134 THE PREPARATION OF VACCINES 



dead or living, present in a culture, for the dead as well as the 

 living contain the toxins which may stimulate the therapeutic 

 capacities of the body. The method consists in making a 

 mixture of blood (whose content in red blood corpuscles is 

 known) with the bacterial culture, and comparing the number of 

 bacteria with the number of corpuscles. The observer first 

 estimates the red cells in his blood ; a capillary pipette with a 

 rubber nipple and with a mark near its capillary extremity is 

 then taken, blood is sucked up to the mark, then an air-bubble, 

 and then an equal volume of the bacterial emulsion diluted 

 according to the empirical estimate the observer forms of its 

 strength. The blood and bacterial emulsion are then thoroughly 

 mixed by being drawn backwards and forwards in the wide 

 part of the pipette, a drop is blown out on to a slide, and a 

 blood film is spread which may be stained by Leishman's 

 method. The bacteria and blood corpuscles are now separately 

 enumerated in a series of fields in different parts of the 

 preparation. If a dilution has been taken in which a large 

 number of bacteria are present, an artificial field may be used, 

 made by drawing with the oil pencil a small square on a circular 

 cover-glass, and dropping the latter on to the diaphragm of the 

 microscope eye-piece. Suppose, now, that the observer's blood 

 contained 5,000,000 red cells per c.mm., that to the bacterial 

 emulsion three volumes of diluent had been added, and that in 

 the fields examined there were 500 red cells and 600 bacteria. 

 It is evident that in the undiluted culture for 500 red cells there 

 would have been 2400 bacteria. Now 500 : 2400 : ; 5,000,000 : 

 24,000,000, which last figure is the number of bacteria per 

 c.mm. of the emulsion. 



It has been found in the case of certain bacteria, e.g. the 

 members of the coli-typhoid and cholera groups, that when an 

 emulsion of these is mixed with whole blood, the serum of the 

 latter may have a bacteriolytic or an agglutinating action on the 

 organisms, which interferes with the counting. To obviate the 

 inaccuracies or difficulties thus introduced, Harrison has modified 

 Wright's method by substituting, in a given quantity of blood, 

 normal saline for the serum. The method is as follows : A 

 capillary pipette has a mark made upon it, to which blood is 

 sucked up and quickly expelled into a small tube containing a 

 little "75 per cent, sodium citrate solution ; any remaining blood is 

 washed out of the pipette with the same fluid. The tube is then 

 centrifuged to deposit the corpuscles, the supernatant fluid 

 carefully removed, and the corpuscles are washed by centrifuging 

 twice or thrice with normal saline, care being always taken not 



