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PubMed1994

Direct vs. indirect effects of trypan blue on the ectodermal cells of the 11.5-day rat embryo?

The effect of trypan blue on the 11.5-day rat conceptus after intravitelline vessel administration is described. For comparison, conceptuses injected with varying volumes of Hanks' BSS have also been studied. Trypan blue significantly retarded the growth and development of conceptuses after 6 hours incubation in vitro. The SEM revealed rounded ectodermal cells, some of which appeared disrupted. These cells seemed to cause some of the intersomitic grooves to disappear, making a number of the somites indistinct from the outside. Unlike cells of uninjected embryos, the surfaces of the affected ectodermal cells lacked microvilli and their perimeters were lined with microvilli-like structures which appeared matted together. It was concluded that trypan blue affected the embryo directly probably by disturbing its fluid and ionic balance.

Animals↗
PubMedDate not supplied

Effects of trypan blue and related compounds on production and activity of streptolysin S.

Most dyes related to trypan blue inhibited hemolytic activity of oligonucleotide-streptolysin S (SLS) complex, an exotoxin produced by Streptococcus pyogenes. Order of the inhibition was: trypan blue greater than benzo blue 2B greater than Congo red greater than Evans blue greater than benzo purpurine 4B greater than thiazine red greater than trypan red. When resting streptococcal cells were incubated with these dyes, significant amount of the hemolysin was produced. The carrier (or inducing) activity for SLS was further manifested in growing cell system and the potency of the compounds was as follows: Congo red greater than benzo blue 2B greater than trypan blue greater than Evans blue greater than Benzo purpurine 4B greater than or equal to thiazine red greater than trypan red. In this system, Congo red was more effective than oligonucleotide fraction rich in guanyl residue. Chromotrope 2B, H acid and o-tolidine were ineffective, as the carrier as well as the inhibitor. Based on these results, structure-function relationship among SLS, the carrier and the inhibitor was discussed.

Animals↗
PubMed2004

Rapid and reliable DNA extraction techniques from trypan-blue-stained mycorrhizal roots: comparison of two methods.

Two improved DNA extraction techniques from trypan-blue-stained root fragments were developed and compared for rapid and reliable analyses. In Method A, 1 cm trypan-blue-stained mycorrhizal root fragments were individually isolated, crushed by bead beating, and purified with Chelex-100 (Bio-Rad). In Method B, DNA extraction was carried out using an UltraClean microbial DNA isolation kit (MoBio Laboratories). DNA was extracted from the mycorrhizal roots of four plant species, quantified by UV absorbance, and PCR-amplified with primers specific to arbuscular mycorrhizal fungi. Although PCR inhibitors might still exist when using Method A, appropriate dilution and employment of nested-PCR overcame this problem. Method B removed PCR inhibitors, but sometimes, depending on the mycorrhizal colonization within the root fragments, it also required nested PCR. In conclusion, both methods enabled us to handle many samples in a short time. Method B provided greater reliability and Method A provided better cost performance. Both techniques can be useful for PCR-based applications to identify species and estimate species composition after measuring mycorrhizal colonization rate with trypan blue staining.

DNA Primers↗
PubMed1979

Fine structural analysis of the effect of trypan blue on the visceral endoderm of the mouse egg cylinder.

The effects of a maternal injection of trypan blue on primitive streak mouse embryos were studied with electron microscopy. Commercial trypan blue was purified by descending paper chromatography, and pregnant females received an intraperitoneal injection of the collected blue fraction on the evening of the 7th day of gestation. Ultrastructurally, the changes in the visceral endoderm were apparent 10 min after the injection and included an increase in the number of fuzzy-coated vesicles in the apical cytoplasm. By 20 min the apical cytoplasm of the extraembryonic visceral endodermal cells was filled with many fuzzy-coated vesicles and numerous vacuoles of various size and electron density. 30 min after the injection, the extraembryonic visceral endodermal cells were relatively smooth lacking a microvillous border and evidence of endocytic activity was rare. Many embryonic visceral endodermal cells were observed in various stages of degeneration although the underlying embryonic ectoderm appeared unaffected. Morphologically, it appears that trypan blue exerts its first effect by altering the endocytic activity of the visceral endoderm.

Animals↗
PubMed1984

Predictive value of trypan blue exclusion viability measurements for colony formation in a human tumor cloning assay.

