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Current Protocols in Immunology Nov 2015The protocol described in this appendix allows for light microscopic quantitation of cell viability. Cells are suspended in PBS containing trypan blue and then examined...
The protocol described in this appendix allows for light microscopic quantitation of cell viability. Cells are suspended in PBS containing trypan blue and then examined to determine the percentage of cells that have clear cytoplasm (viable cells) versus cells that have blue cytoplasm (nonviable cells).
Topics: Animals; Cell Survival; Cells, Cultured; Cytoplasm; Humans; Microscopy; Staining and Labeling; Trypan Blue
PubMed: 26529666
DOI: 10.1002/0471142735.ima03bs111 -
Romanian Journal of Ophthalmology 2016The term "chromovitrectomy" has been coined to define the use of vital dyes in vitreoretinal surgery. The basic concept for the application of vital dyes during... (Review)
Review
The term "chromovitrectomy" has been coined to define the use of vital dyes in vitreoretinal surgery. The basic concept for the application of vital dyes during vitreoretinal surgery is to assist in highlighting preretinal membranes and tissues which are very thin and semitransparent and thus difficult to detect. Various dyes are currently being used in routine clinical procedures, however, the ideal staining agent has not yet been found. The vital dyes indocyanine green, infracyanine green, and brilliant blue stain the internal limiting membrane, trypan blue and triamcinolone acetonide help to visualize the epiretinal and vitreous membranes. New dyes with a better safety profile than the synthetic ones are important for optimizing the outcome of modern ophthalmic surgery and natural dyes, such as lutein, offer a potentially safer and more efficient method of identifying intraocular structures such as vitreous and ILM. Any dye, which is intravitreally injected has the potential to become toxic.
Topics: Benzenesulfonates; Coloring Agents; Epiretinal Membrane; Humans; Indocyanine Green; Intravitreal Injections; Staining and Labeling; Triamcinolone Acetonide; Trypan Blue; Vitrectomy; Vitreoretinal Surgery; Vitreous Body
PubMed: 29450324
DOI: No ID Found -
Ophthalmic Surgery, Lasers & Imaging... Oct 2018Vital dyes contain complex molecules with chromophores that stain living tissues and have greatly enhanced identification and removal of transparent vitreoretinal... (Review)
Review
Vital dyes contain complex molecules with chromophores that stain living tissues and have greatly enhanced identification and removal of transparent vitreoretinal tissues during surgery. Several "chromovitrectomy" dyes are frequently used by vitreoretinal specialists, including indocyanine green, trypan blue, brilliant blue G, and triamcinolone acetonide; other dyes are also under investigation. Trypan Blue was approved by the U.S. Food and Drug Administration (FDA) for epiretinal membrane removal, and preservative-free triamcinolone acetonide was approved by the FDA for intraocular use. However, currently available chromovitrectomy dyes have their limitations, and of particular concern for some of them is the possibility for acute and chronic toxicity to the neurosensory retina and retinal pigmented epithelium. The potentially irreversible acute toxicity and other limitations, such as lack of long-term safety profiles, highlight the need for further advancements. [Ophthalmic Surg Lasers Imaging Retina. 2018;49:788-798.].
Topics: Coloring Agents; Humans; Indocyanine Green; Intraoperative Period; Macula Lutea; Retinal Diseases; Trypan Blue; Vitreoretinal Surgery; Vitreous Body
PubMed: 30395665
DOI: 10.3928/23258160-20181002-07 -
SN Applied Sciences 2021Zein-based materials were used to remove Trypan blue from water under flow conditions and in batch tests. In flow tests, zein dissolved at pH = 13 was injected in...
