Diverse antigen specificity of erythrocyte-reactive monoclonal autoantibodies from NZB mice.
The specificities of a panel of erythrocyte-reactive MoAbs derived from NZB mice with autoimmune haemolytic anaemia (AIHA) were determined by immunoprecipitation and immunoblotting. Of the eight antibodies, two (IgG1 MoAb 105-2H and IgG2a MoAb 34-3C) immunoprecipitated a 105-kD component identified as the erythrocyte anion channel band 3. A similar band was also immunoprecipitated by the IgG2b MoAb 34-2B when used at relatively high concentrations, but none of the remaining hybridoma antibodies precipitated any labelled erythrocyte components. In immunoblotting experiments only 34-2B reacted with band 3, indicating that the epitope recognized by this MoAb is robust and differs from the determinant(s) recognized by 105-2H and 34-3C. The remaining MoAbs to react by immunoblotting were the IgM antibodies IE10 and 4C8, both of which bound to a doublet corresponding to band 4.1 from the internal erythrocyte membrane skeleton. Of the three MoAbs which gave negative results in immunoprecipitation and immunoblotting, the IgM antibodies 103-7E and 106-10E reacted poorly with intact erythrocytes by flow cytometry, but the IgG1 antibody 31-9D bound well. ELISAs demonstrated that all four IgM MoAbs are polyreactive, since they bound to histones from a panel of nuclear antigens, and additionally 103-7E reacted with phosphatidyl choline. It is concluded that band 3 is an important autoantigen in NZB AIHA. However, since 3/5 haemolytic MoAbs failed to participate this antigen, either these antibodies represent minor components of the total autoantibody response, or responses to diverse possibly non-protein surface antigens also contribute to the pathogenesis of the disease.
de Sá Oliveira
G G
GG
Department of Immunology, University College, London Medical School, UK.
Izui
S
S
Ravirajan
C T
CT
Mageed
R A
RA
Lydyard
P M
PM
Elson
C J
CJ
Barker
R N
RN
D000744:Anemia, Hemolytic, Autoimmune / Q000276:immunology*; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000032:analysis; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001323:Autoantibodies / Q000276:immunology*; D004912:Erythrocytes / Q000276:immunology*; D005434:Flow Cytometry; D015151:Immunoblotting; D051379:Mice; D008814:Mice, Inbred NZB / Q000276:immunology*; D010743:Phospholipids / Q000276:immunology; D011233:Precipitin Tests
D001457:Anion Exchange Protein 1, Erythrocyte; D000911:Antibodies, Monoclonal; D001323:Autoantibodies; D010743:Phospholipids