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Updated: May 4, 2026
Scalable High Throughput Selection From Phage-displayed Synthetic Antibody Libraries
Published on: January 17, 2015
[Construction and identification of a full-length human secretory antibody library in Pichia pastoris]
Qiang Liu1, Jing Zhang1, Xiaoai Zhang2
1Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, PLA Academy of Military Medicine, Beijing 100850, China.
Objective:
To clone and express functional, recombinant full-length human antibodies in partial N-glycoengineered Pichia pastoris yeast.
Methods:
We constructed double promoter series expressing the constant regions of the light and heavy chain genes using the secretory expression vector pPICZαA; the obtained vector was named pPICZαA-CH;-CL;. Human peripheral blood mononuclear cells (PBMCs) were isolated and RT-PCR was performed using 44 pairs of degenerate primers to generate variable region genes of the heavy and light chains (VH; and VL;). Then, they were inserted into the pPICZαA-CH;-CL; vector. The full-length human antibody library pPICZαA-VH;-CH;-VL;-CL; was transformed into partial N-glycoengineered Pichia pastoris, GS115Y, by electroporation. Sequencing analysis of 20 clones was carried out and the Results were submitted to the IgBLAST-imgt website to determine accuracy and diversity of the antibody library.
Results:
The pPICZαA-CH;-CL; vector was sequenced and the expressions of CH; and CL; were determined by immunoblotting. The full-length human antibody library pPICZαA-VH;-CH;-VL;-CL; was constructed with a library capability of 10(5);. The IgBLAST-imgt analysis showed that the antibodies were functional as predicted.
Conclusion:
We successfully constructed a full-length human secretory antibody library in N-glycoengineered Pichia pastoris, which allows the screening of functional antibodies.
