Chemical properties of leucocytic pyrogen: I. Partial purification of rabbit leucocytic pyrogen

Abstract

Rabbit leucocytes, if properly stimulated, will yield an endogenous or leucocytic pyrogen that is chemically and biologically distinct from exogenous pyrogens such as bacterial endotoxins. Just how the leucocytic pyrogen is produced is not yet known but the present study was undertaken in an effort to learn more of its chemical properties. Crude leucocytic pyrogen, obtained from sterile peritoneal exudates, was dialyzed against distilled water, freeze-dried, and applied first to a DEAE column to remove much of the protein, then to a phosphorylated cellulose column which adsorbed leucocytic pyrogen while allowing virtually all of the remaining protein to be eluted. The leucocytic pyrogen in turn was eluted by a stepwise increase in pH with a resulting thirty-fold increase in pyrogenic activity per gram of protein as compared with the crude pyrogen. This pyrogenic protein material is inactivated by 60°C for 5 minutes and by trypsin digestion. It is electrophoretically homogenous at pH 5.0 and 8.0 and yields at least nine amino acids on hydrolysis. The methods outlined above enable rabbit leucocytic pyrogen to be obtained in a relatively pure form so that further chemical characterization is now possible.