1 88 PHAGOCYTOSIS OPSONINS AND BACTERIOTROPINS. 



made by means of the ordinary edge of a slide, and stained with carbol- 

 thionin, Leishman's or Jenner's stain. 



The entire smear is divided up (with a blue grease pencil) into eight equal 

 subdivisions, by one transverse line drawn parallel to the long diameter of 

 the slide at its middle and five vertical lines, one at each edge of the smear, 

 one in the center and one equally distant between the edge and the central 

 line. It is also advantageous to employ an eye piece, the field of which has 

 been divided or made very much smaller by the insertion of a small paper 

 screen with a small central opening representing the size of the desired field. 

 Five or six fields are then counted in each of the eight subdivided areas. 

 The number of red blood cells seen in each field are enumerated in one 

 vertical column, the number of organisms in the same field in another column. 

 In this manner an average of the entire slide is obtained. 



By means of a simple proportional sum, the number of bacteria per 

 cubic centimeter of emulsion is estimated, e.g., the number of red blood cells 

 counted is 850 and the number of bacteria 1020. The red blood corpuscles 

 used in the standardization are known to number 5,000,000 to a cubic 

 millimeter or 5,000 million to a cubic centimeter; therefore the number of 

 bacteria to a cubic centimeter of the unknown emulsion is expressed as 

 follows. 



850 : 1020 : : 5,000,000,000 : No. of bacteria per c.c. of emulsion, 

 .'.6,000,000,000 = the number of bacteria per c.c. of emulsion. 



After the emulsion has been heated for one hour, the tube is unsealed and 

 a drop is expressed into an agar culture tube which is incubated for twenty- 

 four hours to demonstrate whether the emulsion is sterile or not. At the end 

 of this time, if a growth is observed, the emulsion must be heated again for 

 one hour at 60 C. and its sterility again tested for. 



Proper dilution of the emulsion is next undertaken. Small bottles 

 containing 25 c.c. of 1/2 per cent, carbolic acid in sterile saline are aseptic- 

 ally closed with rubber caps; for example, it is desirable to make up these 

 25 c.c. with staphylococcus vaccine so that each cubic centimeter contains 

 500 million bacteria, then 



(desired amt. to each c.c.) 

 500,000,000X25 (No. of c.c. desired) 



6,000,000,000 (dose of original emulsion) 



2.08 c.c. or approximately 2 c.c. of the original emulsion must be added to 

 the 25 c.c. (to be exact 23 c.c.) to make up the desired dilution. 



The rubber cap is finally coated with melted paraffin wax. 



For stock vaccines it is best to make up the different vaccines in the 

 following concentrations : 



i. Staphylococcus vaccine prepared from various strains of staphy- 



