Preparation of mouse anti-recombinant SAG1 antigen monoclonal antibody of Toxoplasma gondii by intrasplenic immunization
- VernacularTitle:小鼠脾内免疫法制备重组弓形虫SAG1抗原单克隆抗体
- Author:
Nan WU
;
Danqiu ZHOU
;
Wei RUAN
;
Ligui WU
;
Huizhang ZHANG
- Publication Type:Journal Article
- Keywords:
rSAG1;
intrasplenic immunization;
monoclonal antibody;
sandwich-ELISA;
mice
- From:
Fudan University Journal of Medical Sciences
2010;37(2):184-188
- CountryChina
- Language:Chinese
-
Abstract:
Objective To prepare monoclonal antibody in mice so as to develop an ELISA method for diagnosis of Toxoplasma gondii infection during the initial stage. Methods The mice were immunized by combining routine and intrasplenic immunization with recombinant SGA1 antigen. B lymphocyte hybridization technique was applied to prepare the anti-SAG1 McAbs. Positive clones were screened using ELISA and subcloned to establish cell lines. Ascites was induced to produce the McAbs. Then the McAbs were purified by protein G chromatograph column. The specificity of McAbs was identified by Western blot and sandwich-ELISA. Sensitivity of the McAbs was determined using sandwich-ELISA. Comparasion was carried out between PCR and sandwich-ELISA method. Results Two positive clones were obtained and named as 3B6, 10C4, both could identify the native and recombinant SAG1 antigens. The sensitivity of 3B6, 10C4 was 31.3 ng and 62.5 ng, respectively. There was no cross reaction between the McAbs and positive sera from patients with schistosomiasis, ancylostomiasis or malaria. By using PCR and ELISA, the positive infection rate of T. gondii was 63.2% and 47.4%, respectively. Conclusions Therefore, mouse anti-rSAG1 antigen McAbs have been prepared successfully and primarily applied to early stage diagnosis of T. gondii infection.
