 may play an important role in the pathogenesis of brain edema in FHF.

DETECTION OF NONEXUDATIVE CHOROIDAL NEOVASCULARIZATION IN AGE-RELATED MACULAR DEGENERATION WITH OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY.

PubMed

Palejwala, Neal V; Jia, Yali; Gao, Simon S; Liu, Liang; Flaxel, Christina J; Hwang, Thomas S; Lauer, Andreas K; Wilson, David J; Huang, David; Bailey, Steven T

2015-11-01

To evaluate eyes with age-related macular degeneration and high-risk characteristics for choroidal neovascularization (CNV) with optical coherence tomographic (OCT) angiography to determine whether earlier detection of CNV is possible. Eyes with drusen, pigmentary changes, and with CNV in the fellow eye were scanned with a 70-kHz spectral domain OCT system (Optovue RTVue-XR Avanti). The split-spectrum amplitude-decorrelation angiography (SSADA) algorithm was used to distinguish blood flow from static tissue. Two masked graders reviewed scans for CNV, defined as flow in the outer retinal/sub-RPE slab. Choroidal neovascularization flow area repeatability and between-grader reproducibility were calculated. Of 32 eyes, 2 (6%) were found to have Type 1 CNV with OCT angiography. The lesions were not associated with leakage on fluorescein angiography or fluid on OCT. One case was followed for 8 months without treatment, and the CNV flow area enlarged slightly without fluid buildup on OCT or vision loss. Between-grader reproducibility of the CNV flow area was 9.4% (coefficient of variation) and within-visit repeatability was 5.2% (pooled coefficient of variation). Optical coherence tomographic angiography can detect the presence of nonexudative CNV, lesions difficult to identify with fluorescein angiography and OCT. Further study is needed to understand the significance and natural history of these lesions.

Prevalidation of a human cornea construct as an alternative to animal corneas for in vitro drug absorption studies.

PubMed

Hahne, Matthias; Zorn-Kruppa, Michaela; Guzman, Gustavo; Brandner, Johanna M; Haltner-Ukomado, Eleonore; Wätzig, Hermann; Reichl, Stephan

2012-08-01

The use of ophthalmic drugs has increased consistently over the past few decades. Currently, most research is conducted using in vivo and ex vivo animal experiments; however, they have many disadvantages, including ethical concerns, high costs, the questionable extension of animal results to humans, and poor standardization. Although several cell culture-based cornea models have been developed, none have been validated and accepted for general use. In this study, a standardized, three-dimensional model of the human cornea (Hemicornea, HC) based on immortalized human corneal cells and cultivated in serum-free conditions was developed for drug absorption studies and prevalidated using compounds with a wide range of molecular characteristics (sodium fluorescein, rhodamine B, fluorescein isothiocyanate-labeled dextran, aciclovir, bimatoprost, dexamethasone, and timolol maleate). The HC model was independently cultured in three different laboratories, and the intralaboratory and interlaboratory reproducibility was analyzed and compared with the rabbit cornea. This analysis showed that the HC has a barrier in the same range as excised animal corneas, although with a higher reproducibility and lower variability. Because of the demonstrated transferability, the HC represents a promising in vitro alternative to the use of ex vivo tissue and offers a well-defined and standardized system for drug absorption studies. Copyright © 2012 Wiley Periodicals, Inc.

Development of a convenient ex vivo model for the study of the transcorneal permeation of drugs: histological and permeability evaluation.

PubMed

Pescina, Silvia; Govoni, Paolo; Potenza, Arianna; Padula, Cristina; Santi, Patrizia; Nicoli, Sara

2015-01-01

In this paper, an ex vivo model for the study of the transcorneal permeation of drugs, based on porcine tissues, was evaluated. The setup is characterized by ease of realization, absence of O₂ and CO₂ bubbling and low cost; additionally, the large availability of porcine tissue permits a high throughput. Histological images showed the comparability between porcine and human corneas and confirmed the effectiveness of the isolation procedure. A new de-epithelization procedure based on a thermal approach was also set up to simulate cornea permeability in pathological conditions. The procedure did not affect the integrity of the underlying layers and allowed the characterization of the barrier properties of epithelium and stroma. Six compounds with different physicochemical properties were tested: fluorescein, atenolol, propranolol, diclofenac, ganciclovir and lidocaine. The model highlighted the barrier function played by epithelium toward the diffusion of hydrophilic compounds and the permselectivity with regard to more lipophilic molecules. In particular, positively charged compounds showed a significantly higher transcorneal permeability than negatively charged compounds. The comparability of results with literature data supports the goodness and the robustness of the model, especially taking into account the behavior of fluorescein, which is generally considered a marker of tissue integrity. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.

