PURIFICATION AND CHARACTERIZATION OF AN ANTIGEN INVOLVED IN NEUTROPHIL CHEMOTAXIS AND DE-GRANULATION USING A MONOCLONAL-ANTIBODY

An anti-neutrophil monoclonal antibody, which inhibited human neutrophil chemotaxis and degranulation without any detectable effect on phagocytosis or oxidative metabolism was previously described. This antibody was termed NCD 1. The number of NCD 1-binding sites per neutrophil was determined. Approximately 25,000 NCD 1 IgG-binding sites per cell were found with an equilibrium Kd of 6.5 .mu.M for antibody binding. NCD 1 Fab bound to .apprx. 39,000 sites/cell with a Kd of 16.5 .mu.M. 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 [kilodalton] glycoprotein which was not susceptible to reduction by 2-mercaptoethanol. The antigen was not internalized following phagocytosis of opsonized sheep erythrocytes by neutrophils.