570 PROTOZOA 



Malarial parasites, first cultivated in vitro by Bass and Johns 

 {J own. Exper. Med., xvi, 1912, p. 567). To 10 c.c. of malarial 

 blood from a vein add 0-1 c.c. of a 50 per cent, solution of dextrose 

 and defibrinate. Distribute the blood in test-tubes and incubate 

 at 40° C. 



Parasites will be found in some of the red cells which settle to 

 the bottom of the serum. 



Sub-culture every few days, but normal blood treated as above 

 may be used for the medium. 



frypanosomes, Leishmania and many other flagellates are 

 cultivated in blood agar, of which the N. N. N. medium devised 

 by NovY, MacNeal, Nicolle, is the most important. It is made 

 by heating together : — 



Water 900 c.c. 



Agar 14 grm. 



NaCl 6 „ 



About 5 c.c. of this mixture is put into each test-tube, which 

 is plugged and sterilised in the ordinary way. When it has 

 cooled to 50° C. and is still liquid, 2 or 3 c.c. of sterile rabbit's 

 blood are added to each tube, which is rapidly revolved in order 

 to mix the two fluids without forming bubbles. Each tube is 

 then cooled so that the blood agar forms a sloped surface, and 

 inoculation is made into the condensation fluid at the bottom. 

 The flagellates multiply in this fluid but sub-cultures must be 

 made every few days. 



III. CULTIVATION OF INTESTINAL PROTOZOA 



1117. The numerous media, which have recently been used 

 for cultivating entamoebte, ciliates and flagellates from the ali- 

 mentary canal of man and other vertebrates, all consist of a 

 saline solution containing nutrient material, such as serum, 

 ascitic fluid or egg. They may be used liquid {e.g., 1—4 below) 

 or solid (by coagulating the egg or serum or by the addition of 

 agar, etc.), and many of the media now most used (§§ 1119, 1120) 

 consist of a liquid and a solid. The following are some of the 

 most important media : — 



1118. Serum-Saline-Citrate medium consists of O'S grm. 

 Loffler's dehydrated blood serum in 100 c.c. distiUed water, 

 containing 0-7 grm. sodium chloride and 1 grm. sodium citrate 

 (Andrews, Journ. Parasit., xii. 1926, p. 148 ; Tanabe, idem, 

 p. 101). To maintain Trichomonas from man, rat and other 

 warm-blooded vertebrates in this medium sub-cultures have to 

 be made every two or three days by introducing a drop of a 

 three-day culture into a fresh tube of medium and incubating at 

 35° to 37° C. 