A reduced graphene oxide-based fluorescence resonance energy transfer sensor for highly sensitive detection of matrix metalloproteinase 2.

PubMed

Xi, Gaina; Wang, Xiaoping; Chen, Tongsheng

2016-01-01

A novel fluorescence nanoprobe (reduced nano-graphene oxide [nrGO]/fluorescein isothiocyanate-labeled peptide [Pep-FITC]) for ultrasensitive detection of matrix metalloproteinase 2 (MMP2) has been developed by engineering the Pep-FITC comprising the specific MMP2 substrate domain (PLGVR) onto the surface of nrGO particles through non-covalent linkage. The nrGO was obtained by water bathing nano-graphene oxide under 90°C for 4 hours. After mixing the nrGO and Pep-FITC for 30 seconds, the fluorescence from Pep-FITC was almost completely quenched due to the fluorescence resonance energy transfer between fluorescein isothiocyanate (FITC) and nrGO. Upon cleavage of the amide bond between Leu and Gly in the Pep-FITC by protease-MMP2, the FITC bound to nrGO was separated from nrGO surface, disrupting the fluorescence resonance energy transfer process and resulting in fluorescence recovery of FITC. Under optimal conditions, the fluorescence recovery of nrGO/Pep-FITC was found to be directly proportional to the concentration of MMP2 within 0.02-0.1 nM. The detection limit of the nrGO/Pep-FITC was determined to be 3 pM, which is approximately tenfold lower than that of the unreduced carboxylated nano-graphene oxide/Pep-FITC probe.

Influence of the target molecule on the oxygen radical absorbance capacity index: a comparison between alizarin red- and fluorescein-based methodologies.

PubMed

Martin, I; Aspée, A; Torres, P; Lissi, E; López-Alarcón, C

2009-12-01

A comparison of alizarin red (AR) and fluorescein (FL) as target molecules in oxygen radical absorbance capacity (ORAC)-like methods is reported. Galangin, apigenin, ferulic acid, and coumaric acid decreased AR initial consumption rate, whereas quercetin, kaempferol, luteolin, caffeic acid, and sinapic acid inhibited its consumption through an induction time, associated with a repair mechanism. On the other hand, all compounds protected FL with a clear induction time. AR was more selective and provides ORAC-AR values considerably smaller for compounds of low reactivity. The ORAC-AR value for luteolin was nearly 200 times that of coumaric acid. However, the ratio of ORAC-FL values for luteolin and coumaric acid was only 1.2. This different selectivity implies that AR provides ORAC values more related to reactivity than FL. ORAC-AR values of infusions were considerably smaller than the corresponding ORAC-FL values. These differences are interpreted in terms of the capacity of FL to generate induction times, irrespective of the reactivity of the additive. It is proposed that comparison of ORAC-AR and ORAC-FL values could afford a rough estimation of the average reactivity of the antioxidants titrated by the ORAC-FL methodology.

Extending applicability of the oxygen radical absorbance capacity (ORAC-fluorescein) assay.

PubMed

Dávalos, Alberto; Gómez-Cordovés, Carmen; Bartolomé, Begoña

2004-01-14

The ORAC-fluorescein (ORAC-FL) method recently validated using automatic liquid handling systems has now been adapted to manual handling and using a conventional fluorescence microplate reader. As calculated for Trolox, the precision of the method was <3.0, expressed as percent coefficient of variation. The accuracy of the method was <2.3, expressed as percent variation of the mean. The detection and quantification limits were those corresponding to 0.5- and 1-microM Trolox standard solutions, respectively. The method has been applied to 10 pure compounds (benzoic and cinnamic acids and aldehydes, flavonoids, and butylated hydroxyanisole), to 30 white, rose, and bottled- and oak-aged red wines, and to 7 commercial dietary antioxidant supplements. All samples exhibited a good linear response with concentration. As seen by other methodologies, the chemical structure of a compound determines its antioxidant activity (ORAC-FL value). Of particular interest were the results with oak-aged red wines from different vintages (1989-2002) that confirm influence of vintage, but not origin of the oak, in the antioxidant activity of wines from the same variety. Dietary antioxidant supplements presented a great variability (170-fold difference) in their antioxidant potency. This work proves applicability of the ORAC-FL assay in evaluating the antioxidant activity of diverse food samples.

[Determination of H2S in Rat Intestinal Perfusion Solution Based on Fluorescence Analysis].

