Calpain 1, Large Subunit (CAPN1) is an intracellular protease that requires calcium for its catalytic activity. Calcium-regulated non-lysosomal thiol-protease which catalyze limited proteolysis of substrates involved in cytoskeletal remodeling and signal transduction. It has broad endopeptidase specificity. Besides, Signal Transducer And Activator Of Transcription 3 (STAT3) has been identified as an interactor of CAPN1, thus a binding ELISA assay was conducted to detect the interaction of recombinant rat CAPN1 and recombinant rat STAT3. Briefly, CAPN1 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to STAT3-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-CAPN1 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50μL stop solution to the wells and read at 450nm immediately. The binding activity of of CAPN1 and STAT3 was shown in Figure 1, and this effect was in a dose dependent manner.