256 INFECTION AND IMMUNITY 



A number of dilutions of the serum are made with sterile neutral 

 bouillon or salt solution, ranging from 1 in 20 to 1 in 500, or higher. It 

 is convenient to make a first solution of I in 20. One c.c. of this mixed 

 with 4 c.c. of broth will give I in 100. One c.c. of the I in 100 dilution 

 with 1 c.c. of broth, 2 c.c. of broth and 4 c.c. of broth will give 1 in 200, 

 1 in 300, and 1 in 500, respectively. Into one cubic centimeter of each 

 of these dilutions there is placed one platinum loopful of a twenty-four- 

 hour agar culture of the microorganism against which the serum is to 

 be tested. Into another test tube is placed 4 c.c. of broth, without 

 serum, and with one loopful of the microorganisms. The mixtures 

 are thoroughly emulsified in each case by rubbing the bacteria against 

 the sides of the tube with the platinum loop. 



Intraperitoneal injections into guinea-pigs are then made of I c.c. 

 of each of the serum-dilution-bacterial-emulsions. A control guinea-pig 

 (better two or three) receives Ic. c. of the broth emulsion — one-fourth 

 as many bacteria, therefore, as the animals receiving the serum 

 dilutions. 



Before making the injections, areas on the lateral abdominal walls 

 of the guinea-pigs are shaved, and small incisions made through the 



Fig. 65. — Capillary Pipette for Removal op Exudate in doing the 



Pfeipper Test. 



skin, down to the muscular layers. The needle of the syringe is then 

 introduced perpendicular to the skin until it has penetrated the peri- 

 toneum, and then carefully slanted to avoid puncturing the gut. The 

 animals need not be strapped down during this procedure and after- 

 ward may be allowed to run about. 



After one-half hour, and again after one hour has elapsed, a drop 

 of peritoneal exudate is removed from each guinea-pig and examined 

 in the hanging drop for granulation and swelling of the bacteria. The 

 method of obtaining the peritoneal exudate is as follows: Small glass 

 tubing is drawn out into capillary pipettes, the ends of the capillaries 

 being again drawn to fine points in a small yellow flame. A number of 

 such pipettes should be prepared before the test is begun. The guinea- 

 pig is then held down upon a table, either by an assistant or by the left 

 hand of the operator, and the point of the pipette pushed through the 

 cut in the abdominal wall into the peritoneum by a sharp, quick thrust- 