ABSTRACT
Zein-based materials were used to remove Trypan blue from water under flow conditions and in batch tests. In flow tests, zein dissolved at pH = 13 was injected in sand columns and subsequently coagulated with CaCl, to create an adsorbent filter which removed over 99% of Trypan blue. Batch tests were conducted using zein powder, zein dissolved at pH = 13 and coagulated with CaCl, FeCl or citric acid, and zein dissolved in ethanol and then coagulated with water. The highest Trypan blue removal was achieved with zein powder (4000 mg Trypan blue/kg sorbent, as determined through spectrophotometry), followed by zein coagulated with FeCl (500 mg Trypan blue/kg sorbent) and with other salts (140 mg Trypan blue/kg sorbent). Differences in the sorption efficiency are attributed to differences in the surface area. The sorption isotherm of Trypan blue onto zein-based sorbents was a Type II isotherm, suggesting physisorption. Desorption of Trypan blue was limited when zein-based coagulated sorbents were immersed in pure water. Trypan blue could be degraded by free laccase in water, as determined through spectrophotometry and electrospray ionization mass spectroscopy (ESI-MS). Trypan blue could also be degraded by laccase when zein-based laccase-containing sorbents were prepared at pH = 10, using FeCl as coagulant.
SUPPLEMENTARY INFORMATION
The online version contains supplementary material available at 10.1007/s42452-020-04107-w.
PubMed: 33442668
DOI: 10.1007/s42452-020-04107-w -
Ophthalmologica. Journal International... 2018To report on the combined use of trypan blue (TB) and brilliant blue G (BBG) for staining the epiretinal membrane (ERM) and internal limiting membrane (ILM) during... (Clinical Trial)
Clinical Trial
PURPOSE
To report on the combined use of trypan blue (TB) and brilliant blue G (BBG) for staining the epiretinal membrane (ERM) and internal limiting membrane (ILM) during vitrectomy and to describe the histopathological findings.
METHODS
10 surgical specimens were removed from 10 eyes with macular pucker during vitrectomy using a commercially available combination of TB and BBG for ERM and ILM staining and peeling. Specimens were evaluated using light and transmission electron microscopy.
RESULTS
In all cases the combination of TB and BBG was useful for identifying and delineating ERM and ILM. No complications related to the use of the dye were observed during or after surgery. Glial cells were present in all specimens. Hyalocytes were observed in 6 cases and myofibroblasts in 3 of them. In 7 cases native vitreous collagen fibrils were found on the ILM, while in 5 specimens newly formed collagen was present. No clinical evidence of toxicity was observed during the 3-month follow-up.
CONCLUSION
The combined use of TB and BBG appeared to be very useful intraoperatively to improve the visualization of ERM and ILM, thus facilitating their complete removal. Anatomical and histopathological findings demonstrated the safety and the efficacy of this vital dye.
Topics: Aged; Aged, 80 and over; Basement Membrane; Coloring Agents; Epiretinal Membrane; Female; Follow-Up Studies; Humans; Indicators and Reagents; Intraoperative Period; Macula Lutea; Male; Microscopy, Electron, Transmission; Prospective Studies; Rosaniline Dyes; Single-Blind Method; Time Factors; Trypan Blue; Visual Acuity; Vitrectomy
PubMed: 29393272
DOI: 10.1159/000485986 -
Tropical Medicine & International... Sep 2022This systematic review was undertaken to answer the research question: "In children with primary cataracts, what are the outcomes (posterior continuous curvilinear... (Review)
Review
OBJECTIVE
This systematic review was undertaken to answer the research question: "In children with primary cataracts, what are the outcomes (posterior continuous curvilinear capsulorhexis + posterior chamber intraocular lens implantation) of surgery when performed with and without trypan blue staining of the posterior lens capsule?"
METHODS
An electronic search in six biomedical databases was conducted to identify randomised controlled trials that compared trypan blue with no stain during surgery in children 0-16 years with primary cataracts. Titles and abstracts of studies published between 1946 and 2021 in English language were screened. Data extraction, risk of bias assessment and synthesis of findings were done by two independent reviewers, while conflicts were discussed and resolved with a third.
