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[High-level expression of 6B11 ovarian carcinoma anti-idiotypic antibody scFv genes in E. Coli]
1Gynecologic Oncology Center, People's Hospital Beijing Medical University, Beijing 100034.
Objective:
To express high-level 6B11 ovarian carcinoma anti-idiotypic antibody single chain Fv (scFv) genes in E. Coli.
Methods:
Using PCR cloning technique, We cloned 6B11scFv genes into bacterial expression vector pKPL-3a. Recombinant plasmid vector pL-6B11scFv was transformed into E. Coli pop2136 with temperature control to produce proteins. Renaturated proteins were purified on DEAE-Sepharose Fast Flow ion exchange column with salt gradient elution. Immunoactivity was determined with ELISA and inhibition ELISA tests respectively.
Results:
6B11scFv genes were expressed as inclusion bodies in the cytoplasm. The purity of 6B11scFv turned out by SDS-PAGE was more than 95%. The expressed proteins after purification specifically reacted with anti-ovarian carcinoma monoclonal antibody COC166-9 and efficiently inhibited the reaction between primary ovarian carcinoma antigen OC166-9 and COC166-9.
Conclusion:
We successfully expressed high-level 6B11scFv genes in E. Coli. Expressed proteins showed pretty good immunoactivity.
