Il 18

The release rates of LDH (C0016; Beyotime Biotechnology Co., Ltd.), Scr (C011-2; Nanjing Jiancheng Bioengineering Institute, Nanjing, Jiangsu, China) and BUN (C013-2; Nanjing Jiancheng Bioengineering Institute) were measured with corresponding test kits; IL-1β (SEA563Mu; Wuhan USCN Business Co., Ltd., Wuhan, China) and IL-18 (SEA064Mu; Wuhan USCN Business Co., Ltd.) were measured with enzyme-linked immunosorbent assay kits; and caspase-1 activity was measured with the Caspase 1 Activity Assay kit (C1101; Beyotime Biotechnology Co., Ltd.). All tests were performed according to the manufacturer’s instructions.

The ELISA was performed according to the manufacturer's instructions. Briefly, microtiter plates pre-coated with a monoclonal antibody against human KIM-1 (#CSB-E08807H, Cusabio, China), NGAL (#KIT036, Bioporto, Denmark), IL-18 (E90064Hu, USCN Life Science, Houston, TX, USA) or L-FABP (E91566Hu, USCN Life Science) were added with 100 µL of urine samples or standards for 1 hr at 37℃. After removed the liquid, each well was incubated with a 100 µL biotinylated monoclonal antibody for 1 hr at 37℃. The solution was aspirated and washed 3 times, followed by 100 µL avidin-conjugated HRP treatments for 1 hr at 37℃. After aspirate and washing 5 times, 100 µL tetramethylbenzidine substrate was added for color development in the dark room, which was read after 10-30 min at 450 nm with ELISA reader (Benchmark Plus, BioRad, Hercules, CA, USA). All measurements were made in triplicate.