58 
Scientific Proceedings (42). 
and corpus striatums which are spead over a series of fresh agar 
plates, and put in the thermostat for 24 hours. It is peculiarly- 
important to have conditions such as to insure sterility of this 
tissue medium because of the possibility of the amebas using up 
any organism that might get into the plates before it could be 
detected. 
This crushed sterile brain is now spread in small amounts 
over the plates containing the amebas, and placed, some in thermo- 
stat, and some at room temperature. If in 24 hours the amebas 
show slight or no growth as may be the case in the first culture 
generations, a loop or two of sterile normal salt solution may be 
added, then in 24 hours at 36 0 C. or 3-5 days at 20 0 C, there is 
generally an abundant growth. 
To rule out the presence of extraneous organisms, at each 
transfer three or four plates are made. To one is added sterile 
normal salt solution, to another sterile broth, to another sterile 
blood, one is left simply with the transplanted material, and to 
the others are added the fresh sterile brain tissue for carrying on 
the culture. In the second or third culture generations, the cul- 
tures transferred on blood or broth or normal salt or plain agar 
encyst or die out. Spreads, making an added important control, 
have been made at practically every transfer and usually stained 
in three different ways. Two other strains of parasitic amebas 
are being tried and they seem to be growing, though it is too soon 
to make a positive statement. Liver tissue and spleen tissue are 
being tried, and the amebas seem to be growing on the liver tissue 
and not on the crushed spleen. 
On the brain-streaked agar plates the one strain of ameba 
is now growing abundantly in the fourteenth culture generation, 
with normal brain and in the twentieth culture generation with 
rabies brain, grown at 36 0 C. and transplanted every 2-5 days. 
