Anti-CD18 Anticorpo [MEM-48] (APC) (A86011)

The cysteine-rich repeats in the stalk region of integrin beta subunits appear to convey signals impinging on the cytoplasmic domains to the ligand-binding headpiece of integrins. We have examined the functional properties of mAbs to the stalk region and mapped their epitopes, providing a structure-function map. Among a panel of 14 mAbs to the beta(2) subunit, one, KIM127, preferentially bound to alpha(L)beta(2) that was activated by mutations in the cytoplasmic domains, and by Mn(2+). KIM127 also bound preferentially to the free beta(2) subunit compared with resting alpha(L)beta(2). Activating beta(2) mutations also greatly enhanced binding of KIM127 to integrins alpha(M)beta(2) and alpha(X)beta(2). Thus, the KIM127 epitope is shielded by the alpha subunit, and becomes reexposed upon receptor activation. Three other mAbs, CBR LFA-1/2, MEM48, and KIM185, activated alpha(L)beta(2) and bound equally well to resting and activated alpha(L)beta(2), differentially recognized resting alpha(M)beta(2) and alpha(X)beta(2), and bound fully to activated alpha(M)beta(2) and alpha(X)beta(2). The KIM127 epitope localizes within cysteine-rich repeat 2, to residues 504, 506, and 508. By contrast, the two activating mAbs CBR LFA-1/2 and MEM48 bind to overlapping epitopes involving residues 534, 536, 541, 543, and 546 in cysteine-rich repeat 3, and the activating mAb KIM185 maps near the end of cysteine-rich repeat 4. The nonactivating mAbs, 6.7 and CBR LFA-1/7, map more N-terminal, to subregions 344-432 and 432-487, respectively. We thus define five different beta(2) stalk subregions, mAb binding to which correlates with effect on activation, and define regions in an interface that becomes exposed upon integrin activation.

Six monoclonal antibodies were prepared that recognize a broadly expressed antigen composed of noncovalently associated 95 kDa and 180 kDa subunits. Antibodies MEM-25, MEM-83 and MEM-95 are shown to react with a determinant(s) present on the isolated larger subunit identified as the CD11a glycoprotein, MEM-48 reacts with the isolated smaller subunit which is identical to the CD18 glycoprotein. Antibodies MEM-30 and MEM-94 recognize probably a determinant in the CD11a/CD18 complex (i.e., LFA-1 antigen) dependent on association of the constituent chains. Cytotoxic activity of NK cells in inhibited by antibodies MEM-25, MEM-95 and less strongly by MEM-30 and MEM-94.