Pure Cultures of Parasitic Amebas. 
57 
human amebic dysentery, grew with comparative ease and some 
abundance on agar plates streaked with rabies brains. 
I then tried normal brains as controls and found that this strain 
of ameba grew with almost equal ease on them ; so that the problem 
was changed for the time from that of a possible aid in determining 
the nature of the Negri bodies to that of the growth of pure cul- 
tures of amebas. 
Only four investigators (Kartulis 1893, Casagrandi & Barba- 
gallo 1897, Tsugitani 1898) have claimed to obtain amebas 
growing free from other living organisms and none of these have 
been quite clear in regard to their technic nor have they apparently 
grown their organisms for more than two or three culture genera- 
tions, which, as controls show, does not rule out the factor of sub- 
sistence upon food stored up in the amebas themselves. 
The majority of investigators who have tried to obtain pure 
cultures of amebas all make emphatic statements as to the im- 
possibility of obtaining such cultures by any methods tried. 
The present work, the details of which will be given in the full 
report, may be divided into 3 parts: (1) Obtaining living amebas 
free from other living organisms, (2) Obtaining sterile brain tissue, 
(3) Making successive transplants of amebas and brain tissue and 
proving that every transplant is free from other living organisms. 
Each step requires many controls. 
In obtaining living amebas free from other organisms a method 
has been used which has been employed successfully by other 
investigators, differing only in detail. The stock culture of ameba 
and bacterium is planted in small amount in the center of a nutrient 
agar plate from which the water of condensation has been dried. 
On such a plate at 25 0 to 30 0 C, in 48 hours, the amebas usually 
spread beyond the bacteria. If they do not, a drop of normal salt 
solution will help them do it in another 24 hours, and there is then 
a wide-spreading zone of motile amebas minus bacteria. Platinum 
loopfuls of these edge amebas are transplanted to a series of agar 
plates, and thoroughly controlled for presence of bacteria. 
The brain tissue is obtained by removing under strictly aseptic 
precaution the cerebrums of normal rabbits or guinea pigs, drop- 
ping them into 5 per cent, carbolic acid for one to two minutes, 
then placing them on agar plates and removing Amnion's horns 
