Establishment and application of blocking ELISA method for detection of against Duck reovirus σC antibody

Duck reovirus disease is a duck-transmitted disease characterized by homorrhage and necrosis of the livers and spleens. Blocking ELISA has high sensitivity and simple operation, which can be used as a serological test for duck infection, to improve the detection efficiency of DRV infection. Duck reovirus σC protein is the main protein involved in the body's immune system, which can induce the production of neutralizing antibodies. Currently, there is no established blocking ELISA detection method based on this protein. This study aimed to produce and characterize σC monoclonal antibodies with the ultimate goal of developing a monoclonal antibody-based blocking ELISA for DRV antibody detection. This established detection method had no response to the serum after infection with ARV and MDRV, so it can also be used to distinguish DRV from ARV and MDRV infections. To evaluate the performance of the detection method, 87 serum samples (53 negative and 34 positive samples) were analyzed and receiver-operating characteristic (ROC) analysis was applied to determine the Cut-Off value. According to ROC analysis, the area under the curve (AUC) was 0.9789 (95% confidence interval: 95.13 to 100%), the Cut-Off value was set to 15.49%. The coefficients of inter- and intra-batches were < 10%, indicating the good repeatability of the method. The maximum dilution of positive standard serum detected by this blocking ELISA method was 1:512 and did not cross-react with other duck-derived viruses, illustrating strong specificity. In conclusion, the σC mAb-based blocking ELISA detection method established in this study has high sensitivity and specificity, which provides a reliable and useful tool for on-site surveillance and epidemiological studies in duck flocks.