2.11. Western immunoblotting

Cells or tissues (isolated glomeruli) were collected with RIPA buffer, and protein concentration was determined by bicinchoninic acid (BCA) assays (Beyotime, China). Equal amounts of protein (30 μg per lane) were separated by SDS‐PAGE and then transferred to PVDF membranes (Millipore Corp, USA). The membranes were then incubated with a primary antibody (LRH‐1: 1:500, GLS2: 1:1000, GAPDH: 1:5000) in 5% milk overnight at 4°C. Next, the membranes were labelled with an Alexa Fluor 680/790‐labelled (1:10,000, LI‐COR Biosciences, USA) goat anti‐mouse/goat anti‐rabbit secondary antibody, followed by scanning via an LI‐COR Odyssey Infrared Imaging System.