A Remarkable Tale: Increased Retinal Fluorescence Following Application of Permanent Marker Ink to Mouse Tails.

Purpose: Unusually high retinal fluorescence was observed during in vivo imaging among a few mice in our colony. Their tails had been marked with red Sharpie ink, a common practice for identification. They were observed licking the ink off their tails, suggesting possible oral intake of a fluorescent ink component that might accumulate in the retina. One component of red marker ink is the fluorophore Rhodamine 6G (R6G), suggesting that it might be responsible. Methods: We tested this hypothesis by tail marking C57Bl/6J mice with one line or four lines of red ink. Retinal fluorescence was monitored in vivo with confocal scanning laser ophthalmoscope (cSLO) imaging at baseline and from 6 hours to 2 weeks after marking. Retinas and brains were sectioned for histology. A second cohort of mice received an oral gavage of R6G at the concentration estimated in red ink, and retinal histology was performed. Results: Tail marking with red ink led to a transient, dose-dependent increase in retinal fluorescence in cSLO images, peaking at 6 hours. Histology revealed that this fluorescence was predominantly in the photoreceptor outer segments. This fluorescence localization was reproduced with oral R6G alone. Fluorescence in the brain was concentrated in the choroid plexus and ventricles. Conclusions: These results suggest that R6G can cross the gut-blood, blood-retina, and blood-cerebrospinal fluid barriers. This raises important technical considerations for experimental mouse identification and has potential applications for fluorescence imaging and central nervous system drug targeting.