PubMed

Hou, Jun-feng; Li, Xin-xia; Shen, Xue-ru; Huojia, Miliban; Guan, Ming

2015-08-01

Under alkaline conditions, Fluorescein mercury has strong fluorescence, however, when it met S(2-), its fluorescence would quench, in view of the above, a fluorescence method for determination of H2S in biological samples was established. In the 0.1 mol · L(-1) NaOH dilution, when the concentration of fluorescein Mercury and Na2S was 5.0 × 10(-5) and 1.0 × 10(-5) mol · L(-1) respectively, the fluorescence intensity of system was determined at 522 nm. The results showed that, at the range of 4.0 × 10(-7)~2.0 × 10(-6) mol · L(-1), the concentration decreasing of H2S and fluorescence intensity had good linear relationship, r=0.9980, the RSD of precision test was 4.59% (n=7), the detection limit was 3.5 × 10(-8) mol · L(-1), the content of H2S in the sample were 1.01 × 10(-6) and 1.15 × 10(-6) mol · L(-1), and the recovery rate was 95.8%~101.0%, the method has the advantages of simple operation, high sensitivity, good selectivity, can accurately determine of H2S in intestinal perfused solution, and provides the basis for the determination of endogenous H2S.

Successful application of the dual-vector system II in creating a reliable phage-displayed combinatorial Fab library.

PubMed

Song, Suk-yoon; Hur, Byung-ung; Lee, Kyung-woo; Choi, Hyo-jung; Kim, Sung-soo; Kang, Goo; Cha, Sang-hoon

2009-03-31

The dual-vector system-II (DVS-II), which allows efficient display of Fab antibodies on phage, has been reported previously, but its practical applicability in a phage-displayed antibody library has not been verified. To resolve this issue, we created two small combinatorial human Fab antibody libraries using the DVS-II, and isolation of target-specific antibodies was attempted. Biopanning of one antibody library, termed DVFAB-1L library, which has a 1.3 x 10(7) combinatorial antibody complexity, against fluorescein-BSA resulted in successful isolation of human Fab clones specific for the antigen despite the presence of only a single light chain in the library. By using the unique feature of the DVS-II, an antibody library of a larger size, named DVFAB-131L, which has a 1.5 x 10(9) combinatorial antibody complexity, was also generated in a rapid manner by combining 1.3 x 10(7) heavy chains and 131 light chains and more diverse anti-fluorescein-BSA Fab antibody clones were successfully obtained. Our results demonstrate that the DVS-II can be applied readily in creating phage-displayed antibody libraries with much less effort, and target-specific antibody clones can be isolated reliably via light chain promiscuity of antibody molecule.

Optimizing Fluorescein Isothiocyanate Dextran Measurement As a Biomarker in a 24-h Feed Restriction Model to Induce Gut Permeability in Broiler Chickens

PubMed Central

Baxter, Mikayla F. A.; Merino-Guzman, Ruben; Latorre, Juan D.; Mahaffey, Brittany D.; Yang, Yichao; Teague, Kyle D.; Graham, Lucas E.; Wolfenden, Amanda D.; Hernandez-Velasco, Xochitl; Bielke, Lisa R.; Hargis, Billy M.; Tellez, Guillermo

2017-01-01

Fluorescein isothiocyanate dextran (FITC-d) is a 3–5 kDa marker used to measure tight junction permeability. We have previously shown that intestinal barrier function can be adversely affected by stress, poorly digested diets, or feed restriction (FR), resulting in increased intestinal inflammation-associated permeability. However, further optimization adjustments of the current FITC-d methodology are possible to enhance precision and efficacy of results in future. The objective of the present study was to optimize our current model to obtain a larger difference between control and treated groups, by optimizing the FITC-d measurement as a biomarker in a 24-h FR model to induce gut permeability in broiler chickens. One in vitro and four in vivo independent experiments were conducted. The results of the present study suggest that by increasing the dose of FITC-d (8.32 versus 4.16 mg/kg); shortening the collection time of blood samples (1 versus 2.5 h); using a pool of non-FITC-d serum as a blank, compared to previously used PBS; adding a standard curve to set a limit of detection and modifying the software’s optimal sensitivity value, it was possible to obtain more consistent and reliable results. PMID:28470003

Vaginal delivery of carboplatin-loaded thermosensitive hydrogel to prevent local cervical cancer recurrence in mice.