Specimens (173) were evaluated to assess the predictive value of cell viability as measured by Trypan Blue exclusion on colony formation in a human tumor cloning system. Overall, there was no significant correlation between Trypan Blue exclusion-determined viability and colony formation in soft agar for these specimens (p = 0.43). This lack of significant correlation was maintained for primary, metastatic, solid, and fluid tumor specimens. These data suggest that, in this soft agar system, specimens containing cells with low Trypan Blue exclusion viability should not be excluded from testing, and that it may not be advantageous to plate on the basis of "viable cells" as measured by Trypan Blue exclusion.

Cell Survival↗
PubMed1983

Assessment of adult skeletons to detect prenatal exposure to trypan blue in mice.

A Skeletal Variant Assay System (SVAS) consisting of a group of 88 spontaneously occurring qualitative variations of the adult mouse skeleton was studied in CD-1 mice which had been exposed in utero by way of three daily ip injections of their dams on days 7-9 of gestation with trypan blue. Treatment groups received daily doses of 0.25 cc of 0, .037, .075, .15, or .30% trypan blue dissolved in 0.9% NaCl. Two separate series of experiments were performed, and skeletons were examined at 62 +/- 2 days postnatal. Sixty-six and 58 of the variants occurred in the two series, respectively. Frequencies of occurrence of substantial numbers of variants differed from Untreated (UNTD) and Vehicle-Treated (VEH) values in a dose-related manner for both series. At the high dose 18 and 22 variants occurred with significantly different (P less than .01) frequencies from UNTD in the two series. Contrasting high-dose animals with vehicle controls revealed significant differences in 24 and 17 variants. There were 13 and 14 variants in the two series, respectively, which differed from both UNTD and VEH. If one considers differences at P less than .01 in one comparison and P less than .05 in the other, then 22 and 18 variants qualify as being significantly different from both controls in the two series. Agreement between the two series was excellent regarding which variants were affected. Several differed significantly from both UNTD and VEH in both series of experiments. Among these were a number which appeared more or less specific to trypan blue exposure. They include Dyssymphysis of the Atlas, Carpal Fusions, and Tarsal Fusions. Although increased frequency of an Interfrontal bone is seen with several treatments, the magnitude of the response and the low doses at which it is elicited are unique to trypan blue exposure. Numerous low-dose effects are striking in this set of experiments, making the SVAS a very sensitive indicator of trypan blue exposure. In addition to the variants mentioned, a large cluster of cervical (C) vertebrae variants, including dyssymphyses, fusions, imperfect transverse foramina of C1 and C2, and accessory transverse foramina of C3-C6, as well as vertebral fusions at various levels (especially cervical, sacral, and caudal), appear to be the principal effects of exposure to this compound. In addition, rib malformations at the high dose level, and increased frequency of occurrence of 27-presacral vertebrae at all dose levels, were important effects.(ABSTRACT TRUNCATED AT 400 WORDS)

Abnormalities, Drug-Induced↗
PubMed1992

Heparin, dextran and trypan blue allosterically modulate M2 muscarinic receptor binding properties and interfere with receptor-mediated inhibition of adenylate cyclase.

The influences of heparin, dextran and trypan blue on muscarinic receptor binding properties and inhibition of adenylate cyclase were investigated in homogenates of the rat heart. These compounds caused a concentration-dependent enhancement in the specific binding of the muscarinic antagonist [3H]N-methylscopolamine ([3H]NMS) when measured at a radioligand concentration of approximately 0.5 nM in magnesium-containing, low ionic strength buffer. The maximal enhancements of [3H]NMS binding were 2.89-, 1.68- and 1.43-fold increases for heparin, dextran and trypan blue, respectively; the EC50 values for this effect were 0.12, 0.033 and 4.6 microM, respectively. The effects of heparin, dextran and trypan blue on [3H]NMS binding were attributed mainly to an increase in the overall affinity of muscarinic receptors for [3H]NMS, and were greatly attenuated by 100 mM NaCl. These effects were qualitatively similar to those produced by GTP. Heparin, dextran and trypan blue also affected the binding of the muscarinic agonist oxotremorine-M in a manner similar to that of GTP; that is, in the presence of these compounds, agonist affinity was decreased. Our experiments also showed that heparin and dextran attenuate the inhibition of adenylate cyclase activity caused by oxotremorine-M in myocardial homogenates without influencing basal adenylate cyclase activity. We conclude that heparin and dextran interfere with the muscarinic receptor-G protein coupling in the rat heart.

