COMPLEMENT FIXATION' IN SYPHIIJS 427 



antigen suggested by Noguchi and Bronfenbrenner (1911). Many 

 modifications such as those of Tschernogubow (1908), Noguchi 

 (1909), Hecht (1909), Stem (1909) and Bauer (1909) were 

 introduced. At the present time these are for the most part of 

 historic interest only and the student is referred to the original 

 articles for further information. 



Third Stage Introduced by Cholesterinized Antigen. — The 

 third stage, according to Kolmer (1922), began with the discovery 

 by Browning, Cruikshank and McKenzie (1910) that cholesterol 

 added to lecithin antigen improved it and also the discovery of 

 H. Sachs that a cholesterinized alcoholic extract of guinea pig 

 heart is a better antigen than a plain alcoholic extract. These 

 observations were soon confirmed by Mcintosh and Fildes (1912), 

 AValkcr and Swift (1913), and Kolmer, Laubaugh and Williams 

 (1914). In all Wassermann tests for many years it has been cus- 

 tomary to use three antigens, a cholesterinized, a plain alcoholic 

 extract of normal heart and an acetone-insoluble lipoid antigen. 

 False positives have been encountered occasionally when only 

 cholesterinized antigens were employed, but fairly accurate results 

 are obtained Avhen all three antigens are used and the results 

 carefully interpreted. 



Fourth Stage: Introduction of Improved Antigen. — While 

 Kolmer speaks of only three stages in the development of the 

 AVassermann reaction, it seems desirable to mention two or three 

 additional stages of development. 



A fourth stage representing definite improvement in the antigen 

 occurred when Neymann and Gager (1917) made their recom- 

 mendations as to antigen preparation. These recommendations in- 

 cluded a change from guinea pig heart, which contains Forss- 

 mann's antigen, to beef heart, which does not contain the hetero- 

 ]>hile antigen. Since occasionally one encounters a normal serum 

 with a higli heterophile antibody titer, it is conceivable that false 

 poxitives may, on rare occasions, be obtained. They further recom- 

 mended the preparation of an alcoholic extract of beef heart using 

 methods of grinding and drying, preliminary and final extrac- 

 tions of the tissue and later cholesterinizing part of the extract, 

 that are quite similar to the more recent nu'thods recommended 

 by Kolmer. They further suggested that a cholesterinized and 

 an acetone-insoluble lipoid antigen be prepared from portions 



