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PubMed2002

Trypan blue capsular staining to "find" the leading edge of a "lost" capsulorhexis.

PURPOSE: To describe an effective surgical technique to visualize an obscured leading edge of a capsulorhexis. DESIGN: Prospective, interventional case series. METHODS: In six eyes of six patients in which the capsulorhexis was started but could no longer be seen during surgery, the anterior chamber was irrigated to remove all viscoelastic, and 0.3-ml trypan blue 0.06% was applied onto the anterior lens capsule to stain and visualize the leading edge of the capsulorhexis. RESULTS: In all patients, the leading edge of the capsulorhexis was quickly visualized by using the dye. No adverse reactions were observed up to 1 year after surgery. CONCLUSION: Trypan blue staining of the anterior lens capsule is an effective and apparently safe technique to "find" a "lost" capsulorhexis during surgery.

Capsulorhexis↗
PubMed2005

In vitro quantitative determination of ophthalmic irritancy by the chorioallantoic membrane test with trypan blue staining as alternative to eye irritation test.

The damage provoked by some substances on the chicken egg's chorioallantoic membrane (CAM) is used as an alternative assay to determine ocular irritation. There is good prediction of the eye irritation when compared to the in vivo Draize method. Nevertheless, this assay has some limitations, such as subjectivity. Hagino et al. developed an objective evaluation technique using the amount of trypan blue absorbed at the site of treatment as an indicator of injury to the CAM. The present work was aimed at the determination of ocular irritation of 21 substances (chemicals and cosmetics). We used the spectrophotometric quantification by trypan blue staining of the damage produced on CAM, of fertile chicken eggs. Results were compared with the values obtained by the traditional Draize assay. We observed a good correlation (r=0.835; p<0.0001) between the amount of dye absorbed by the CAM and the Draize eye irritation test score. The r values were 0.688; p<0.05 for cosmetics and 0.925; p<0.0001 for chemicals. Three chemicals turned as false positive and one cosmetic substance as false negative. The CAM-TBS assay is inexpensive, simple and provides an in vitro alternative method to predict the damage that chemical substances or cosmetics can cause to the ocular structures.

Animals↗
PubMed1976

Pathogenesis of trypan-blue-induced spina bifida.

Congenital malformations induced in rats by the application of trypan blue were studied at variable times of their development of congenital malformations is a continuous process of structural and functional adaptation of the development embryo to focal tissue degeneration and necrosis (cell death). The primary degeneration is a transitory phenomenon, because the dead cells are phagocytosed by surviving cells. However, the damage triggers off the process of abnormal development.

Animals↗
PubMed1987

Heat-induced K+ loss, trypan blue uptake, and cell lysis in different cell lines: effect of serum.

Experiments were performed with three different cell lines, mouse fibroblast LM cells, HeLa S3 cells, and Ehrlich Ascites Tumor (EAT) cells, to establish the possible importance of hyperthermic-induced alterations in cellular K+ content in the mechanism of cell killing by heat. At different time points after the hyperthermic treatment, the K+ content in the cells, the uptake of the dye Trypan Blue (TB), and cell lysis were assayed. Heat-induced K+ loss preceded TB uptake which was followed by the heat-induced cell lysis. Lysis was assayed as disappearance of cells by counting the cells at different time points in a hematocytometer. The presence of serum during and after the heat treatment was of considerable importance with respect to K+ loss and TB uptake. K+ loss and TB uptake after the heat treatment were less when serum was present during and after hyperthermia. To protect against cell lysis, however, the serum had to be present during a preincubation period of 24 h. Clonogenic ability was not affected by the presence of serum. It is concluded that the intracellular K+ level of hyperthermic-treated cells is not a direct cause for cell killing and that heat-induced alterations in the cell leading to cell lysis are different from the processes decreasing cellular K+ content and permeabilizing the plasma membrane for trypan blue.

Animals↗
PubMed1980

Specific tumor immunity induced with mitomycin C-treated syngeneic tumor cells (MCT). Effects of carrageenan and trypan blue on MCT-induced immunity in mice.