Adenylyl Cyclase Inhibitors↗
PubMed2000

The quantitive chlorioallantoic membrane test using trypan blue stain to predict the eye irritancy of liquid scintillation cocktails.

The chorioallantoic membrane-trypan blue staining assay (CAM-TBS) is used to evaluate the potential ocular irritation caused by liquid scintillation cocktails constituted by complex mixtures, including surfactants and other potential irritants. The harmful effect of these substances is determined by the amount of trypan blue adsorbed by the CAM. In the membrane previously treated with the scintillation liquids this amount was concentration dependent only in certain cases, irrespective of the water solubility of the mixtures. In general, it showed a high correlation (r=0.986) with the scores obtained in the Draize eye irritation test. In the present study, only two cocktails presented values of trypan blue adsorption higher than those recorded for their eye irritation in vivo, irrespective of the solvent nature. Unlike the classical HET-CAM procedure, this assay is objective and allows the evaluation of opaque and coloured substances without interfering in the determination of irritancy. Despite these advantages, the method is not suitable for complex mixtures of products that induce ocular irritation in small quantities.

Allantois↗
PubMed2005

[Vitreous surgery with trypan blue staining of membranes in PVR retinal detachment surgery].

BACKGROUND: In our clinic we used trypan blue staining routinely for removal of internal limiting membranes (ILM) and epiretinal membranes (ERM) during vitreoretinal surgery for PVR retinal detachment. PATIENTS AND METHODS: We treated 27 eyes of 27 patients with severe PVR retinal detachment with vitreoretinal surgery. After complete vitrectomy and removal of epiretinal membranes we injected 0.15 % trypan blue solution and proceeded with further removal of the visualized remaining epiretinal membranes and ILM. We performed relaxing retinotomies in 14 eyes. Follow-up lasted six months. RESULTS: After the first surgery the retina was reattached in 24 of 27 eyes and after the second surgery in all eyes. The final visual acuity was: hand movements in 3 eyes, 1/50 or better in 24 eyes (eight of them had a visual acuity of 0.1 - 0.5). No case of macular pucker was observed during the follow-up. CONCLUSIONS: Trypan blue staining during vitreoretinal surgery in PVR retinal detachment therapy allows complete removal of ILM, ERM, and peripheral vitreous. It upgrades the quality of the surgery.

Female↗
PubMed1984

[Studies on the pharmacological bases of fetal toxicity of drugs. (V) Effect of different administration routes of trypan blue in rats].

The teratogenicity of trypan blue (TB) after both maternal and intrauterine administrations was studied in Wistar rats, and the following results were obtained: 1) The teratogenic dose of TB by maternal subcutaneous injection was found to be greater than 50 mg/kg, and the critical period was until day 10 of pregnancy (sperm = day 0). No teratogenicity was detected by oral administration of 250 mg/kg TB. 2) TB (250 micrograms/uterine horn) was injected into the uterine cavity on day 4 or 6 of pregnancy. An increase of intrauterine death without malformations was observed in both TB-treated groups. 3) TB was injected into the exocoelom. The incidences of malformed fetuses were 53% in the group injected with 2.5 micrograms/embryo TB on day 10 and 32% and 57% in the groups injected with TB at 1.0 and 2.5 micrograms/embryo on day 11, respectively. An increase of intrauterine death was observed in these groups. Types of malformations observed in these groups were abnormal tail, spina bifida and deformity of vertebrae, and were almost similar to those observed by maternal subcutaneous injection of TB. 4) A trace of TB was found microscopically in the frozen section of embryo after both subcutaneous and intraexocoelom injections of TB. These results suggest that TB acts directly on embryos to produce malformations.

Abnormalities, Drug-Induced↗
PubMed2006

Hypothermia protects cultured human retinal pigment epithelial cells against trypan blue toxicity.

PURPOSE: To determine whether trypan blue (TB) is toxic to cultured human retinal pigment epithelial (ARPE-19) cells, and whether hypothermia can protect ARPE-19 cells against TB toxicity. METHODS: ARPE-19 cells were cultured and exposed to balanced salt solution as a control, while other cells were exposed to 0.05, 0.2, and 0.5% TB dye at 37 and 4 degrees C for 5 and 30 min. The percentage of ARPE-19 cells that survived was determined by resazurin 1 day after the exposure. RESULTS: A statistically significant decrease in the percentage of ARPE-19 cells surviving was found after exposure to 0.2 and 0.5% TB at any temperature or for any exposure duration (p < 0.01). The percentage of RPE cells surviving at 0.05% was not significantly different from that of controls except for a 30-min exposure at 37 degrees C. The percentage of cells surviving at 4 degrees C for a 5-min exposure to 0.5% TB and a 30-min exposure to 0.2 and 0.5% was significantly higher than that at 37 degrees C under each condition (p < 0.01 for all). CONCLUSIONS: These results indicate that TB is toxic to human RPE cells, and the toxicity is dose- and exposure duration-dependent. Exposing the cells at 4 degrees C had a protective effect against higher concentrations or longer exposure durations of TB compared to exposure at 37 degrees C.

Cell Count↗
PubMed2006

Efficacy of trypan blue in posterior capsulorhexis with optic capture in pediatric cataracts [ISRCTN48221688].

BACKGROUND: To evaluate the efficacy of trypan blue (0.06%) in posterior capsulorhexis with optic capture in pediatric cataracts. METHODS: In this prospective randomized controlled study, trypan blue dye assisted posterior capsulorhexis with optic capture was performed in 18 eyes (group 1) and no dye was used for posterior capsulorhexis (group 2) in 17 eyes. RESULTS: The mean size of the posterior capsulorhexis was 4.6 +/-1.77 mm and 4.0 +/- 0.93 mm in the group 1 and 2 respectively. Optic capture was possible in 17 eyes in the group 1 and 11 eyes in the group 2. CONCLUSION: Trypan blue facilitates posterior capsulorhexis with optic capture of AcrySof IOL in cases of pediatric cataracts.

Acrylates↗
PubMed2005

A prospective study on trypan blue capsule staining under air vs under viscoelastic.

AIM: To compare the safety and effectiveness of trypan blue capsule staining under air vs under viscoelastic. METHODS: A total of 52 consecutive patients planned for phacoemulsification of white mature cataract were randomly assigned to trypan blue staining under air or under viscoelastic. Perioperative changes in best-corrected visual acuity (BCVA), central corneal thickness (CCT), and endothelial cell density (ECD) were compared between the two groups. The differences in operating and phacoemulsification times, staining pattern, and complications between the two groups were also recorded. RESULTS: Phacoemulsification of white mature cataract was performed in 50 (96%) eyes. The median preoperative BCVA was hand movement. No significant differences in the baseline characteristics were found between the two groups. At 3 months after phacoemulsification, the median BCVA improved to 0.8. The mean CCT returned to preoperative level by 1 month postoperatively and the mean ECD loss was 11.9% 3 months postoperatively. No significant differences in median BCVA, mean phacoemulsification and operation times, mean CCT, and mean ECD were found between the two groups. CONCLUSIONS: Trypan blue staining of the anterior lens capsule under air or under viscoelastic were similarly effective and safe methods for the phacoemulsification of white mature cataract.

Aged↗
PubMed2005

[Role of trypan blue in epiretinal membrane surgery].

INTRODUCTION: Idiopathic epiretinal membrane (ERM) results from detachment of the posterior hyaloid. Vitrectomy and peeling are the treatment of choice. Trypan blue 0.15% (TB) stains the epiretinal membrane and the internal limiting membrane (ILM), facilitating surgery and thus allowing selective and complete removal with less retinal damage. We conducted a study evaluating the advantages of selective staining associated with surgery, the incidence of postoperative damage, and the influence on final visual acuity. MATERIAL AND METHODS: Fourteen patients were operated on by the same surgeon, who carried out a complete vitrectomy, then fluid-air exchange for dye injection of undiluted 0.2 ml of Trypan blue 0.15% in front of the membrane, which was then easily distinguished from the area under the retina. Preoperative visual acuity, postoperative damage, as well as eye status at the end of follow-up are discussed. Transmission electron microscopy was used to determine if epiretinal membrane removal was complete and selective. Angiography was systematically used to study the state of retina before and after surgery. RESULTS: Preliminary results show that trypan blue is useful and facilitates complete removal of ERM and ILM. It may reduce long-term complications and recurrent membrane formation. Trypan blue is a staining agent that is quick and easy to use. It presents no short-term toxic effects. Functional results in 6 months show improvement of visual acuity and disappearance of metamorphopsia. CONCLUSION: Idiopathic epiretinal membrane surgery with tryptan blue staining is a better alternative than indocyanine green staining.

