Ab193761

The levels of HSP70 and midkine in human sera and synovial fluid from healthy individuals and patients with OA, in conditioned medium and cell lysates of human C28I2 cells treated with 25 nM ProTx II or 1 µM PF-04856264 were measured by ELISA according to the manufacture’s instructions (Abcam, ab133060, ab193761). Before ELISA analysis, human synovial fluids were digested with hyaluronidase at 1 unit per 100 µl synovial fluid for 1 h at 37 °C. The levels of PRG4 and MMP13 in supernatant of human OA cartilage explants were measured by ELISA according to the manufacturer’s instructions, respectively (R&D systems, DY9829-05; Abcam, ab100605). The levels of HSP70 and midkine in mouse sera and conditioned medium collected from primary chondrocytes were measured by ELISA according to the manufacture’s instructions (Abcam, ab133061, ab279416).

The levels of midkine, IL-10, and IFN-γ in serum samples or conditional medium were evaluated by ELISA kit (Abcam # ab193761, # ab279416, # ab185986, #ab255729 and #ab174443, USA) according to manufacturer’s instructions. Briefly, samples were incubated with antibody-coated plates for 1 h at room temperature and washed for five times. Then plates were incubated with biotinylated antibody, streptavidin antibody, substrate solution, and stop solution consecutively. The absorbance at 450 nm was measured by a microplate reader. All samples were done in triplicates.