PubMed

Wang, Xue; Wang, Jin; Wu, Wenbin; Li, Hongjun

2016-11-01

Local tumor recurrence after cervical cancer surgery remains a clinical problem. Vaginal delivery of thermosensitive hydrogel may be suited to reduce tumor relapse rate with more efficacy and safety. A pilot study was carried out to evaluate the efficacy of carboplatin-loaded poloxamer hydrogel to prevent local recurrence of cervical cancer after surgery. In vivo vaginal retention evaluation of 27% poloxamer hydrogel in mice was proven to be a suitable vaginal drug delivery formulation due to its low gelation temperature. A mimic orthotopic cervical/vaginal cancer recurrence model after surgery was established by injecting murine cervical cancer cell line U14 into the vaginal submucosa to simulate the residual tumor cells infiltrated in the surgical site, followed by drug administration 24 h later to interfere with the formation/recurrence of the tumor. By infusing fluorescein sodium-loaded hydrogel into the vagina of mice, a maximized accumulation of fluorescein sodium (Flu) in the vagina was achieved and few signals were observed in other organs. When used in the prevention of the cervical cancer formation/recurrence in mice, the carboplatin-loaded poloxamer hydrogel exhibited great efficacy and systemic safety. In conclusion, thermosensitive hydrogel presents a simple, practical approach for the local drug delivery via vagina against cervical cancer recurrence.

Five essential oils from aromatic plants of Cameroon: their antibacterial activity and ability to permeabilize the cytoplasmic membrane of Listeria innocua examined by flow cytometry.

PubMed

Nguefack, J; Budde, B B; Jakobsen, M

2004-01-01

To investigate the antibacterial effect of five essential oils (EO) extracted from aromatic plants (Cymbopogon citratus, Ocimumbasilicum, Ocimum gratissimum, Thymus vulgaris and Zingiber officinale) of Cameroon against strains of Listeria monocytogenes, L. innocua and Staphylococcus aureus. The ability of selected EO to permeabilize the cytoplasmic membrane of L. innocua was also examined. The antibacterial activity of the EO determined by the agar diffusion method showed that T. vulgaris had the highest activity followed by O. gratissimum and C. citratus. Lowest activity was recorded from Z. officinale and O. basilicum. Significant differences in sensitivity between strains of Listeria and S. aureus were observed. Flow cytometry of L. innocua stained with carboxy-fluorescein diacetate showed that the fluorescence intensity of cells exposed to EO decreased faster than nonexposed cells, indicating that EO permeabilized the cytoplasmic membrane with the leakage of carboxy-fluorescein. Almost all the EO tested showed antibacterial activity to a different extent. The antibacterial effect was due to permeabilization of the cytoplasmic membrane. This study has identified the preservative potential of the EO examined. The use of sensitive method, such as flow cytometry, is advantageous for quick generation of data on the antibacterial effect of EO.

Endoscopic endonasal dacryocystorhinostomy combined with canaliculus repair for the management of dacryocystitis with canalicular obstruction.

PubMed

Tu, Yunhai; Qian, Zhenbin; Zhang, Jiao; Wu, Wencan; Xiao, Tianlin

2015-01-01

Purpose. The aim of this study is to propose a simple and efficient combination surgery for the management of dacryocystitis with canalicular obstruction. Methods. A retrospective noncomparative case series of dacryocystitis with canalicular obstruction has been studied. Twelve patients with dacryocystitis and canalicular obstruction underwent a conventional endoscopic endonasal dacryocystorhinostomy (EE-DCR) combined with a modified canalicular repair. Postoperative observations included slit lamp, fluorescein dye disappearance test, lacrimal syringing, lacrimal endoscopy, and nasal endoscopy. Results. After 6-18 months of postoperative follow-up, the symptoms of epiphora and mucopurulent discharge disappeared completely in 10 patients, and occasional or intermittent epiphora remained in 2 patients. All of the twelve patients showed an opened intranasal ostium and normal fluorescein dye disappearance test. Patent bicanalicular irrigation was achieved in 9 patients. One patient had a partial and the other two had a complete reobstruction by lacrimal irrigation to their repaired lower canaliculus; however, all of them had a patent lacrimal irrigation to upper canaliculus. The functional success rate for the combination surgery is 83% (10/12), and anatomical success rate is 75% (9/12). Conclusion. EE-DCR combined with modified canalicular repair is a simple and efficient method for the management of dacryocystitis with canalicular obstruction.

Acetate but not propionate induces oxidative stress in bakers' yeast Saccharomyces cerevisiae.

