Influence of extracellular Ca2+ and Mg2+ on chemotactic factor-induced neutrophil aggregation

The influences of extracellular Ca2+ and Mg2+ on chemotactic factor-induced rabbit polymorphonuclear neutrophil (PMN) aggregation was studied using a recently described Coulter counter assay technique. Compared with those of other PMN functional assays (adhesion, chemotaxis, degranulation, and phagocytosis), the cation requirements for cell aggregation appear unique. Chemotactic factor-induced aggregation did not occur in the absence of either cation. When the concentration of both cations was increased equivalently, aggregation increased. The effect plateaued at 2.8 mM of Ca2+ and Mg2+. When the concentration of one cation alone was increased, aggregation peaked. Further increases inhibited maximal aggregation. By systematic variation of the cation concentrations, optimal aggregation was found at Ca2+ and Mg2+ concentrations of 2.8 mM. At these concentrations, significant aggregation was induced with chemotactic doses of bacterial factor, the chemotactic fragment of human C5, and the synthetic chemotactic tripeptide, formyl-met-leu-phe. Thus, under these conditions, chemotactic factor-induced aggregation of neutrophils may be a useful indicator of interactions of chemotactic factors with neutrophils.