Efficient construction of a large nonimmune phage antibody library: The production of high-affinity human single-chain antibodies to protein antigens

被引:341
作者
Sheets, MD
Amersdorfer, P
Finnern, R
Sargent, P
Lindqvist, E
Schier, R
Hemingsen, G
Wong, C
Gerhart, JC
Marks, JD
机构
[1] Univ Calif Berkeley, Dept Mol & Cell Biol, Berkeley, CA 94720 USA
[2] Univ Calif Berkeley, Dept Publ Hlth, Berkeley, CA 94720 USA
[3] Univ Calif San Francisco, San Francisco Gen Hosp, Dept Anesthesia & Pharmaceut Chem, San Francisco, CA 94110 USA
[4] Univ Wisconsin, Dept Biomol Chem, Madison, WI 53706 USA
[5] Univ Calif San Francisco, Dept Stomatol, San Francisco, CA 94143 USA
关键词
single-chain Fv; phage display; antibody libraries; human mAbs;
D O I
10.1073/pnas.95.11.6157
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
A large library of phage-displayed human single-chain Fv antibodies (scFv), containing 6.7 x 10(9) members, was generated by improving the steps of library construction. Fourteen different protein antigens were used to affinity select antibodies from this library. A panel of specific antibodies was isolated with each antigen, and each panel contained an average of 8.7 different scFv, Measurements of antibody-antigen interactions revealed several affinities below 1 nM, comparable to affinities observed during the secondary murine immune response, In particular, four different scFv recognizing the ErbB2 protein had affinities ranging from 220 pM to 4 nM. Antibodies derived from the library proved to be useful reagents for immunoassays, For example, antibodies generated to the Chlamydia trackomatis elementary bodies stained Chlamydia-infected cells, but not uninfected cells, These results demonstrate that phage antibody libraries are ideally suited for the rapid production of panels of high-affinity mAbs to a wide variety of protein antigens, Such libraries should prove especially useful for generating reagents to study the function of gene products identified by genome projects.
引用
收藏
页码:6157 / 6162
页数:6
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