Monoclonal Antibodies Against the Whitefly-Transmitted Tomato Yellow Leaf Curl Virus and Their Application in Virus Detection

Abstract

Tomato yellow leaf curl virus (TYLCV) is a species of the family Geminiviridae, causing serious yield losses in tomato production. The coat protein (CP) gene of TYLCV isolate SH2 was expressed in Escherichia coli BL21 (DE3) using pET-32a as the expression vector. The recombinant protein was purified through Ni+-NTA affinity column and used to immunize BALB/c mice. Three hybridoma cell lines (2B2, 2E3 and 3E10) secreting monoclonal antibodies (MAbs) against TYLCV CP were obtained by fusing mouse myeloma cells (Sp 2/0) with spleen cells from the immunized BALB/c mouse. The titers of ascitic fluids of three MAbs ranged from 10−6 to 10−7 in indirect-ELISA. Isotypes and subclasses of all the MAbs belonged to IgG1, k light chain. Triple antibody sandwich enzyme-linked immunosorbent assay (TAS-ELISA) showed that the MAb 3E10 could react with five begomoviruses infecting tomato, while the other two (2B2 and 2E3) mainly reacted with TYLCV. TAS-ELISA was set up using the MAb 3E10, and the established method could successfully detect virus in plant sap at 1:2 560 (w/v, g mL−1). Detection of field samples showed that begomoviruses were common in tomato crops in Zhejiang Province, China.