Production of recombinant humanized anti-HBsAg Fab antibody by fermentation.
- Author:
Ning DENG
1
;
Jun-Jian XIANG
;
Wen-Yin CHEN
;
Sheng XIONG
;
Xun-Zhang WANG
;
Kuan-Yuan SU
Author Information
1. Life Science and Technology College in Jinan University, Guangzhou 510632, China.
- Publication Type:Journal Article
- MeSH:
Fermentation;
Hepatitis B Antibodies;
biosynthesis;
isolation & purification;
Hepatitis B Surface Antigens;
immunology;
Humans;
Immunoglobulin Fab Fragments;
biosynthesis;
isolation & purification;
Pichia;
genetics;
Recombinant Proteins;
biosynthesis;
isolation & purification
- From:
Chinese Journal of Biotechnology
2004;20(5):800-804
- CountryChina
- Language:Chinese
-
Abstract:
In order to produce recombinant human anti-HBsAg Fab antibody in Pichia pastoris, the recombinant yeast was fermented using fed-batch system in a 30 L bioreactor. The fermentation temperature was 30 degrees C, the pH was 5.0 approximately 5.3, and the DO was 20% approximately 30%. The recombinant Fab antibody was purified from crude culture supernatant by ion exchange and analyzed by SDS-PAGE and western blot and ELISA. When the absorbance (OD600) of broth reach 300 at the end of fed-batch phase, the induced phase was initiated. The results showed that recombinant human anti-HBsAg Fab antibody was high-level expressed in recombinant Pichia pastoris using a fed-batch fermentation system. Both chains of the Fab were successfully expressed upon methanol induction. After 192 h of induction, the expression level of recombinant Fab (soluble) reached 412 mg/L. The recombinant Fab antibody was purified effectively by ion-exchange chromatography from the fermentation supernatant to a purity of 95%. And the affinity activities of the purified recombinant Fab antibdy and fermentation supernatant were detected, and both of them showed high affinity activities. The results demonstrated that recombinant human anti-HBsAg Fab antibody could be high level produced by fed-batch fermentations in Pichia pastoris. Which can be efficiently used in industrial production.
