Enhancement of Hemolytic Activity of Complement by Polyethylene Glycols. ∗

Abstract

In the course of an investigation designed to test the ability of various substances to prevent the deterioration of complement, it was found that certain of the polyethylene glycols greatly enhance the hemolytic activity of complement. The polyethylene glycols, a series of polymers with the general formula HOCH₂(CH₂OCH₂)xCH₂OH, are soluble in water and in many of the aromatic hydrocarbons. The compounds with average molecular weight up to 700 are liquids; those with average molecular weight above 1000 are wax-like solids, sold under the trade name “Carbowax”.† This paper constitutes a preliminary report on the effect of the polyethylene glycol carbowax-4000 on the hemolytic activity of complement, as determined by complement titrations in the presence and absence off this compound.

Procedure. Complement titrations were carried out according to the 50% hemolysis end-point method developed by Mayer et al., 1 and Kent et al. 2 The reaction mixtures, 20 ml in volume, had the following composition: 1) 1 part complement, the dilution so adjusted as to yield final dilution values ranging from 1:3300 to 1:950; 2) 1 part standardized suspension of sensitized erythrocytes; 3) either 2 parts of buffered saline, 3 or 2 parts of an 8% solution of carbowax-4000 in buffered saline. The sensitized erythrocytes were well mixed with either buffered saline or carbowax-4000 solution, and then the complement added dropwise with constant agitation. All components were kept in ice-water until time of incubation. The reaction mixtures thus prepared were incubated at 37°C for 60 minutes, with frequent agitation. After centrifugation of the mixtures, readings of per cent hemolysis in the supernates were made at 5500 Å in a Coleman spectrophotometer, an optical density of .500 representing complete hemolysis. Titrations were carried out in duplicate. Controls were set up of 1) sensitized erythrocytes and carbowax-4000.