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C. M. Hutnik, D. Yuen, H. Liu, A. Proulx, J. Gonder; A Comparison of the in vitro Safety of Intraocular Dyes. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4884.
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Another group recently reported that Brilliant blue G (BBG) was a safe dye for for internal limiting membrane (ILM) peeling in vitreoretinal surgery based upon an in vivo animal study. In vitro toxicity wasnot reported.The purpose of the present study was tocompare the in vitro toxicity of BBG to indocyanine green (ICG), the current gold standard, as well as to trypan blue (TB) and evans blue (EB). A demonstration of in vitro safety would further support a potential paradigm shift in the first choice of adjuvant dye used for ILM peeling.
The toxicity of four dyes was tested in two different cell cultures. These were a commercially available human retinal pigment epithelial cell line known as ARPE-19 and a primary mixed murine retinal ganglion/ muller glial cell culture (RGC). The dose-dependent toxicity of the dyes was determined by exposing the two different cell cultures to each dye, at four different concentrations, for a short exposure time of 3 minutes. In another experiment a medium exposure time of 30 minutes was performed. Cell viability was measured using the MTT assay. Time-dependent toxicity of the dyes was also studied. All four dyes, each diluted to 1/500th of stock concentration, were applied to the ARPE-19 cells for a prolonged exposure, to simulate the possible long lasting persistence of dye in the vitreous cavity after vitreoretinal surgery. Cell viability was measured at the 2, 24, 48 and 72 hour time point using the MTT assay. Finally, the possible influence of osmolarity was assessed.
After 3 minutes of dye exposure, only BBG caused significant loss of ARPE-19 cell viability. Longer exposure times of 30 minutes resulted in toxicity from both BBG and TB, with the toxicity of BBG being greater than that of TB. Prolonged exposure at 1/500th of stock concentration of either BBG or ICG produced equivalent loss of cell viability. None of the dyes produced any significant loss of RGC/glial cell viability after 3 minutes of exposure. However, 30 minutes of exposure to BBG, TB and EB demonstrated toxicity, with the toxicity of BBG being greater than that of TB and EB. Osmolarity was not a factor in any of the experiments.
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