Application of colloidal gold immunoassay strips for the rapid detection of potato Y virus

Abstract

Potato Y virus (PVY) is a bacterial virus that seriously jeopardizes the growth of tobacco. In order to achieve rapid detection of PVY, monoclonal antibodies to PVY-CP protein were prepared and characterized, and colloidal gold immunochromatographic test strips that can be used to detect PVY were established. In this study, we constructed the PVY-CP protein expression plasmid pET28a-PVY-CP, transformed it into Escherichia coli BL21 (DE3) receptor cells to induce the expression of the target protein, and then further purified it as an immunogen to screen hybridoma cells that can stably secrete monoclonal antibody against PVY through cell fusion with hybridoma cells, prepared ascites, and purified the monoclonal antibody by using the caprylic acid saturated ammonium sulfate method. The monoclonal antibody was purified using the method of ammonium octanoate saturated sulfate, labeled with colloidal gold, and the colloidal gold immunochromatographic test strip was established by optimizing the reaction conditions, and the detectability, accuracy and specificity of the test strip were evaluated. The detection limit of the test strip for PVY-CP protein was 1 μg/mL, and there was no cross-reactivity with tobacco bunchy top virus, tomato spotted wilt virus, and tobacco mosaic virus. Comparison of the prepared colloidal gold test strips and the RT-PCR method for actual samples showed that the total conformity rate of the two was 86.67%, and the positive conformity rate was 90%.