A monoclonal antibody with broad anti-HLA-DR activity fails to bind to DRw11/Dw5: possible effect of a unique polymorphism on the beta 1 domain alpha-helix.
We describe the generation and characterization of a murine monoclonal antibody with broad anti-HLA-DR beta activity, but which does not recognize DRw11/Dw5. A BALB/c mouse was immunized with a human alloreactive T-cell clone, with the intention of generating monoclonal anti-T-cell-receptor antibodies. In the course of screening the resulting hybridomas, a clone was detected which secreted an antibody (MP2) with anti-HLA-DR activity. This was shown by flow cytometry as well as immunoprecipitation followed by gel electrophoresis. Flow cytometry experiments using transfectants bearing hybrid human/murine class II molecules demonstrated that MP2 binds to the DR beta chain. MP2 bound to a wide range of Epstein-Barr-Virus-transformed cell lines and transfectants expressing different DR beta 1 and DR beta 3 subtypes: the only exceptions were three transfectants expressing DRw11/Dw5. One of these (RGT1) was shown to be functionally normal in DRw11-restricted alloreactive and antigen-specific systems. The specificity of MP2 was confirmed in functional assays: it was able to inhibit the recognition of DR1 and DRw15 but not DRw11 by alloreactive T-cell clones. Previously reported sequence data show that the beta chain of DRw11 differs from all other DR and DQ beta chains at position 58 by a glutamic acid for alanine substitution. This may account for the inability of DRw11/Dw5 to be recognized by MP2. The data emphasize the value of transfectants in defining precisely the allelic and chain specificity of anti-major-histocompatibility-complex (MHC) antibodies, and illustrate the influence that inaccessible residues can have on the conformation of MHC molecules.
Mackworth-Young
C G
CG
Department of Immunology, Royal Postgraduate Medical School, London, England.
Kevany
M
M
Lombardi
G
G
Sidhu
S
S
Sharrock
C
C
Batchelor
J R
JR
Lechler
R I
RI
D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D006684:HLA-DR Antigens / Q000737:chemistry / Q000235:genetics; D059810:HLA-DR Serological Subtypes; D006801:Humans; D008968:Molecular Conformation; D011110:Polymorphism, Genetic
D000911:Antibodies, Monoclonal; D006684:HLA-DR Antigens; D059810:HLA-DR Serological Subtypes; C080766:HLA-DR11 antigen