The separation and identification by monoclonal antibodies of dog IgG fractions

J Immunol Methods. 1993 May 26;161(2):193-203. doi: 10.1016/0022-1759(93)90295-i.

Abstract

Four fractions of IgG from normal dog serum have been successfully isolated by gel filtration followed by protein A and protein G affinity chromatography using the fast protein liquid chromatography (FPLC) system. Protein A chromatography produced three peaks: peak 1 was fallthrough material consisting of components which did not bind to protein A, peak 2 consisted of bound material eluting at pH 6, and peak 3 contained bound material eluting at pH 3.5. The three peaks were then subjected individually to protein G affinity chromatography. Peak 1 from protein A chromatography produced a fallthrough peak followed by a weakly binding component which eluted at pH 8, and was called peak w. Peak 2 from protein A chromatography bound to protein G and eluted as a single peak at pH 3.8, and was called peak x. Peak 3 from protein A chromatography emerged as two separate peaks (y and z) off the protein G column; peak y bound and eluted at pH 4.1, and peak z bound weakly to protein G and emerged as a broad band at pH 8. Peaks w, x, y and z have been named gamma w, gamma x, gamma y and gamma z, respectively, and there purified IgG fractions were used to immunize mice for the preparation of monoclonal antibodies (McAbs). To date, two sets of McAbs have been produced: one which recognizes an epitope present in both gamma w and gamma z fractions and another set of McAbs which recognizes an epitope in the gamma x and gamma y fractions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal / immunology*
  • Antibody Specificity
  • Chromatography, Affinity
  • Chromatography, Gel
  • Dogs / immunology*
  • Hybridomas
  • Hydrogen-Ion Concentration
  • Immunoglobulin G / immunology
  • Immunoglobulin G / isolation & purification*
  • Male
  • Mice
  • Mice, Inbred BALB C

Substances

  • Antibodies, Monoclonal
  • Immunoglobulin G