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

Am J Ophthalmol. 2004 Jul;138(1):64-9. doi: 10.1016/j.ajo.2004.02.061.


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%).

Publication types

  • Comparative Study

MeSH terms

  • Cell Line
  • Cell Survival / drug effects
  • Coloring Agents / toxicity*
  • Dose-Response Relationship, Drug
  • Humans
  • Indocyanine Green / toxicity*
  • Light
  • Osmolar Concentration
  • Pigment Epithelium of Eye / drug effects*
  • Pigment Epithelium of Eye / radiation effects
  • Time Factors
  • Trypan Blue / toxicity*


  • Coloring Agents
  • Trypan Blue
  • Indocyanine Green