PubMed

Semchyshyn, Halyna M; Abrat, Oleksandra B; Miedzobrodzki, Jacek; Inoue, Yoshiharu; Lushchak, Volodymyr I

2011-01-01

The influence of acetic and propionic acids on baker's yeast was investigated in order to expand our understanding of the effect of weak organic acid food preservatives on eukaryotic cells. Both acids decreased yeast survival in a concentration-dependent manner, but with different efficiencies. The acids inhibited the fluorescein efflux from yeast cells. The inhibition constant of fluorescein extrusion from cells treated with acetate was significantly lower in parental strain than in either PDR12 (ABC-transporter Pdr12p) or WAR1 (transcriptional factor of Pdr12p) defective mutants. The constants of inhibition by propionate were virtually the same in all strains used. Yeast exposure to acetate increased the level of oxidized proteins and the activity of antioxidant enzymes, while propionate did not change these parameters. This suggests that various mechanisms underlie the yeast toxicity by acetic and propionic acids. Our studies with mutant cells clearly indicated the involvement of Yap1p transcriptional regulator and de novo protein synthesis in superoxide dismutase up-regulation by acetate. The up-regulation of catalase was Yap1p independent. Yeast pre-incubation with low concentrations of H₂O₂ caused cellular cross-protection against high concentrations of acetate. The results are discussed from the point of view that acetate induces a prooxidant effect in vivo, whereas propionate does not.

Evaluation of Pump Pulsation in Respirable Size-Selective Sampling: Part II. Changes in Sampling Efficiency

PubMed Central

Lee, Eun Gyung; Lee, Taekhee; Kim, Seung Won; Lee, Larry; Flemmer, Michael M.; Harper, Martin

2015-01-01

This second, and concluding, part of this study evaluated changes in sampling efficiency of respirable size-selective samplers due to air pulsations generated by the selected personal sampling pumps characterized in Part I (Lee E, Lee L, Möhlmann C et al. Evaluation of pump pulsation in respirable size-selective sampling: Part I. Pulsation measurements. Ann Occup Hyg 2013). Nine particle sizes of monodisperse ammonium fluorescein (from 1 to 9 μm mass median aerodynamic diameter) were generated individually by a vibrating orifice aerosol generator from dilute solutions of fluorescein in aqueous ammonia and then injected into an environmental chamber. To collect these particles, 10-mm nylon cyclones, also known as Dorr-Oliver (DO) cyclones, were used with five medium volumetric flow rate pumps. Those were the Apex IS, HFS513, GilAir5, Elite5, and Basic5 pumps, which were found in Part I to generate pulsations of 5% (the lowest), 25%, 30%, 56%, and 70% (the highest), respectively. GK2.69 cyclones were used with the Legacy [pump pulsation (PP) = 15%] and Elite12 (PP = 41%) pumps for collection at high flows. The DO cyclone was also used to evaluate changes in sampling efficiency due to pulse shape. The HFS513 pump, which generates a more complex pulse shape, was compared to a single sine wave fluctuation generated by a piston. The luminescent intensity of the fluorescein extracted from each sample was measured with a luminescence spectrometer. Sampling efficiencies were obtained by dividing the intensity of the fluorescein extracted from the filter placed in a cyclone with the intensity obtained from the filter used with a sharp-edged reference sampler. Then, sampling efficiency curves were generated using a sigmoid function with three parameters and each sampling efficiency curve was compared to that of the reference cyclone by constructing bias maps. In general, no change in sampling efficiency (bias under ±10%) was observed until pulsations exceeded 25% for the DO cyclone. However, for three models of pumps producing 30%, 56%, and 70% pulsations, substantial changes were confirmed. The GK2.69 cyclone showed a similar pattern to that of the DO cyclone, i.e. no change in sampling efficiency for the Legacy producing 15% pulsation and a substantial change for the Elite12 producing 41% pulsation. Pulse shape did not cause any change in sampling efficiency when compared to the single sine wave. The findings suggest that 25% pulsation at the inlet of the cyclone as measured by this test can be acceptable for the respirable particle collection. If this test is used in place of that currently in European standards (EN 1232–1997 and EN 12919-1999) or is used in any International Organization for Standardization standard, then a 25% pulsation criterion could be adopted. This work suggests that a 10% criterion as currently specified in the European standards for testing may be overly restrictive and not able to be met by many pumps on the market. Further work is recommended to determine which criterion would be applicable to this test if it is to be retained in its current form. PMID:24064963

Evaluation of pump pulsation in respirable size-selective sampling: part II. Changes in sampling efficiency.