Mitomycin C treatment of 3-methylcholanthrene-induced syngeneic tumor cells in male mice of the highly inbred strain CBA/WEHI was a convenient and effective method for producing viable but nondividing tumor cells. A single sc injection of 1 x 10(6) mitomycin C-treated syngeneic tumor cells (MCT) induced a specific antitumor immune response that could mediate rejection of small subcutaneous tumor grafts and substantially inhibit growth of larger grafts in normal mice. The response was evident between 3 and 7 days after immunization and persisted for at least 120 days. Adoptive transfer experiments with peritoneal exudate cells from MCT-immunized mice indicated that a cellular basis for resistance was likely in the absence of a detectable humoral component. Administration of MCT to mice with established tumor grafts did not, however, affect tumor growth. Experiments with carrageenan and trypan blue showed that neither induction nor expression of cytotoxic effector cells in MCT-immunized mice was inhibited by these substances, though carrageenan and trypan blue augmented tumor growth in normal mice.

Animals↗
PubMed1990

Increased attachment and confluence of skin epidermal cells in culture induced by ascorbic acid: detection by permeation of trypan blue across cultured cell layers.

In culture epidermal cells from the skin of newborn rats became attached to Millipore filters coated with type IV collagen much better than to filters coated with type I collagen. Ascorbic acid markedly increased the attachment and viability of epidermal cells seeded on type I collagen, but had no significant effect on cells seeded on type IV collagen. It was also found to enhance the synthesis of type IV collagen by the cells, which, we concluded, enabled the cells to become well attached to type I collagen. This conclusion was supported by studies on the penetration of trypan blue through the cell layers. There was a lag in penetration through cell layers cultured both with and without ascorbic acid on Millipore filters coated with either type I or IV collagen, indicating that the cells were confluent over the whole surface of the filters. The lag was much longer in the cultures with ascorbic acid, indicating greater confluence and tighter attachment of cells due to production of type IV collagen. The penetration was found to be due to destruction of the confluent cell layers by its cytotoxic effect. The time lag before penetration of trypan blue is a good index of the confluence and attachment of cultured cells to collagen layers.

Animals↗
PubMedDate not supplied

Gamma-glutamyl transferase (GGT) activity and biochemical characterization of rat visceral yolk-sac during gestation with or without trypan blue exposure.

Yolk-sacs from untreated Sprague-Dawley rat conceptuses were removed on gestational days (GD) 9 to 18 and examined for gamma-glutamyl transferase (GGT), alkaline phosphatase (AP), lactate dehydrogenase (LDH), and glutamic-oxaloacetic transaminase (GOT) activities. All enzyme activities were found to increase through gestation in total activity as well as in specific activity. Protein (PRO) and urea nitrogen (UN) were also found to increase through gestation whereas triglyceride (TRI) increased steadily from GD 9 to 13 and then appeared to plateau through GD 19. Additional rats were treated on GD 8 with 75 mg trypan blue (TB)/kg body weight and yolk-sacs taken on GD 9 to 18. Yolk-sac GGT and GOT activities from TB-treated rats were significantly higher than the respective controls during early gestation but recovered to or were lower than control levels during midgestation. GGT activity in treated rats was significantly higher than the controls on GD 15 and 16, and both GGT and GOT were significantly lower than controls on GD 17 and 18. AP activity in the TB-treated yolk-sacs was significantly lower than that in controls during early and midgestation but was not significantly different from the control values late in gestation. Triglyceride concentration was not affected early in gestation but significantly decreased on GD 16 and 18. Thus, the yolk-sac enzymes monitored, which are associated with nutrition and normal growth, increased in activity through GD 18. The yolk-sac toxicant, trypan blue, significantly affected enzymatic activity at various time periods during gestation and resulted in significant changes in yolk-sac protein and triglyceride content.

Alkaline Phosphatase↗
PubMed2006

[Risk of contamination of trypan blue dye after first use].

