THE AGGLUTINATION REACTION 191 



to one hour before use. Some strains of an organism are better 

 than others, and old laboratory strains are generally much more 

 sensitive to agglutination than recently isolated ones. 



Dilution of the serum. This may be carried out in various ways, 

 with the haemocytometer pipette, with a pipette with rubber teat 

 as used for opsonin work (Fig. 35, a, p. 215), or with a platinum 

 loop. With the pipette a little serum is aspirated up so as to 

 occupy 1^-2 cm. of the stem, and the upper limit is marked with a 

 grease pencil or ink. A bubble of air is then admitted so that an 

 air-space is left between the end of the pipette and the lower end of 

 the column of serum. The end of the pipette is then immersed in 

 a watch-glass of salt solution, and the salt solution is aspirated up 

 to the mark, another bubble of air is admitted, and the process is 

 repeated again and again ; so that, finally, the pipette contains 

 1 volume of serum and 4-14 volumes of salt solution, each volume 

 being separated from the next one by an air-bubble. The contents 

 of the pipette are then expelled into a watch-glass and thoroughly 

 mixed, and further dilution of this dilution is performed in the same 

 manner. Two or three dilutions are usually made e.g. 1 in 15, 

 1 in 25, and 1 in 50. A platinum loop may also be employed as a 

 measure ; a loopful of the serum is deposited in a watch-glass, and 

 by spotting round it nine or fourteen loops of salt solution a dilution 

 of 1 in 10 or 1 in 15 is prepared, or any other dilution in a similar 

 manner. 



The microscopic test. Two or three hanging-drop slides are 

 vaselined, and two or three cover-glasses cleaned. One loopful of 

 a dilution of serum is placed on each cover-glass, and to each is 

 added a loopful of the broth culture of the organism e.g. typhoid 

 and well mixed up, and the specimens are mounted as hanging 

 drops. Starting with three dilutions of serum e.g. 1 in 15, 1 in 25, 

 and 1 in 50 the dilutions in the specimens will be 1 in 30, 1 in 50, 

 and 1 in 100 respectively. Should only one dilution of serum have 

 been made e.g. 1 in 15 if on each cover-glass one loopful of this 

 be placed, and to the first be added one loopful, to the second two 

 loopfuls, and to the third three loopfuls of typhoid culture, then the 

 final dilutions in the three specimens will be 1 in 30, 1 in 45, and 

 1 in 60 respectively. 



Care should be taken that the hanging-drop cultures are quite 

 sealed with the vaseline, so that evaporation is prevented. The 

 hanging drops are then examined microscopically, a -in. objective 

 sufficing for typhoid. In the case of typhoid the following phenomena 

 will be observed : The motility of the majority of the bacilli is 

 instantaneously or very quickly arrested, and in a few minutes they 