RESULTS
A total of 115 of 153 articles were screened after de-duplication. Of these, 113 were excluded while 2 randomised controlled trials involving 56 eyes of 42 participants were included in the review. The risk of bias was similar across all domains in both. Staining of the capsule led to complete posterior capsulorhexis and optimal placement of the implant in >90% of study eyes, while the control arms had 65%-80% for both outcomes.
CONCLUSION
Use of trypan blue in paediatric cataract surgery probably leads to better outcomes, but more well-conducted randomised controlled trials on this important topic are needed.
Topics: Capsulorhexis; Cataract; Cataract Extraction; Child; Coloring Agents; Humans; Lens Capsule, Crystalline; Randomized Controlled Trials as Topic; Trypan Blue
PubMed: 35859347
DOI: 10.1111/tmi.13801 -
Biosensors Nov 2022Simple staining of cells is a widely used method in basic medical diagnostics, education, and research laboratories. The stains are low-cost, but the extensive...
Simple staining of cells is a widely used method in basic medical diagnostics, education, and research laboratories. The stains are low-cost, but the extensive consumption results in excessive toxic waste generation. Thus, to decrease the amount of toxic waste resulting from the cell staining procedure is a need. In this study, we developed a magnetically driven and compartmentalized passive microfluidic chip to perform simple staining of human eukaryotic cells, K562 cells, and lymphocyte cells derived from patients. We demonstrated simple staining on cells with trypan blue, methylene blue, crystal violet, and safranin for high, medium, and low cell densities. The stained cells were imaged using a bright field optical microscope and a cell phone to count cells on the focal plane. The staining improved the color signal of the cell by 25-135-pixel intensity changes for the microscopic images. The validity of the protocol was determined using Jurkat and MDA-MB-231 cell lines as negative controls. In order to demonstrate the practicality of the system, lymphocyte cells derived from human blood samples were stained with trypan blue. The color intensity changes in the first and last compartments were analyzed to evaluate the performance of the chip. The developed method is ultra-low cost, significantly reduces the waste generated, and can be integrated with mobile imaging devices in terms of portability. By combining microfabrication technology with cell staining, this study reported a novel contribution to the field of microfluidic biosensors. In the future, we expect to demonstrate the detection of pathogens using this method.
Topics: Humans; Lab-On-A-Chip Devices; Staining and Labeling; Gentian Violet; Trypan Blue; Methylene Blue
PubMed: 36421132
DOI: 10.3390/bios12111013 -
Asia-Pacific Journal of Ophthalmology... Nov 2020Chromovitrectomy, the intraocular application of dyes to assist visualization of preretinal tissues during vitreoretinal surgery, was introduced to avoid ocular... (Review)
Review
Chromovitrectomy, the intraocular application of dyes to assist visualization of preretinal tissues during vitreoretinal surgery, was introduced to avoid ocular complications related to internal limiting membrane peeling, inadequate removal of the vitreous, and incomplete removal of epiretinal membranes. Since 2000, chromovitrectomy has become a popular approach among vitreoretinal specialists. The first vital dye used in chromovitrectomy, indocyanine green, facilitated identification of the fine and transparent internal limiting membrane. Following indocyanine green, trypan blue was introduced to identify epiretinal membranes, and triamcinolone acetonide stained the vitreous well. Recently, additional natural dyes such as lutein and anthocyanin from the açaí fruit have been proposed for intraocular application during vitrectomy. The main goal of this review was to study the role of vital stains in chromovitrectomy and report the latest findings in the literature.
Topics: Coloring Agents; Epiretinal Membrane; Humans; Indocyanine Green; Ophthalmologic Surgical Procedures; Staining and Labeling; Trypan Blue; Vitrectomy; Vitreoretinal Surgery; Vitreous Body
PubMed: 33252365
DOI: 10.1097/APO.0000000000000344 -
PloS One 2018To investigate the photochemical degradation of trypan blue (TB) and to identify decomposition products.
PURPOSE
To investigate the photochemical degradation of trypan blue (TB) and to identify decomposition products.
