DETERMINATION OF THERMAL DEATH-POINT. 71 



then having taken a bulb and sterilised the point and broken it 

 off with sterile forceps, the bulbous end is to be rapidly passed 

 through the flame of a Bunsen burner four or five times to 

 expel some of the air, and the sterile point of the shank is 

 to be dipped into the fluid in 

 the dish, and as the bulb cools 

 the fluid runs slowly up the 

 shank and falls into the bulb 



below. It is well not tO fill FIG. 36. Sternberg's bulb adapted for 



the bulb more than one-quarter, thermal death-point test. 



as a great bulk of fluid is to be avoided, interfering as it does with 

 the delicacy of the test. Removing the bulb from the fluid, its 

 point is carefully sealed in the flame and it is then deposited in 

 a small galvanised sheet-iron box perforated with many small 

 holes, or into a stout, finely meshed wire box; both bulb and 

 box are then to be placed in a water bath with enough water 

 in it to submerge the box to the depth of at least one inch, and 

 kept for the required time at a constant temperature. [In 

 testing vegetative forms of bacteria, it is recommended to begin 

 with an exposure of five minutes at 50 C., then ten minutes at 

 50 C., and so on, for every five succeeding degrees up to 65. 

 Spores are tested in boiling water with exposures varying from 

 one minute up to twenty, or more.] After conditions of time 

 and temperature have been fulfilled, the bulb is removed, the 

 shank wiped dry, the point broken off by forceps under sterile 

 precautions, and the shank grasped by the forceps near the bulb, 

 which is now held uppermost so as to permit of the ready dis- 

 charge of the contents. This step is accomplished by introduc- 

 ing the shank of the bulb into a tube of previously melted agar, 

 whose temperature is 42 C., and, bringing the upper empty end 

 of the bulb near to the lowermost portion of Bunsen flame, 

 expansion of the air at once drives the contents into the agar, 

 when they are to be well mixed and poured into a sterile Petri's 

 dish, and incubated for 72 hours, and examined for evidences 

 of growth. Caution must be observed in expelling the contents 

 of the bulb, lest the flame come into direct contact and vitiate 

 the experiment. 



The Counting of Colonies. An approximate estimate of the 

 number of bacteria present in a given amount of a fluid (say, 

 water) can be arrived at by counting the number of colonies 



