744 THE ANIMAL PARASITES OF MAN 



which sterile autolysed tissue had been added. The sterile tissue, 

 such as brain or liver, was kept in a sterile thermostat at a temperature 

 of 40 C. for ten to twenty days. The surface of the agar should 

 be broken up before use, and the medium must not be too acid (not 

 over i '5 per cent.). They consider that autolysed tissue is necessary 

 for the growth of Entamcebae, and that naturally associated bacteria 

 aid growth by autolysing the tissues. 



(2) Culture Media for the growth of Protozoa parasitic in the 

 Blood. MacNeal and Novy/ in 1903, used a mixture of blood and 

 agar for the cultivation of trypanosomes such as T. lewisi and T. brucei. 

 They employed varying proportions of the blood and agar, a medium 

 consisting of two parts of defibrinated rabbit's blood mixed with one 

 part of agar being useful. The trypanosomes grew in the water of 

 condensation. Some of the authors' earlier formulae contained 

 different proportions of blood and agar with a little peptone, while 

 one of these media contained meat extract, agar, peptone, salt and 

 sodium carbonate. The temperature, like the proportion of blood 

 and agar, varied with the trypanosome investigated, but the optimum 

 was 25 C. 



Mathis 2 (1906) somewhat simplified the technique of Novy and 

 MacNeal. He collected the blood of a suitable animal, such as rabbit, 

 cow or dog, strict asepsis not being essential. The blood was 

 defibrinated in the ordinary way. One part of blood was added 

 to two parts of agar at 50 C. The mixture was sterilized several 

 times by heating to 75 C. or 100 C. Slopes were made and the 

 water of condensation was inoculated with a little blood containing 

 the trypanosomes. Blood may be obtained from a superficial vein 

 or from the heart. 



Novy-MacNeal-NicoUe or N. N-. X. Medium. In 1908 C. Nicolle 3 

 brought forward a modification of the Novy-MacNeal (N.N.) 

 medium. The formula is as follows : 



... ... ... ... 14 grm. 



Sea salt ... ... ... ... ... 6 ,, 



Water ... ... ... ... ... 900 ,, 



Apparently pure sodium chloride can be substituted equally 

 well for sea salt. The mixture is placed in tubes and sterilized in 

 an autoclave. To each tube one-third of its volume of rabbit 

 blood, taken by aseptic puncture of the heart, is added. The salt 

 agar is kept liquid at 45 C. to 50 C. and the blood is added to 

 the mixture. The culture medium so prepared is maintained for 

 five days at 37 C., and then for a few days at room temperature. 



1 See Sleeping Sickness Bulletin (1909), i, No. 8, p. 287. 



- C. R. Soc. BioL, Ixi, p. 550. 3 c% R Acadf Sci ? cxlyi> p _ 42 



