Discrete monoclonal antibodies define functionally important epitopes in the CD200 molecule responsible for immunosuppression function.

BACKGROUND: Both murine and human CD200 fusion proteins (CD200Fc) act as immunosuppressants after engagement of cell-bound receptors (CD200R). Anti-CD200 monoclonal antibodies (mAbs) augment activity in mixed leukocyte cultures (MLCs) (increased cytotoxic T lymphocyte/cytokine production) after neutralization of endogenous CD200 activity. Previous studies documented critical regions in the N-terminal domains of both CD200 and CD200R1 for ligand:receptor binding and defined a number of synthetic CD200 and CD200R peptides that antagonize that interaction.
METHODS: We used a panel of mAbs to mouse and human CD200Fc to compare the rank activities of antibodies for binding (flow cytometric analysis [FACS] or enzyme-linked immunoadsorbent assay [ELISA]) to CD200 with their abilities to augment immune reactivity in MLCs.
RESULTS: Only mAbs defining epitopes in the N-terminal domain could augment MLC reactivity (or block immunosuppression by soluble CD200Fc), whereas mAbs targeting C-domain epitopes, although reactive in ELISA or FACS (targeting cell surface CD200), were inactive in MLCs.
CONCLUSION: In addition to defining the importance of N-terminal epitopes for CD200 function, rank comparison of mAbs for FACS staining of CD200 expressed on various cell types indicates heterogeneity in expressed CD200.