PURPOSE: To determine the potential risk of contamination of a trypan blue bottle (TB) after first use and after being stored under different temperature and humidity conditions, as well as to identify possible contamination factors, most frequently involved microorganisms and simultaneously evaluate bacteriostatic and bactericide properties of the dye. METHODS: An experimental and prospective study was carried out, in which 30 TB bottles were divided into 3 groups (A: control, B: refrigerator storage and C: cabinet storage). The dye was suctioned and cultivated in agar blood plates and Sabouraud agar tube. In group A, TB was cultivated immediately after the opening of the bottles (temperature zero - T0), in groups B and C cultivation occurred in T0, T1 (1 day), T2 (2 days), T7 (7 days) and T10 (10 days) after the opening of the bottles. On the 10th day, groups B and C bottles were also submitted to scraping of their inner walls after opening. Concurrently, the inhibitory action measurement test was conducted on the TB dye for the study of bacteriostatic and bactericidal activity. RESULTS: Cultivation procedures conducted in T0 presented no contamination. Among T1 and T10, added to the scraping there was only one contaminated bottle stored in the refrigerator. The encountered microorganism was Aspergillus niger. It has been proven that the dye does not show bactericide and bacteriostatic properties against the bacteria which were tested. CONCLUSIONS: Under this study's conditions there was no contamination of the bottles stored in cabinets and 1 bottle (10%) stored in the refrigerator showed contamination after opening and initial use. The source of contamination may possibly be the outer part of the product. TB does not show bactericidal and bacteriostatic properties against the tested bacteria and in the applied concentration.

Aspergillus niger↗
PubMed1985

Schistosoma mansoni: suramin and trypan blue in vitro and the ultrastructure of feeding schistosomules.

The in vitro effects of suramin and trypan blue on schistosomules of Schistosoma mansoni were examined by light and electron microscopy. The drugs were administered to 12-day cultures of schistosomules produced by the penetration method. The larvae were maintained on mouse red blood cells for 5 days prior to addition of the drugs. At the concentrations used, the morphological changes attributable to the drugs were identical for the two drugs. The first signs of anomaly were observable at 8 hr after exposure. At this time, the digestive system showed signs reminiscent of early starvation effects such as alterations of the Golgi and an increase of autophagy. After 36 hr, a rapid disintegration of the gastrodermis became evident. It was hypothesized that the drugs initially inhibit the digestive enzymes and, subsequently, labilize the luminal plasma membrane of the gastrodermis. By 48 hr after exposure, the larvae were dead.

Animals↗
PubMed2001

Ocular toxicity study of trypan blue injected into the vitreous cavity of rabbit eyes.

BACKGROUND: To evaluate the ocular toxicity of trypan blue (TB) injected into the vitreous cavity of rabbit eyes. TB is a dye that could be useful for staining epiretinal membranes during vitrectomy surgery. METHODS: Ten New Zealand White (NZW) rabbits underwent gas-compression vitrectomy. Rabbits were divided into three groups to receive injections of 0.1 ml basic salt solution, 0.1 ml of a 0.06% TB solution or 0.1 ml of a 0.2% TB solution. Ocular toxicity was assessed by slit-lamp biomicroscopy, ophthalmoscopy, electroretinography and histology. RESULTS: Transient posterior capsule opacification was noted in all animals. No significant reductions in a-wave or b-wave amplitudes were found in any of the animals. Light and electron microscopic examination of the inferior retina in the 0.2% TB-treated eyes showed damaged photoreceptors and marked disorganization. Immunohistochemical staining for rhodopsin was strongly reduced in those sections and staining for proliferation with Ki-67 was positive. No histological abnormalities were found in the upper retina of the 0.2% TB-treated eyes or in any part of the retina of the 0.06% TB-treated or control eyes. No histological abnormalities were found in any of the anterior chamber angle specimens. CONCLUSIONS: Although no signs of toxicity were found after the prolonged presence of TB at a concentration of 0.06% in the vitreous cavity of rabbit eyes, marked damage occurred in the lower retina of 0.2% TB-treated eyes. The short-term presence of TB at a concentration of 0.06% in the vitreous cavity is harmless to the rabbit eye but a higher concentration of TB could be unsafe.

Animals↗
PubMed2006

[Unintentional staining of lens posterior capsule with trypan blue during extracapsular cataract extraction: case report].