PubMed

Lee, Eun Gyung; Lee, Taekhee; Kim, Seung Won; Lee, Larry; Flemmer, Michael M; Harper, Martin

2014-01-01

This second, and concluding, part of this study evaluated changes in sampling efficiency of respirable size-selective samplers due to air pulsations generated by the selected personal sampling pumps characterized in Part I (Lee E, Lee L, Möhlmann C et al. Evaluation of pump pulsation in respirable size-selective sampling: Part I. Pulsation measurements. Ann Occup Hyg 2013). Nine particle sizes of monodisperse ammonium fluorescein (from 1 to 9 μm mass median aerodynamic diameter) were generated individually by a vibrating orifice aerosol generator from dilute solutions of fluorescein in aqueous ammonia and then injected into an environmental chamber. To collect these particles, 10-mm nylon cyclones, also known as Dorr-Oliver (DO) cyclones, were used with five medium volumetric flow rate pumps. Those were the Apex IS, HFS513, GilAir5, Elite5, and Basic5 pumps, which were found in Part I to generate pulsations of 5% (the lowest), 25%, 30%, 56%, and 70% (the highest), respectively. GK2.69 cyclones were used with the Legacy [pump pulsation (PP) = 15%] and Elite12 (PP = 41%) pumps for collection at high flows. The DO cyclone was also used to evaluate changes in sampling efficiency due to pulse shape. The HFS513 pump, which generates a more complex pulse shape, was compared to a single sine wave fluctuation generated by a piston. The luminescent intensity of the fluorescein extracted from each sample was measured with a luminescence spectrometer. Sampling efficiencies were obtained by dividing the intensity of the fluorescein extracted from the filter placed in a cyclone with the intensity obtained from the filter used with a sharp-edged reference sampler. Then, sampling efficiency curves were generated using a sigmoid function with three parameters and each sampling efficiency curve was compared to that of the reference cyclone by constructing bias maps. In general, no change in sampling efficiency (bias under ±10%) was observed until pulsations exceeded 25% for the DO cyclone. However, for three models of pumps producing 30%, 56%, and 70% pulsations, substantial changes were confirmed. The GK2.69 cyclone showed a similar pattern to that of the DO cyclone, i.e. no change in sampling efficiency for the Legacy producing 15% pulsation and a substantial change for the Elite12 producing 41% pulsation. Pulse shape did not cause any change in sampling efficiency when compared to the single sine wave. The findings suggest that 25% pulsation at the inlet of the cyclone as measured by this test can be acceptable for the respirable particle collection. If this test is used in place of that currently in European standards (EN 1232-1997 and EN 12919-1999) or is used in any International Organization for Standardization standard, then a 25% pulsation criterion could be adopted. This work suggests that a 10% criterion as currently specified in the European standards for testing may be overly restrictive and not able to be met by many pumps on the market. Further work is recommended to determine which criterion would be applicable to this test if it is to be retained in its current form.

Vascular Patterns and Perfusion of Mucogingival Tissues and their Relation to Periodontal Flap Design

DTIC Science & Technology

1987-05-01

flaps were mosL vulnerable to necrosis . Sutures placed with minimal tension did not adversely affect blood perfusion of surgically replaced flaps. vi * C...perfusion change with narrow flaps most severely affected. In general, narrow thin flaps were most vulnerable to necrosis . Sutures placed with minimal...Day Narrow Pedicle ............. 75 B. Fluorescein Angiography of Envelope Flap Immediately Post Surgery and Necrosis of Marginal Third of Six

Preventing Vision Loss from Blast Injuries with Regenerative Biomaterial

DTIC Science & Technology

2011-12-01

photorefractive keratectomy ( PRK ) procedure, which can be used to monitor epithelial healing rates. It was shown that the curved silk film bandage could...dissolution time on epithelial regeneration The PRK injury model was utilized to assess the rabbit cornea’s initial response to the application of the silk...dissolution time as compared to trial 1 within water. The PRK debridement procedure was performed and tracked over time with fluorescein staining. The films

Rapid method for identification and enumeration of oral Actinomyces.

PubMed Central

Marucha, P T; Keyes, P H; Wittenberger, C L; London, J

1978-01-01

Serotype-specific antisera prepared against whole cells of Actinomyces viscosus, A. naeslundii, and A. israeli were labeled with fluorescein dye and used to detect and quantitate antigenically related microorganisms in human dental plaque. By relating the DNA content of the dental plaque microflora to the number of Actinomyces present in the plaque samples, a reproducible method was developed for specifically enumerating five serotypic representatives of this genus found in human plaque. PMID:711333

Studies of Altered Response to Infection Induced by Thermal Injury.

