Relation of the Phospholipins to the Reactivity of Antipneumococcus Sera

Hardy and Gardiner's method 1 for the removal of lipoids from sera is based upon the fact that extraction of protein solutions by alcohol or alcohol-ether at low temperatures does not cause denaturation. Using this method, Hartley 2 has shown that the removal of lipoids from certain antisera apparently abolishes their in vitro reactivity. Felton 3 has extracted antipneumococcus horse serum in a similar manner, and has demonstrated that the removal of lipoids does not diminish protective action.

These findings have been confirmed for Type I antipneumococcus horse serum. Extracted sera fail to agglutinate homologous type pneumococci and to give a precipitate with the specific capsular polysaccharide. In vivo, however, they show the presence of protective antibody in unaltered concentration.

Lipoid extraction was carried out in the following manner: The antiserum was introduced, with stirring, into 10 volumes of absolute alcohol at —10°C. After 6 hours′ extraction the precipitate was collected by centrifugation at a temperature below —2°C., and again extracted with an amount of chilled absolute alcohol equal to that first used. After 12 hours the precipitate was collected in the same manner, and was then extracted with anhydrous ether for 10 hours at —10°C. and for a second time with anhydrous ether for 10 hours at room temperature. The precipitate was then collected and freed of ether by vacuum distillation, and finally was dissolved in an amount of saline equal to the original serum volume. The resulting solution does not differ in appearance from untreated serum.

Although failing entirely in vitro, the extracted antipneumococcus horse serum brought about agglutination in vivo as well as unextracted serum. As a consequence of this observation extracted immune horse serum was injected intraperitoneally into mice and the peritoneal fluid was withdrawn after 30 minutes.