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B. Ruiz-Perez, T. Dowie, R. Gee, P. Isom, M. Manzo, E. Skokanova; Transscleral Delivery of a 12.4 kDa Protein by Ocular Iontophoresis. Invest. Ophthalmol. Vis. Sci. 2008;49(13):1813.
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To test delivery of cytochrome C, a 12.4 kDa protein, into the eye by iontophoresis.
For in vitro testing, anodal iontophoresis was performed using a two chamber Ussing apparatus. 40 mg/mL of cytochrome C in H2O was used in the donor chamber and phosphate buffered saline solution was used in the receptor chamber. Visual inspection and the Agilent Bioanalyzer® were used to detect the cytochrome C. For ex vivo testing, cadaver adult rabbit eyes were dosed with 40 mg/mL of cytochrome C using the EyeGate II device for 15 and 20 mA*mins. Tissues were harvested and a qualitative measurement of cytochrome C was obtained via its chromophore on a UV spectrophotometer at 220 nM. For in vivo testing, four rabbits were dosed with 40 mg/mL of cytochrome C using the EyeGate II device; two rabbits received 15 mA*mins (anodal iontophoresis) while the two control rabbits received 0 mA*mins (passive). Eyes from control and treatment groups were harvested immediately after dosing and 2 hours post-dosing.
In vitro experiments showed a slight red coloring of the cytochrome C in the passive receptor chambers; red coloring was observed to be more intense in receptor chambers after anodal iontophoresis was applied. 3% of the cytochrome C was delivered by iontophoresis while only 0.27% of the protein was delivered passively. Additionally, after anodal iontophoresis in cadaver eyes, cytochrome C was detected in all ocular tissues harvested. Both the 15 and 20 mA/mins treated eyes showed similar distribution of cytochrome C between tissues with the exception of the conjunctiva which showed slightly higher levels at 20 mA/mins. Moreover, in the in vivo rabbit model, the red coloring of the cytochrome C was clearly observed within each iontophoretically treated eye while no red color was observed in the passively treated eyes.
These data indicate successful transscleral delivery of the 12.4 kDa cytochrome C protein. Additionally, in vitro results show an 11.2 fold increase in iontophoretic delivery over passive delivery alone.
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