METHODS
Defined solution samples of TB and a mixture with lutein/zeaxanthin were exposed to blue light. Thermal degradation processes were ruled out using controls not subjected to irradiation. All samples were analyzed using optical microscopy, UV/Vis spectroscopy, matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and nuclear magnetic resonance (NMR) spectrometry. Degradation kinetics were determined based on changes in absorbance; intermediates were identified by analyzing mass differences of characteristic fragment ion peaks within the fragmentation patterns, and assignments were verified by NMR.
RESULTS
TB demonstrated a photochemical degradation, which can be triggered by lutein/zeaxanthin. Intermediates vary depending on the presence of lutein/zeaxanthin. The self-sensitized photodegradation of TB occurs under generation of dimethyl sulfate and presumed formation of phenol. In contrast, within the presence of lutein/zeaxanthin the decomposition of TB indicates the formation of methoxyamine and sulfonyl arin. Thermal degradation processes were not observed.
CONCLUSIONS
TB demonstrated a photodegradation that may be triggered by lutein/zeaxanthin and results in the formation of cytotoxic decomposition products. Our findings contribute to understand degradation mechanisms of TB and may elucidate previous clinical and experimental observations of cellular toxicity after TB application.
Topics: Kinetics; Light; Lutein; Photochemistry; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Trypan Blue; Zeaxanthins
PubMed: 29634764
DOI: 10.1371/journal.pone.0195849 -
Journal of Nanobiotechnology Oct 2021There is a huge body of literature data on ZnOnanoparticles (ZnO NPs) toxicity. However, the reported results are seen to be increasingly discrepant, and deep...
BACKGROUND
There is a huge body of literature data on ZnOnanoparticles (ZnO NPs) toxicity. However, the reported results are seen to be increasingly discrepant, and deep comprehension of the ZnO NPs behaviour in relation to the different experimental conditions is still lacking. A recent literature overview emphasizes the screening of the ZnO NPs toxicity with more than one assay, checking the experimental reproducibility also versus time, which is a key factor for the robustness of the results. In this paper we compared high-throughput real-time measurements through Electric Cell-substrate Impedance-Sensing (ECIS®) with endpoint measurements of multiple independent assays.
RESULTS
ECIS-measurements were compared with traditional cytotoxicity tests such as MTT, Neutral red, Trypan blue, and cloning efficiency assays. ECIS could follow the cell behavior continuously and noninvasively for days, so that certain long-term characteristics of cell proliferation under treatment with ZnO NPs were accessible. This was particularly important in the case of pro-mitogenic activity exerted by low-dose ZnO NPs, an effect not revealed by endpoint independent assays. This result opens new worrisome questions about the potential mitogenic activity exerted by ZnO NPs, or more generally by NPs, on transformed cells. Of importance, impedance curve trends (morphology) allowed to discriminate between different cell death mechanisms (apoptosis vs autophagy) in the absence of specific reagents, as confirmed by cell structural and functional studies by high-resolution microscopy. This could be advantageous in terms of costs and time spent. ZnO NPs-exposed A549 cells showed an unusual pattern of actin and tubulin distribution which might trigger mitotic aberrations leading to genomic instability.
CONCLUSIONS
ZnO NPs toxicity can be determined not only by the intrinsic NPs characteristics, but also by the external conditions like the experimental setting, and this could account for discrepant data from different assays. ECIS has the potential to recapitulate the needs required in the evaluation of nanomaterials by contributing to the reliability of cytotoxicity tests. Moreover, it can overcome some false results and discrepancies in the results obtained by endpoint measurements. Finally, we strongly recommend the comparison of cytotoxicity tests (ECIS, MTT, Trypan Blue, Cloning efficiency) with the ultrastructural cell pathology studies.
Topics: A549 Cells; Apoptosis; Cell Survival; Cloning, Molecular; Electric Impedance; Humans; Lung; Metal Nanoparticles; Microscopy, Confocal; Microscopy, Electron; Toxicity Tests; Trypan Blue; Zinc Oxide
PubMed: 34620157
DOI: 10.1186/s12951-021-01033-w