428 RESEARCH IN PROTOZOOLOGY 



Subsequently, rapid strides were made during the course of 

 some very careful work from which evolved the standardization 

 of the complement fixation test for diagnosing dourine of horses, 

 caused by T. cqidperdmn. Some of the men associated with this 

 work are Winkler and Wyschelessky (1911), Mohler, Eichhorn 

 and Buck (1913), Watson (1914, 1915, i92o),Wehrbein (1914), 

 Waldman and Knuth (1920), Dahmen (1922), Bessemans (1921, 

 1922) and Bessemans and Leynen (1922, 1923). Of especial 

 interest are the following details : As test antigen Mohler, Eich- 

 horn and Buck used freshly prepared extracts of spleens of rats 

 heavily infected with T. evansi or of trypanosomes isolated by 

 taking the blood cells with saponin. Watson (1920) decided 

 that the test was specific in diagnosing the disease and was often 

 positive before symptoms were apparent or during latent stages. 

 As test antigen, he used trypanosomes isolated by washing and 

 centrifuging first in citrated saline and later in saline, preserved 

 in twice their volume of an extracting fluid (0.85 per cent NaCl, 

 90 parts; pure neutral glycerin, 10 parts; and formol, i part) 

 and kept on ice in one cubic centimeter ampules. Reynolds and 

 Schoening (1918) modified this technique by laking the blood 

 cells in distilled water. Dahmen, in testing the suitability of 

 alcoholic and carbolized saline extracts, concluded that the active 

 components were different in each and that the alcoholic was 

 more potent. In making the latter, the concentrated trypanosomes 

 were dried at 50° C. and pulverized ; o.i gm. of powder was shaken 

 an hour in one cubic centimeter of ether ; the residue, after filtra- 

 tion, was allowed to stand in one cubic centimeter of absolute 

 alcohol two days at 37° C. ; and finally, the supernatant was 

 slowly diluted with five to ten volumes of water. Bessemans 

 (1922Z7), in a very detailed study of the anticomplementary 

 and antigenic power of organ and isolated trypanosome test anti- 

 gens extracted in alcohol and ether, found that emulsions of try- 

 panosomes isolated by centrifugation and diluted in the alcoholic 

 or aqueous extracts of Mohler or Watson were most efificient. 



Some serological method is especially needed in Chagas' disease 

 where definite diagnosis can only be made by finding trypanosomes 

 in the blood during the short acute stage of the disease or at 

 autopsy by finding the leishmanial forms in the tissues. Very 

 promising results were reported by Machado and Guerreiro 

 (1913), Villcla and Bicalho (1923) and Lacorte (1927). In all 



