THE TECHNIQUE OF SERUM REACTIONS 309 



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 thrusting motion. A column of peri- 

 toneal fluid will run into the glass tubing by capillary attraction; 

 this can then be blown out upon a cover-slip for hanging-drop 

 examination or may be blown upon a slide, smeared, and examined 

 after staining. The reaction is regarded as positive if within thirty 

 minutes to an hour the peritoneal exudates of the animals receiving 

 immune sera contain only swollen or disintegrated microorganisms, 

 while in that of the control animals only well-preserved and unde- 

 generated bacteria are found. In dealing with typhoid bacilli and 

 cholera spirilla, in connection with which the test is most often 

 used, active motility in the controls is of much help. Should there 

 be extensive degeneration of the bacteria in the exudate of the con- 

 trol animals the test is of no value. 



2. Identification of a microorganism by observing its susceptibility 

 to lysis in a known immune serum in vivo: 



The technique for this test is practically the same as that of 

 the preceding except that in this case we require a potent known 

 immune serum and normal serum for control. It is necessary, 

 furthermore, that by previous tests we should know the degree of 

 dilution in which the immune serum will cause complete bacteriolysis 

 of the microorganism used in its production. Thus, if we are em- 

 ploying a typhoid immune serum and are about to test by this 

 method an unknown Gram-negative bacillus, we must know the 

 titer of the serum for the typhoid bacillus itself. 



Mixtures are then made of dilutions of this serum and definite 

 quantities of the microorganism to be tested. It is best, always, 

 to employ from ten to one hundred times the amount of immune 

 serum which suffices to produce lysis with its homologous micro- 

 organism. Thus, if the serum has been found to be active in dilu- 

 tions of 1:1,000, it is employed in the test in dilutions at 1:1,000, 

 1 :100, and 1 :10. These dilutions are then injected into guinea-pigs 

 in quantities of 1 e.c. together with the bacteria to be tested, and 

 control guinea-pigs are injected with undiluted normal serum mixed 

 with the bacteria and with salt solution and the bacteria. The 

 exudates are then observed in the same way as in the preceding 

 experiment. 



