MONOCLONAL-ANTIBODY AGAINST PIII OF FILAMENTOUS PHAGE - AN IMMUNOLOGICAL TOOL TO STUDY PIII FUSION PROTEIN EXPRESSION IN PHAGE DISPLAY SYSTEMS

被引:35
作者
TESAR, M
BECKMANN, C
ROTTGEN, P
HAASE, B
FAUDE, U
TIMMIS, KN
机构
[1] GBF NAT RES CTR BIOTECHNOL, DEPT MICROBIOL, D-38124 BRAUNSCHWEIG, GERMANY
[2] GBF NAT RES CTR BIOTECHNOL, DEPT APPL GENET, D-38124 BRAUNSCHWEIG, GERMANY
来源
IMMUNOTECHNOLOGY | 1995年 / 1卷 / 01期
关键词
MONOCLONAL ANTIBODY; FILAMENTOUS PHAGE M13; PHAGE DISPLAY; PIII-GENE; FUSION PROTEIN EXPRESSION;
D O I
10.1016/1380-2933(95)00005-4
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
A monoclonal antibody directed against the gene 3 product (pIII) of filamentous phage M13 was produced to study pIII-fusion protein expression in E. coli and its incorporation in the phage capsid. The protein was gel-purified from E. coli expression cultures harboring the genetic information of pIII under the control of an inducible lac promoter. To study pIII-fusion protein expression, phage display systems were applied in which either the whole pIII or the C-terminal half was used (McCafferty et al. (1990) Nature (London) 348, 552-554; Szardenings and Collins (1990) Gene 94, 1-7; Barbas and Lerner (1991) In: Methods: Companion to Methods in Enzymology, Combinatorial Immunoglobulin Libraries on the Surface of Phage (Phabs): Rapid Selection of Antigen-Specific Fabs, Vol.2, Academic Press, Orlando, pp. 119-124). In all cases, the monoclonal antibody was able to detect the native and the recombinant protein in E. coli and on the phage tip using non-denaturing (ELISA) and denaturing (SDS-PAGE, immunoblot analysis) conditions. All selected pIII-specific monoclonal antibodies were found to be directed against epitopes within amino acids 198 to 406 of pIII, which is necessary for capsid incorporation and therefore included in all pIII-mediated phage display designs.
引用
收藏
页码:53 / 64
页数:12
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