242 



ANTITOXINS 



of the fourth day Nos. 1, 2, 3, and 4 were dead and Nos. 5 and 6 were 

 alive, the serum would contain 200 units of antitoxin in a cubic centi- 

 meter. These injections are best given with precision syringes, the one 

 devised by Hitchens being particularly serviceable (Fig. 74). The syr- 

 inges are sterilized, and the needles are dipped in sterile vaselin to 

 plug them. The mixtures are made in the barrel of the syringe, and 

 sufficient sterile salt solution is placed in the side-arm to bring the total 

 volume of the injection up to 4 c.c., and to wash in all traces of toxin and 

 antitoxin. The mixtures are allowed to stand for at least fifteen min- 

 utes (Park) before being injected (Fig. 75).. The pigs must be of proper 

 weight i. e., about 250 to 300 grams; the abdominal wall is shaved, 



FIG. 75. A BATTERY OF KITCHENS SYRINGES. 



and the injection given directly in the median abdominal line. The 

 animals are placed two in a cage, and carefully observed for four or 

 five days for symptoms of toxemia and edema about the site of injection. 



Romer's Method of Determining Small Amounts of Diphtheria Antitoxin. The 



principle of this method is based upon the observation that, when very small amounts 

 of diphtheria toxin are injected intracutaneously into the abdominal skin of guinea- 

 pigs, small areas of edema and necrosis result in about forty-eight hours. When 

 such injections are made with mixtures of toxin and antitoxin, the presence of free 

 toxin is indicated by such tissue changes. It is chiefly used in determining the anti- 

 toxin content of human serums after active immunization with the toxin-antitoxin 

 mixtures of von Behring. (See p. 770.) 



Technic. I conduct this test in the following manner: The "limes-necrosis" 

 (L n ) dose of a toxin is first determined, which is the amount of toxin which, together 

 with jinny f a unit of standard antitoxin, will still produce a minimal amount of ne- 

 crosis in forty-eight hours after intracutaneous injection into .guinea-pigs. A series of 

 dilutions of the L+ dose of a toxin is made, ranging from 1 : 5 to 1 : 100, and 0.2 

 c.c. of each mixed with 0.2 c.c. of antitoxin so diluted that each 0.1 c.c. contains 



