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[Construction of phage display antibody library to MCF-7 cells and screening of single-chain antibodies against breast cancer cells].
Sheng Wu Gong Cheng Xue Bao. 2004 Sep; 20(5):667-72.SW

Abstract

The aim of this study is to construct a phage display single-chain variable fragment (scFv) library against breast cancer cells and screen the specific antibodies against MCF-7 cells from the library. The BALB/C mice were immunized with MCF-7 cells. Total RNA of spleens was isolated. The heavy-chain (VH) and light-chain variable region genes (VL) of the antibodies were amplified by RT-PCR and joined into a single chain by overlapping PCR with a linker DNA encoding the peptide (Gly4Ser)3. The assembled scFv fragments were cloned into the phagemids(pCANTAB5E) and the recombinant phagemids were used to transform competent E. coli TG1. The transformed TG1 cells were infected by helper phage M13KO7 and the recombinant phagemids were rescued. The scFv fusion proteins were displayed on the surfaces of the recombinant phages. A phage display antibody library of repertoire of 1.2 x 10(6) clones was constructed. The specific antibodies against MCF-7 cells were enriched by 75 times after five rounds of affinity selection. Ten recombinant phages clones that exhibited specific binding to MCF-7 cells were identified. The specificity of those phage clones was analyzed by reactivity against HepG2 cells and Hela cells by ELISA. One of the selected phage clones against MCF-7cells was used to infect E. coli TOP10 to produce the soluble scFv antibodies after induction with IPTG. The strategy of construction and screening of antibody library directed against the whole tumor cells described in this report should be generally applicable to generate tumor cell-specific antibodies.

Authors+Show Affiliations

Institute of Radiation Medicine, Academy of Military Medical Sciences, Beijing 100850, China.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

English Abstract
Journal Article

Language

chi

PubMed ID

15973987

Citation

Zhao, Yan, et al. "[Construction of Phage Display Antibody Library to MCF-7 Cells and Screening of Single-chain Antibodies Against Breast Cancer Cells]." Sheng Wu Gong Cheng Xue Bao = Chinese Journal of Biotechnology, vol. 20, no. 5, 2004, pp. 667-72.
Zhao Y, Wang QM, Fu XQ, et al. [Construction of phage display antibody library to MCF-7 cells and screening of single-chain antibodies against breast cancer cells]. Sheng Wu Gong Cheng Xue Bao. 2004;20(5):667-72.
Zhao, Y., Wang, Q. M., Fu, X. Q., Chen, J. Z., Fan, G. C., & Chen, H. P. (2004). [Construction of phage display antibody library to MCF-7 cells and screening of single-chain antibodies against breast cancer cells]. Sheng Wu Gong Cheng Xue Bao = Chinese Journal of Biotechnology, 20(5), 667-72.
Zhao Y, et al. [Construction of Phage Display Antibody Library to MCF-7 Cells and Screening of Single-chain Antibodies Against Breast Cancer Cells]. Sheng Wu Gong Cheng Xue Bao. 2004;20(5):667-72. PubMed PMID: 15973987.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - [Construction of phage display antibody library to MCF-7 cells and screening of single-chain antibodies against breast cancer cells]. AU - Zhao,Yan, AU - Wang,Qing-Ming, AU - Fu,Xue-Qi, AU - Chen,Ji-Zhong, AU - Fan,Guo-Cai, AU - Chen,Hui-Peng, PY - 2005/6/25/pubmed PY - 2010/4/30/medline PY - 2005/6/25/entrez SP - 667 EP - 72 JF - Sheng wu gong cheng xue bao = Chinese journal of biotechnology JO - Sheng Wu Gong Cheng Xue Bao VL - 20 IS - 5 N2 - The aim of this study is to construct a phage display single-chain variable fragment (scFv) library against breast cancer cells and screen the specific antibodies against MCF-7 cells from the library. The BALB/C mice were immunized with MCF-7 cells. Total RNA of spleens was isolated. The heavy-chain (VH) and light-chain variable region genes (VL) of the antibodies were amplified by RT-PCR and joined into a single chain by overlapping PCR with a linker DNA encoding the peptide (Gly4Ser)3. The assembled scFv fragments were cloned into the phagemids(pCANTAB5E) and the recombinant phagemids were used to transform competent E. coli TG1. The transformed TG1 cells were infected by helper phage M13KO7 and the recombinant phagemids were rescued. The scFv fusion proteins were displayed on the surfaces of the recombinant phages. A phage display antibody library of repertoire of 1.2 x 10(6) clones was constructed. The specific antibodies against MCF-7 cells were enriched by 75 times after five rounds of affinity selection. Ten recombinant phages clones that exhibited specific binding to MCF-7 cells were identified. The specificity of those phage clones was analyzed by reactivity against HepG2 cells and Hela cells by ELISA. One of the selected phage clones against MCF-7cells was used to infect E. coli TOP10 to produce the soluble scFv antibodies after induction with IPTG. The strategy of construction and screening of antibody library directed against the whole tumor cells described in this report should be generally applicable to generate tumor cell-specific antibodies. SN - 1000-3061 UR - https://www.unboundmedicine.com/medline/citation/15973987/[Construction_of_phage_display_antibody_library_to_MCF_7_cells_and_screening_of_single_chain_antibodies_against_breast_cancer_cells]_ L2 - http://www.diseaseinfosearch.org/result/960 DB - PRIME DP - Unbound Medicine ER -