DTIC Science & Technology

1984-05-01

beginning to examine various prophylactic treatments for their efficacy in reversing a number of burn mediated immune defects. We expect even more interesting...with fluoresceinated antibody (OKM5) and FACS sorting or negative selection by Sephadex G-l0 passage and/or antibody and -3- complement treatment ... infectious episode. This mitogen hyperimmunity is typical of a normal immune system dealing with an infectious challenge. The Group II patients have a

Fluorescein angiography versus optical coherence tomography for diagnosis of uveitic macular edema.

PubMed

Kempen, John H; Sugar, Elizabeth A; Jaffe, Glenn J; Acharya, Nisha R; Dunn, James P; Elner, Susan G; Lightman, Susan L; Thorne, Jennifer E; Vitale, Albert T; Altaweel, Michael M

2013-09-01

To evaluate agreement between fluorescein angiography (FA) and optical coherence tomography (OCT) results for diagnosis of macular edema in patients with uveitis. Multicenter cross-sectional study. Four hundred seventy-nine eyes with uveitis from 255 patients. The macular status of dilated eyes with intermediate uveitis, posterior uveitis, or panuveitis was assessed via Stratus-3 OCT and FA. To evaluate agreement between the diagnostic approaches, κ statistics were used. Macular thickening (MT; center point thickness, ≥ 240 μm per reading center grading of OCT images) and macular leakage (ML; central subfield fluorescein leakage, ≥ 0.44 disc areas per reading center grading of FA images), and agreement between these outcomes in diagnosing macular edema. Optical coherence tomography (90.4%) more frequently returned usable information regarding macular edema than FA (77%) or biomicroscopy (76%). Agreement in diagnosis of MT and ML (κ = 0.44) was moderate. Macular leakage was present in 40% of cases free of MT, whereas MT was present in 34% of cases without ML. Biomicroscopic evaluation for macular edema failed to detect 40% and 45% of cases of MT and ML, respectively, and diagnosed 17% and 17% of cases with macular edema that did not have MT or ML, respectively; these results may underestimate biomicroscopic errors (ophthalmologists were not explicitly masked to OCT and FA results). Among eyes free of ML, phakic eyes without cataract rarely (4%) had MT. No factors were found that effectively ruled out ML when MT was absent. Optical coherence tomography and FA offered only moderate agreement regarding macular edema status in uveitis cases, probably because what they measure (MT and ML) are related but nonidentical macular pathologic characteristics. Given its lower cost, greater safety, and greater likelihood of obtaining usable information, OCT may be the best initial test for evaluation of suspected macular edema. However, given that ML cannot be ruled out if MT is absent and vice versa, obtaining the second test after negative results on the first seems justified when detection of ML or MT would alter management. Given that biomicroscopic evaluation for macular edema erred frequently, ancillary testing for macular edema seems indicated when knowledge of ML or MT status would affect management. Proprietary or commercial disclosure may be found after the references. Copyright © 2013 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.

Thousand-fold fluorescent signal amplification for mHealth diagnostics

PubMed Central

Balsam, Joshua; Rasooly, Reuven; Bruck, Hugh Alan; Rasooly, Avraham

2013-01-01

The low sensitivity of Mobile Health (mHealth) optical detectors, such as those found on mobile phones, is a limiting factor for many mHealth clinical applications. To improve sensitivity, we have combined two approaches for optical signal amplification: (1) a computational approach based on an image stacking algorithm to decrease the image noise and enhance weak signals, and (2) an optical signal amplifier utilizing a capillary tube array. These approaches were used in a detection system which includes a multi-wavelength LEDs capable of exciting many fluorophores in multiple wavelengths, a mobile phone or a webcam as a detector, and capillary tube array configured with 36 capillary tubes for signal enhancement. The capillary array enables a ~100X increase in signal sensitivity for fluorescein, reducing the limit of detection (LOD) for mobile phones and webcams from 1000 nM to 10 nM. Computational image stacking enables another ~10X increase in signal sensitivity, further reducing the LOD for webcam from 10 nM to 1 nM. To demonstrate the feasibility of the device for the detection of disease-related biomarkers, Adenovirus DNA labeled with SYBR Green or fluorescein was analyzed by both our capillary array and a commercial plate reader. The LOD for the capillary array was 5ug/mL, and that of the plate reader was 1 ug/mL. Similar results were obtained using DNA stained with fluorescein. The combination of the two signal amplification approaches enables a ~1000X increase in LOD for the webcam platform. This brings it into the range of a conventional plate reader while using a smaller sample volume (10ul) than the plate reader requires (100 ul). This suggests that such a device could be suitable for biosensing applications where up to 10 fold smaller sample sizes are needed. The simple optical configuration for mHealth described in this paper employing the combined capillary and image processing signal amplification is capable of measuring weak fluorescent signals without the need of dedicated laboratories. It has the potential to be used to increase sensitivity of other optically based mHealth technologies, and may increase mHealth’s clinical utility, especially for telemedicine and for resource-poor settings and global health applications. PMID:23928092

Thousand-fold fluorescent signal amplification for mHealth diagnostics.