Report of a complication related to the use of 0.1% trypan blue during capsular staining of the anterior crystalline lens capsule in an extracapsular cataract extraction with intraocular lens implantation. The capsular dye was injected using an air-bubble technique, inadvertently, it was done using high pressure dispersing the dye through the zonules leading to a complete lost of red reflex, the following steps of the procedure becoming more difficult after the cataract extraction and causing a temporary disturbance of visual acuity in the postoperative period. In order to avoid this complication, the authors also describe the use of "Modified air-bubble technique" where dispersive viscoelastic is placed into the iridolenticular space 360 degrees to create a protection barrier.

Adult↗
PubMed1982

[Studies on the pharmacological bases of fetal toxicity of drugs. (II). Effect of trypan blue on the pregnant rats and their offspring].

We performed studies on the mechanisms for the teratogenicity of trypan blue (TB) that is known as a potent teratogenic dye in rodents. TB was administered to Wistar rats at various doses and at different stages of gestation. Teratogenicity and embryolethality were observed by a single (50 and 250 mg/kg) subcutaneous injection and three (25 and 50 mg/kg) subcutaneous injections of TB daily from day 7 of pregnancy. The incidences of malformations in groups given 50 and 250 mg/kg on day 7 were 12 and 59%, respectively. Most of the fetuses with external malformations were accompanied with skeletal and/or internal anomalies. The types of frequently occurring malformations were as follows: exencephaly, spina bifida, tail anomaly, vertebral deformity, hydrocephaly and heart anomaly. Fetal toxicity was decreased after treatment with the mixture of TB and normal rat serum. The serum level of TB in pregnant rats increased to 308 micrograms/ml at one hour after subcutaneous treatment with TB 50 mg/kg and decreased rapidly, but remained at 58 micrograms/ml 72 hours later. Lower serum levels of TB were observed in pregnant rats given TB with serum. No fetotoxic effects of serum from pregnant rats treated with TB were observed in recipient rats given the serum on day 7, 8 and 9 of pregnancy.

Abnormalities, Drug-Induced↗
PubMed2002

Deep lamellar keratoplasty with trypan blue intrastromal staining.

This technique was developed to facilitate removal of the deep stromal layers during deep lamellar keratoplasty. A trypan blue 0.02% solution is injected into the stromal fibers, enabling the surgeon to visualize and remove the posterior stromal layers. This decreases the risk of perforation of Descemet's membrane and the endothelium.

Coloring Agents↗
PubMed1984

Lack of uptake of indocyanine green and trypan blue by hepatocellular carcinoma.

Experimental hepatocellular carcinoma (HCC) in rats did not take up intravenously administered indocyanine green (ICG) and trypan blue, while surrounding tissue did. The lack of ICG uptake was also observed by peritoneoscopy in patients with HCC. The contrast between ICG-stained cirrhotic nodules and HCC tumors was intensified with infrared photography. Non-uptake of dyes by HCC cells may enable discrimination between tumors and normal cells.

Animals↗
PubMed2005

Neurotoxic effects of trypan blue on rat retinal ganglion cells.

To facilitate the peeling of internal limiting membrane or epiretinal membrane in vitreoretinal surgery, trypan blue (TB) and indocyanine green (ICG) have been used. However, the cytotoxicity of these dyes have been concerned. The aim of this study was to investigate the neurotoxic effects of TB on rat retinal ganglion cells (RGCs) and compare the effect of TB with ICG. Rat RGCs were purified by a two-step immunopanning procedure. In short-time exposure experiments, purified RGCs cultured for 3 days were exposed to 600 mgL(-1) TB, 1500 mgL(-1) TB and 1500 mgL(-1) ICG for 10 sec to 30 min. The number of viable RGCs was counted after 12 hr in culture. In long-time exposure experiments, purified RGCs were cultured for 3 days in TB solutions ranging from 2 to 800 mgL(-1) or in ICG solutions at concentrations from 2 to 250 mgL(-1). Then the number of viable cells was counted. Exposure to 600 mgL(-1) TB, 1500 mgL(-1) TB and 1500 mgL(-1) ICG for short time from 10 sec to 30 min caused a time-dependent damage to RGCs. There was no significant difference in cytotoxicity to RGCs between TB and ICG in short-time exposure. In long-time exposure, TB as well as ICG showed neurotoxic effect on RGCs in a dose-dependent manner. Fifty percentage inhibitory concentration (IC50) of TB to RGCs was calculated as 115 mgL(-1), while that of ICG was 33 mgL(-1). In conclusion, TB induced neurotoxic effect on RGCs in a dose- and time-dependent manner. During the short-time exposure in surgery, there is likely no difference in neurotoxic effect on RGCs between TB and ICG. However, once the dyes are left in the eyes after surgery, TB may show less toxicity on RGCs than ICG.

