Purification and characterization of an antigen involved in neutrophil chemotaxis and degranulation using a monoclonal antibody.

In a previous study we described an anti-neutrophil monoclonal antibody, which inhibited human neutrophil chemotaxis and degranulation without any detectable effect on phagocytosis or oxidative metabolism (Cotter, T. G., Spears, P. and Henson, P. M., J. Immunol. 1981. 127: 1355). This antibody was termed NCD 1. In this study we determined the number of NCD 1-binding sites per neutrophil. Approximately 25 000 NCD 1 IgG-binding sites per cell were found with an equilibrium dissociation constant (Kd) of 6.5 microM for antibody binding. NCD 1 Fab bound to approximately 39 000 sites per cell with a Kd of 16.5 microM. Affinity chromatography columns prepared by coupling NCD 1 to Sepharose 4B beads were used to purify the antigen which bound this antibody. The antigen was a 110-kDa glycoprotein which was not susceptible to reduction by 2-mercaptoethanol. The antigen was not internalized following phagocytosis of opsonized sheep erythrocytes by neutrophils.