Preparation and preliminary utilization of monoclonal antibodies specifically against an immunogenic fragment in ectodomain of prostate-specific membrane antigen (PSMA)
10.3969/j.issn.1000-484X.2016.02.013
- VernacularTitle:抗PSMA膜外域单克隆抗体的制备及初步应用
- Author:
Rong TAO
;
Zhenhua NI
;
Sheng YANG
;
Chong LIU
;
Min ZHU
;
Jiangfan SHEN
;
Shaohua TU
- Publication Type:Journal Article
- Keywords:
Prostate specific membrane antigen ( PSMA);
Monoclonal antibody;
Prostate cancer
- From:
Chinese Journal of Immunology
2016;(2):205-209
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To prepare monoclonal antibodies specifically against an immunogenic fragment in ectodomain of prostate-specific membrane antigen ( PSMA ).Methods: An polypeptide immunogenic fragment in the ectodomain of PSMA was predicted by biological information technology,and then it was expressed prokaryotically.BALB/c mice were immunized with the prokar-ytically expressed recombinant polypeptide antigen,to prepare the monoclonal antibodies specifically against an immunogenic fragment in ectodomain of PSMA by hybridoma technology,purification of monoclonal antibody by affinity chromatography,characterization of the monoclonal antibodies by Western blot.The radioimmunoimaging in prostate cancer model was performed by using the labeled McAb.Results:Throught the software analysis,we got the antigen fragment in the ectodomain of PSMA containing 310aa sequences higher specificity, artificially synthesized gene sequence of the region, and constructed a prokaryotic expression vector pET-32a-r-ectodomain-PSMA,by prokaryotic expression we obtained the 50 kD target antigen,after hybridization,the three positive hybridoma cell lines (5E6,4A5 and 4D7) were selected by ELISA using target antigen,the isotypes of 5E6 and 4A5 were IgG2a,the isotypes of 4D7 were IgG1,the titer of three monoclonal antibodies was above 1∶256 000.Western blot results showed that the prepared monoclonal anti-bodies could binding specifically to the antigen in the ectodomain of PSMA.Radioimmunoimaging in prostate cancer animal model results further confirm that the prepared monoclonal antibodies could combinate with the antigen in the ectodomain of PSMA in the animal body, and make the tumor imaging.Conclusion: The prepared monoclonal antibodies can specifically recognizes the PSMA antigen,which laid the foundation for the immunodiagnosis and immunotherapy of prostate cancer.
