Construction and screening of natural immune Fab antibody phage library from patients with colorectal cancer
10.3321/j.issn:1673-4254.2001.04.010
- VernacularTitle:人源性大肠癌自然致敏噬菌体抗体Fab呈现库的构建与筛选
- Author:
Bao-Ping WU
1
;
Ya-Li ZHANG
;
Bing XIAO
;
Zhen-Shu HANG
;
Zhuo-Sheng LAI
;
Ya-Dong WANG
;
Chun-Fang GAO
;
Dian-Yuan ZHOU
Author Information
1. 第一军医大学附属第一医院(南方医院)
- From:
Journal of Southern Medical University
2001;21(4):270-274
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct natural immune Fab fragment phage display library and to screen the antibodies against human colorectal cancer. Methods Total RNA in the metastatic lymph nodes from patients with colorectal cancer was extracted routinely, and reverse transcriptase PCR employed to amplify the heavy chain Fd and light chain κ cDNA. The amplification products were inserted in succession into the vector pComb3 to construct human Fab fragment library, which was screened and identified using phage display technique. Results The heavy chain Fd fragment and light chain κ fragment (650 bp in length) were amplified, and the products were inserted into pComb3 vector with a recombination rate of 40%. The constructed phage display Fab library had a capacity of up to 1.48×106, and was enriched approximately 120 times after 3 cycles of panning. Dot immunoblotting found the expression of Fab fragment which could bind to the antigens of human colorectal cancer. Conclusion The phage display library of the antibody Fab fragment from naturally immunized lymph nodes is successfully established, rendering it possible to screen the antibodies against human colorectal cancer.
