26 ISOLATION OF INDICATOR ORGANISMS 



or india-rubber stoppers to enable the solid and the water to- 

 be very thoroughly shaken and mixed. Definite fractions 01 

 the water are then examined in the same way as for an 

 ordinary fluid and as described below. 



The assumption has to be made that the diluting water, 

 after thorough mixture, contains all the bacteria in the solid 

 substance added. This is an inaccurate assumption, since it 

 is not to be anticipated that all the bacteria can be dislodged, 

 the extent of the error varying with the nature of the sub- 

 stance examined. If, however, it is realized that only rough 

 comparable results are in this way obtainable, this method is 

 of great value. The results are most conveniently returned 

 as per gramme of substance. 



The procedure is much simpler when liquids have to be 

 directly dealt with, but certain precautions are necessary. 

 The amounts to be examined are removed by sterile pipette. 

 For fractions of a cubic centimetre it is obvious not only 

 that the gradations must be wide apart (i.e. the bore must 

 be narrow) but also that too small fractions must not be 

 sampled. When no very special accuracy is required, it is 

 possible to measure sufficiently down to -^ c.c. for water and 

 some other liquids, if they are not heavily contaminated, but 

 with fluids such as milk it is not reliable to take so small a 

 quantity as O'l c.c. and errors as great as thirty per cent, may be 

 made in this way. In all cases where possible it is best to obtain 

 these small fractions by making up preliminary dilutions, and this 

 course is absolutely necessary for quantities less than O'l c.c. 



A whole series of such dilutions may have to be prepared. 

 They are most conveniently made by adding 10 c.c. of the 

 original fluid to 90 c.c. of sterile water in a stoppered bottle, 

 mixing very thoroughly. One c.c. of such a first dilution will 

 represent O'l c.c. of the original fluid. In the same way second 

 dilutions can be made from the first, and so on as many as 

 are required. It is preferable to dilute as above rather than 

 to add I c.c. to 99 c.c., as obviously there is far less likelihood 

 of errors from slight inaccuracies in measurement. 



One mark 10 c.c. pipettes are convenient for making the 

 dilutions in bulk. Such pipettes should be made short (9 inches 



