Nonlinear optical corneal collagen crosslinking of ex vivo rabbit eyes

Purpose To determine whether riboflavin-induced collagen crosslinking (CXL) could be precisely achieved in the corneal stroma of ex vivo rabbit eyes using nonlinear optical excitation with a low numerical aperture lens and enlarged focal volume. Setting Gavin Herbert Eye Institute, University of California Irvine, Irvine, California, USA. Design Experimental study. Methods The corneal epithelium was removed and the corneas were soaked in 0.5% riboflavin solution. Using a 0.1 numerical aperture objective, a theoretical excitation volume of 150 μm × 3 μm was generated using 1 W of 760 nm femtosecond laser light and raster scanned with 4.4 μm line separation at varying effective speeds over a 4.50 mm × 2.25 mm area. Corneal sections were examined for collagen autofluorescence. Results Collagen autofluorescence was enhanced 2.9 times compared with ultraviolet-A (UVA) CXL. Also, increasing speed was linearly associated with decreasing autofluorescence intensity. The slowest speed of 2.69 mm/s showed a mean of 182.97 μm ± 52.35 (SD) long autofluorescent scan lines axially in the central cornea compared with 147.84 ± 4.35 μm for UVA CXL. Conclusions Decreasing dwell time was linearly associated with decreasing autofluorescence intensity, approaching that of UVA CXL at a speed of 8.9 mm/s. Using an effective speed of 8.9 mm/s, nonlinear optical CXL could be achieved over a 3.0 mm diameter area in fewer than 4 minutes. Further development of nonlinear optical CXL might result in safer, faster, and more effective CXL treatments. Financial Disclosure None of the authors has a financial or proprietary interest in any material or method mentioned.