PubMed

Balsam, Joshua; Rasooly, Reuven; Bruck, Hugh Alan; Rasooly, Avraham

2014-01-15

The low sensitivity of Mobile Health (mHealth) optical detectors, such as those found on mobile phones, is a limiting factor for many mHealth clinical applications. To improve sensitivity, we have combined two approaches for optical signal amplification: (1) a computational approach based on an image stacking algorithm to decrease the image noise and enhance weak signals, and (2) an optical signal amplifier utilizing a capillary tube array. These approaches were used in a detection system which includes multi-wavelength LEDs capable of exciting many fluorophores in multiple wavelengths, a mobile phone or a webcam as a detector, and capillary tube array configured with 36 capillary tubes for signal enhancement. The capillary array enables a ~100× increase in signal sensitivity for fluorescein, reducing the limit of detection (LOD) for mobile phones and webcams from 1000 nM to 10nM. Computational image stacking enables another ~10× increase in signal sensitivity, further reducing the LOD for webcam from 10nM to 1 nM. To demonstrate the feasibility of the device for the detection of disease-related biomarkers, adenovirus DNA labeled with SYBR green or fluorescein was analyzed by both our capillary array and a commercial plate reader. The LOD for the capillary array was 5 ug/mL, and that of the plate reader was 1 ug/mL. Similar results were obtained using DNA stained with fluorescein. The combination of the two signal amplification approaches enables a ~1000× increase in LOD for the webcam platform. This brings it into the range of a conventional plate reader while using a smaller sample volume (10 ul) than the plate reader requires (100 ul). This suggests that such a device could be suitable for biosensing applications where up to 10 fold smaller sample sizes are needed. The simple optical configuration for mHealth described in this paper employing the combined capillary and image processing signal amplification is capable of measuring weak fluorescent signals without the need of dedicated laboratories. It has the potential to be used to increase sensitivity of other optically based mHealth technologies, and may increase mHealth's clinical utility, especially for telemedicine and for resource-poor settings and global health applications. Published by Elsevier B.V.

Isolation of the ocular surface to treat dysfunctional tear syndrome associated with computer use.

PubMed

Yee, Richard W; Sperling, Harry G; Kattek, Ashballa; Paukert, Martin T; Dawson, Kevin; Garcia, Marcie; Hilsenbeck, Susan

2007-10-01

Dysfunctional tear syndrome (DTS) associated with computer use is characterized by mild irritation, itching, redness, and intermittent tearing after extended staring. It frequently involves foreign body or sandy sensation, blurring of vision, and fatigue, worsening especially at the end of the day. We undertook a study to determine the effectiveness of periocular isolation using microenvironment glasses (MEGS) alone and in combination with artificial tears in alleviating the symptoms and signs of dry eye related to computer use. At the same time, we evaluated the relative ability of a battery of clinical tests for dry eye to distinguish dry eyes from normal eyes in heavy computer users. Forty adult subjects who used computers 3 hours or more per day were divided into dry eye sufferers and controls based on their scores on the Ocular Surface Disease Index (OSDI). Baseline scores were recorded and ocular surface assessments were made. On four subsequent visits, the subjects played a computer game for 30 minutes in a controlled environment, during which one of four treatment conditions were applied, in random order, to each subject: 1) no treatment, 2) artificial tears, 3) MEGS, and 4) artificial tears combined with MEGS. Immediately after each session, subjects were tested on: a subjective comfort questionnaire, tear breakup time (TBUT), fluorescein staining, lissamine green staining, and conjunctival injection. In this study, a significant correlation was found between cumulative lifetime computer use and ocular surface disorder, as measured by the standardized OSDI index. The experimental and control subjects were significantly different (P<0.05) in the meibomian gland assessment and TBUT; they were consistently different in fluorescein and lissamine green staining, but with P>0.05. Isolation of the ocular surface alone produced significant improvements in comfort scores and TBUT and a consistent trend of improvement in fluorescein staining and lissamine green staining. Isolation plus tears produced a significant improvement in lissamine green staining. The subjective comfort inventory and the TBUT test were most effective in distinguishing between the treatments used. Computer users with ocular surface complaints should have a detailed ocular surface examination and, if symptomatic, they can be effectively treated with isolation of the ocular surface, artificial tears therapy, and effective environmental manipulations.