Preliminary characterization of a polymorphonuclear leukocyte stimulant isolated from alkali-treated collagen

This study reports the preliminary characterization of a stimulant released from alkali-treated collagen which activates the respiratory burst of PMNs. The supernatant fraction from alkali-treated collagen (SATC) was precipitated with ammonium sulfate, resuspended, and centrifuged through a sucrose gradient (10-30%, W/V). Proteins were detected throughout the gradient but PMN stimulatory activity was found mainly in fractions 1 and 2 (bottom of gradient), indicating a very high molecular weight. When SATC was layered on a denser sucrose gradient (20-85%, W/V), protein was spread over the upper half of the gradient. The peak of PMN stimulatory activity was found in the middle of the gradient (fractions 8 and 9), indicating a molecular weight similar to the 200,000 dalton standard. High pressure liquid chromatography (HPLC) of fractions from this heavier sucrose gradient showed a small peak, followed by a much higher main peak. The small peak, with the highest molecular weight peptide, is responsible for stimulation of the respiratory burst of PMNs (fraction 8 and 9). The source of this stimulant is yet to be determined.