Use of monoclonal antibodies against factor H to investigate the role of a membrane-associated protein antigenically related to H in C3b-receptor function

J Immunol. 1984 Jan;132(1):392-8.

Abstract

Three murine monoclonal IgG1 kappa-antibodies, MAH-1, MAH-2, and MAH-3, were raised against factor H purified from human plasma. In cross-inhibition studies MAH-3 did not compete with MAH-1 and MAH-2, and vice versa, for the binding to H, whereas MAH-1 and MAH-2 inhibited each other. MAH-1 and MAH-2 inhibited the binding of H to C3b attached to an ELISA plate as well as to C3b bound to sheep erythrocytes by means of the classical pathway convertase and of C3b to H attached to an ELISA plate. The determinant defined by MAH-1 and MAH-2 was no longer accessible on H bound to C3b. In contrast, MAH-3 interfere with the binding of H to C3b or vice versa only to a smaller extent but recognized a determinant still accessible on H bound to C3b and was able to agglutinate EAC14o23b-H in an indirect Coombs test. All three antibodies were shown to bind to tonsil cells and Raji cells in an indirect cell ELISA. The membrane-associated molecule detected by these antibodies had an apparent m.w. of 140,000 D in SDS-PAGE. All three antibodies partially inhibited the binding of EAC14o23b to tonsil lymphocytes and, in the presence of 0.1 mM DFP, to Raji cells; binding of EAC14o23bi and EAC14o23d to tonsil cells was not affected. We conclude that MAH-3 recognizes a determinant distinct from the ones recognized by MAH-1 and MAH-2, the latter possibly defining identical epitopes that are located close to the binding site for C3b. The fact that these two distinct epitopes could be detected on a 140,000-D membrane-associated protein from lymphoid cells strongly suggests that this molecule is at least antigenically related to serum H and shares with H a region carrying the binding site for C3b. The rosette inhibition studies imply that this structure is important for the binding of C3b-coated particles to lymphoid cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Monoclonal / immunology*
  • Antigen-Antibody Reactions
  • Antigens, Surface / immunology*
  • Binding, Competitive
  • Cell Line
  • Complement C3b Inactivator Proteins / immunology*
  • Complement C3b Inactivator Proteins / metabolism
  • Complement Factor H
  • Epitopes / immunology
  • Female
  • Humans
  • Lymphocytes / immunology
  • Membrane Proteins / immunology*
  • Mice
  • Mice, Inbred BALB C
  • Molecular Weight
  • Palatine Tonsil / cytology
  • Palatine Tonsil / immunology
  • Receptors, Complement / physiology*
  • Receptors, Complement 3b
  • Rosette Formation

Substances

  • Antibodies, Monoclonal
  • Antigens, Surface
  • CFH protein, human
  • Complement C3b Inactivator Proteins
  • Epitopes
  • Membrane Proteins
  • Receptors, Complement
  • Receptors, Complement 3b
  • Complement Factor H