Aged↗
PubMed2006

Trypan-blue-assisted anterior continuous curvilinear capsulorhexis in a case of ocular pemphigoid.

PURPOSE: To report on trypan-blue-assisted anterior continuous curvilinear capsulorhexis (ACCC) in a case of ocular pemphigoid. METHODS: Interventional case report. RESULTS: Due to the reduced visibility especially in the corneal periphery caused by the ocular pemphigoid, trypan blue 0,06% (Acri.Blue) was used to stain the anterior capsule of the lens. Then anterior continuous curvilinear capsulorhexis was performed. Due to the blue staining, the visualization of the margin of the rhexis was always good and phacoemulsification procedure was successfully performed afterwards. During the follow-up period of 12-months-postsurgically, no exacerbation of the ocular pemphigoid occurred. CONCLUSIONS: The use of trypan blue staining of the anterior capsule enabled the surgeon to perform a safe anterior continuous curvilinear capsulorhexis and a subsequent phacoemulsification in ocular pemphigoid. No progression of the ocular pemphigoid was seen within the 12-months-post-surgery period.

Capsulorhexis↗
PubMed2006

Trypan blue to aid visualization of the vitreous during anterior segment surgery.

We describe the usefulness of intracameral injection of trypan blue to highlight vitreous in the anterior chamber. The technique was used in 3 eyes of 3 patients who had vitreous prolapse in the anterior chamber. Trypan blue 0.06% was injected in the anterior chamber under air to visualize and resect the prolapsed vitreous. Trypan blue stained the clear vitreous and improved visualization for a safe and atraumatic vitrectomy. No complications associated with the dye were found.

Adolescent↗
PubMed2002

Trypan blue uptake by chinese hamster ovary cultured epithelial cells: a cellular model to study Escherichia coli STb enterotoxin.

The thermostable enterotoxin b (STb) produced by enterotoxigenic Escherichia coli strains is responsible for diarrheal diseases mainly in weaning piglets. For now, the only available assay for biological activity of STb toxin was in the animal host (i.e. piglet) or in an animal model (i.e. rat, mouse). In this study, we developed a cellular model for the study of the biological activity of STb enterotoxin. Using a trypan blue vital stain method, we showed that STb-treated cells of three out of the five cell lines tested absorbed more vital stain than their controls. Of all the cell lines tested, the chinese hamster's ovary derivated cells (CHO) were the most sensitive, absorbing 50% more trypan blue than their control. Maximal stain uptake was observed after 2h. We then evaluated the trypan blue uptake for 16 STb mutants, produced in a previous work, on the CHO cell lines in order to compare it with the in vivo rat loop assay data. Interestingly, we observed a good correlation between the two bioassays. In fact, the biological activity observed in the rat could be correlated with the trypan blue uptake by the CHO cells (R(2)=0.78) for STb toxin and the 16 mutants. Using the variance analysis statistical test, we determined that the correlation between the two bioassays is significant (F(c)> or =F(0.005)). These results suggest that the trypan blue uptake bioassay could represent a new method to evaluate the biological activity and facilitate the elucidation of the mechanism of action of E. coli STb enterotoxin.

Amino Acid Sequence↗
PubMed1967

Lysosomal enzyme inhibition by trypan blue: a theory of teratogenesis.

A mechanism of mammalian teratogenesis involving inhibition of embryotrophic nutrition is suggested and exemplified by the action of trypan blue on pregnant rats. There is evidence for the localization of trypan blue in heterolysosomes of the epithelium of the visceral yolk sac, and our experiments indicate that the dye is an inhibitor of a selection of hydrolytic enzymes present in lysosomal fractions from homogenates of rat visceral yolk sac. It seems likely that trypan blue inhibits the intracellular digestion of embryotroph by the visceral yolk-sac epithelium; the conceptus may therefore be deprived of essential nutrients at critical stages of development.

Abnormalities, Drug-Induced↗