[Production of recombinant humanized anti-HBsAg Fab antibody by fermentation].
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.
Life Science and Technology College in Jinan University, Guangzhou 510632, China.
Ning
N
Deng
Jun-Jian
JJ
Xiang
Wen-Yin
WY
Chen
Sheng
S
Xiong
Xun-Zhang
XZ
Wang
Kuan-Yuan
KY
Su
D005285:Fermentation; D006510:Hepatitis B Antibodies / Q000096:biosynthesis* / Q000302:isolation & purification; D006514:Hepatitis B Surface Antigens / Q000276:immunology*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000302:isolation & purification; D010843:Pichia / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification
D006510:Hepatitis B Antibodies; D006514:Hepatitis B Surface Antigens; D007140:Immunoglobulin Fab Fragments; D011994:Recombinant Proteins