Animals↗
PubMed1984

Colorimetric determination of microgram quantities of polylysine by trypan blue precipitation.

A simple and sensitive method for the determination of polylysine in solution is described. Polylysine is quantitatively precipitated with trypan blue. The absorbance of unbound dye in the supernatant is inversely proportional to the concentration of this polyamino acid. The precipitation is identical for all sizes of polylysine of molecular weight 13,000 or higher, and is prevented by the addition of either polyanions or serum. The measurable range of polylysine hydrobromide is between 1 and 10 micrograms/ml, which is about 10-fold lower than that by the published methyl orange precipitation method.

Azo Compounds↗
PubMed2004

Three-step technique for staining the anterior lens capsule with indocyanine green or trypan blue.

A 3-step technique for staining the anterior lens capsule was developed to optimize the use of a capsule dye (trypan blue or indocyanine green) during capsulorhexis. After the injection of a viscoadaptive ophthalmic viscosurgical device (OVD) (sodium hyaluronate 2.3% [Healon 5]), which fills the anterior chamber, balanced salt solution (BSS) is gently injected onto the anterior capsule, creating a wafer-thin, fluid-filled space beneath the OVD. The dye is slowly injected through a 27-gauge cannula (Duckworth & Kent), selectively mixing with the BSS and allowing relatively accurate painting of the anterior capsule, approximating the size and shape of the intended capsulorhexis. This 3-step technique avoids the use of air as well as the uncontrolled dispersion and excessive accumulation of dye within the OVD.

Capsulorhexis↗
PubMed2004

Comparison of the in vitro toxicity of indocyanine green to that of trypan blue in human retinal pigment epithelium cell cultures.

PURPOSE: To compare the in vitro toxicity of indocyanine green (ICG) to that of trypan blue (TB) in human retinal pigment epithelium cell cultures. The use of ICG and TB in macular hole surgery is discussed. DESIGN: In vitro cell biology experimental study. METHODS: The ICG dye and TB were applied to ARPE-19, a commercially available human retinal pigment epithelium cell line. Cultures were established and maintained according to supplier protocols. The ICG dye, TB or Hank's balanced salt solution (controls) were then applied to the cells at varying concentrations and over various exposure periods. Fiberoptic light was also applied to cells to assess for the possibility of a potentiating phototoxic effect. Cell viability fractions were determined using a well-studied mitochondrial dehydrogenase assay. RESULTS: The TB was not toxic to the retinal pigment epithelium cell cultures at any concentration or over any period of exposure, whereas ICG dye demonstrated dose-dependent and exposure-dependent toxicity. The ICG dye was found to be toxic to the cells at all tested concentrations between 5.0 mg/ml (stock concentration, 26.1% cell survival) and 0.5 mg/ml (92.8% cell survival) over a 3-minute exposure. No toxicity to TB was seen at the stock concentration of 1.5 mg/mL. Addition of light to the cultures did not significantly alter cell viability with either dye. Long periods of exposure, 2 hours, 24 hours, and 72 hours, to minute concentrations of either dye did not produce significant cell death. CONCLUSIONS: Indocyanine green demonstrates more toxicity than TB to human retinal pigment epithelium cell cultures. This is independent of any phototoxic potentiating effect of fiberoptic light or solvent toxicity. A clinically useful concentration of 0.5-mg/ml ICG causes low cytotoxicity at 3 minutes' exposure (cell survival 92.8%) and shows no detectable toxicity at 1-minute exposure (cell survival 102%).

Cell Line↗