Comparative Toxicology of Trypan Blue, Brilliant Blue G, and Their Combination Together with Polyethylene Glycol on Human Pigment Epithelial Cells

PURPOSE. To determine the toxicity in ARPE-19 human retinal pigment epithelium cells of trypan blue (TB) at 0.15% and 0.25% concentration, brilliant blue G (BBG) at 0.025% and 0.05%, their combination, and the effect of the addition of 4% polyethyleneglycol (PEG), as an additive for increasing the density and thus improving the staining in internal limiting membrane removal, on the individual dyes and their combinations, and compare the toxicity of the dyes to that of clinically used preparations. METHODS. Cells were exposed for 5 and for 30 minutes to the different preparations. Cell viability was measured with the WST-1 assay measuring intracellular dehydrogenase activity. RESULTS. Solutions containing PEG with BBG (0.025%), TB (0.15%), and mixtures of BBG (0.025%) with TB (0.15% and 0.25%) were the least toxic of the preparations as well as preparations of BBG at 0.025% in phosphate-buffered saline solution, while TB at 0.25% in phosphate-buffered saline solution was the most toxic. The addition of PEG reduced the toxicity of preparations containing TB either alone or in combination with BBG. These results were seen only after an incubation for 30 minutes; for a 5-minute incubation, no toxicity was seen for any of the preparations. CONCLUSIONS. For short incubation times, all dyes appear equally safe. For longer incubation times, TB preparations were more toxic than BBG preparations. The toxicity of TB was reduced by the addition of PEG. Further studies are required to determine the clinical impact of this finding. (Invest Ophthalmol Vis Sci. 2011;52:4085-4090) DOI:10.1167/iovs.10-6336