At days 0, 3, 7, and 14 after laser, each whole retina was dissected from the RPE-choroid complex and protein from these two tissue complexes were isolated separately (n = 3 animals from each group at each time point). Samples were sonicated in lysis buffer (20 mM imidazole HCl, 10 mM KCl, 1 mM MgCl2, 10 mM EGTA, 1% Triton X-100, 10 mM NaF, 1 mM Na molybdate, and 1 mM EDTA with protease inhibitor; Sigma-Aldrich) on ice. VEGF protein levels in the supernatant were determined with the use of an ELISA kit (threshold of detection 3 pg/mL; R&D Systems, Minneapolis, MN) that recognizes all splice variants. Readings were taken with the spectrophotometer at 450 to 570 nm (Emax; Molecular Devices, Sunnyvale, CA) and were normalized to total protein (Bio-Rad, Hercules, CA). Duplicate measurements were performed.