Monoclonal antibody against pIII of filamentous phage: an immunological tool to study pIII fusion protein expression in phage display systems

Immunotechnology. 1995 May;1(1):53-64. doi: 10.1016/1380-2933(95)00005-4.

Abstract

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal*
  • Capsid / immunology*
  • Capsid Proteins
  • Cells, Cultured
  • DNA-Binding Proteins / immunology*
  • Enzyme-Linked Immunosorbent Assay
  • Escherichia coli
  • Immunosorbent Techniques
  • Mice
  • Peptide Library
  • Recombinant Fusion Proteins / biosynthesis*
  • Viral Fusion Proteins / immunology*

Substances

  • Antibodies, Monoclonal
  • Capsid Proteins
  • DNA-Binding Proteins
  • Peptide Library
  • Recombinant Fusion Proteins
  • Viral Fusion Proteins