All corneas were pretreated with 0.1% RB solution for 2 minutes. Corneas were irradiated with green light using a cw KTP frequency (continuous wave potassium titanyl phosphate crystal) doubled solid-state laser (Oculight OR; IRIDEX Corporation, Mountain View, CA, USA) at a wavelength of 532 nm that is strongly absorbed by the RB. The laser light from an optical fiber was expanded to a 1.2-cm-diameter beam to deliver an irradiance of 0.25 W/cm
2 on the central cornea. Corneas were treated for 3.25, 6.5, 10, or 13.25 minutes (to deliver fluences of 50, 100, 150, or 200 J/cm
2, respectively) according to the protocol of Cherfan et al.
2 This protocol was shown not to cause toxicity to stromal keratocytes ex vivo and can be used to stiffen corneas thinner than 400 μm.
2 Thirty-five enucleated rabbit eyes were divided into the following subgroups: (1) nontreated control group (five eyes), (2) RB-stained control group (RB, five eyes), (3) 50-J/cm
2 RGX group (RB50, five eyes), (4) 100-J/cm
2 RGX group (RB100, five eyes), (4) 150-J/cm
2 RGX group (RB150, five eyes), (6) 200-J/cm
2 RGX group (RB200, five eyes), and (7) 3-mW/cm
2 RF-UVA group (five eyes).