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Updated: Aug 8, 2026
Scalable High Throughput Selection From Phage-displayed Synthetic Antibody Libraries
Published on: January 17, 2015
[Construction and characterization of anti-DOTA-Y phage antibody library]
Fei Song1, Jin-liang Xing, Si-he Zhang
1Cell Engineeing Research Center, Fourth Military Medical University, Xi'an 710032, China. chcerc1@fmmu.edu.cn
Aim:
To construct a anti-dodecane-tertraacetic acid-yttrium(DOTA-Y) immune Fab phage antibody library.
Methods:
BALB/c were immunized with BSA-Y-DOTA which was prepared by DOTA-conjugated BSA and chelated with Y. After determination of anti-serum, total RNA was extracted from splenic lymphocytes of immuned mice. The heavy chain Fd and light chain Kappa genes repertoires of immunoglobulin were amplified respectively by RT-PCR, and then the amplified products were cloned into the reconstructive phage vector pComb3M to construct anti-DOTA-Y Fab antibody. And then, the recombination rate, diversity and display of Fab antibody library were identified by restriction endonuclease digestion, DNA sequencing and ELISA.
Results:
BSA-Y-DOTA was prepared successfully, and a higher titer of immune sera was achieved. The amplified gene fragments of Fd and Kappa chain by RT-PCR were correct and the length was with about 650 bp, and were inserted exactly. The sink size of Fab phage display library reached 8 x 10(7), the re-combination rate was about 90%, and it possesed great diversity. In addition, ELISA detection showed that there was Fab expression on the phage library.
Conclusion:
An immune Fab phage antibody library of DOTA-Y has been constructed successfully, which lays a solid foundation for screening specific anti-DOTA-Y antibody.
