MICROSCOPICAL AND PHYSIOLOGICAL EXAMINATION. 29 



one bacterium in each drop. With five of these drops he 

 inoculated in one case five glasses containing boiled milk. 

 The result was that the milk in one of these coagulated, 

 showing that it contained Bacterium lactis, whilst the four 

 other glasses remained unaltered and did not show the 

 presence of bacteria. The same method was subsequently 

 employed by NAEGELI and Friz. 



Air has also been made use of for such a dilution 

 (PASTEUR). A small portion of yeast is dried and ground 

 with powdered gypsum. The resulting fine powder is thrown 

 into the air from a height, a series of vacuum flasks being 

 opened while the particles are falling. Isolated yeast-cells 

 which are distributed in the resulting dust-cloud may possibly 

 enter some of the flasks. 



In comparison with the physiological methods the dilution 

 method now described is a distinct advance; indeed we have 

 thus approached much nearer to the goal. On the other 

 hand, it is clear that, even if the dilution is carried as 

 far as in the case mentioned, in which only one of several 

 flasks shows development, it is not yet proved that this one 

 flask has received only one germ. Thus, there is still great 

 uncertainty, even in cases where the individuals with which 

 we are working can be counted. Moreover, it is difficult 

 to count individual bacteria, and often, indeed, quite im- 

 possible. In all cases the accuracy of such calculations is 

 very questionable. Thus, the problem remains to be solved : 

 How are we to distinguish the flasks which have only received 

 one cell from those which, in spite of the counting, have been 

 infected with several cells ? For the bacteria, no means have 

 as yet been found of solving this difficulty. 



In the case of yeast, this problem was solved by HANSEN, 

 who developed the method to such a degree of perfection as to 

 create, in fact, an exact method (1881). He employed dilution 

 with water, in the following manner : The yeast developed in 

 the flask is diluted with an arbitrary amount of sterilised water, 

 and after vigorous shaking, the number of cells in a small drop 

 of the liquid is determined. The counting, in this case, is 

 effected in a very simple manner by transferring a drop to a 

 cover-glsss, in the centre of which some small squares are en- 



