Solubilisation effect of Nonidet P-40, triton X-100 and CHAPS in the detection of MHC-like glycoproteins

J Immunol Methods. 1988 Aug 9;112(1):133-8. doi: 10.1016/0022-1759(88)90043-9.

Abstract

We have analysed the differential solubilisation effect of three detergents on cell-membrane histocompatibility glycoproteins. Two nonionic detergents (Nonidet P-40 and Triton X-100) which are extensively used in the extraction of MHC proteins and a zwitterionic detergent (CHAPS) which is sulphobetaine derivative of cholic acid were used. An AKR (H-2k) derived spontaneous leukaemic cell line--424--was used as the experimental model. In this tumour cell line a class I-like antigen is expressed but not directly detected by cell-binding radioimmunoassay or immunoprecipitation from NP-40 or Triton X-100 solubilised glycoproteins. However, 46 kDa and 12 kDa bands consistent with the classical H-2 class I pattern were seen by SDS-PAGE after immunoprecipitation with the 34.5.8 anti-H-2Dd MoAb using CHAPS solubilised 424 glycoproteins. The H-2Dd-reactive molecule appears to be associated with at least one of the syngeneic class I specificities (H-2Kk, H-2Dk) and not accessible to react with the specific anti H-2Dd MoAb. The detergents NP-40 and Triton X-100 appear to be less efficient than CHAPS in breaking protein-protein interactions. This property of CHAPS permitted the adequate solubilisation of the novel antigen and its direct detection. The results of this study suggest that the alternative use of a non-denaturing zwitterionic detergent may contribute to the detection and characterisation of MHC-related, membrane-bound proteins of tumours and normal cells.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal
  • Cell Line
  • Cholic Acids
  • Detergents*
  • Electrophoresis, Polyacrylamide Gel
  • H-2 Antigens / isolation & purification*
  • Membrane Glycoproteins / isolation & purification*
  • Mice
  • Mice, Inbred AKR
  • Octoxynol
  • Polyethylene Glycols
  • Precipitin Tests
  • Sodium Dodecyl Sulfate
  • Solubility
  • Surface-Active Agents*

Substances

  • Antibodies, Monoclonal
  • Cholic Acids
  • Detergents
  • H-2 Antigens
  • Membrane Glycoproteins
  • Surface-Active Agents
  • Sodium Dodecyl Sulfate
  • Polyethylene Glycols
  • Octoxynol
  • Nonidet P-40
  • 3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate