Rabbit anti stat3 79d7

Retina and RPE-choroid were lysed in RIPA buffer with protease inhibitor cocktails and phosphatase inhibitors (Sigma-Aldrich). The protein concentration was determined by BCA kit (Thermo Scientific). Thirty microgram protein was loaded on a 10% SDS-PAGE gel and transferred to an Immobilon-FL PVDF membrane (Millipore, Watford, UK). The membranes were incubated sequentially with rabbit anti-STAT3 (79D7, 1:1000) or rabbit anti-pSTAT3 (Tyr705, D3A7, 1:1000) (both from Cell Signalling, Danvers, MA, USA), followed by goat anti-rabbit IgG (1:10,000, Li-COR Biosciences, Cambridge, UK). The membranes were imaged with Odyssey infrared imaging system (Li-COR Biosciences), and analysed by ImageStudioLite Software (Li-COR Biosciences). The mouse anti-β actin (C4, 1:10,000, Santa Cruz, Dallas, Texas, USA) was used as a housekeeping control protein.