The Antitoxins 133 



of these tubes was opened and the contained powder dissolved in 200 cc. of a 

 mixture of 10 per cent, aqueous solution of sodium chlorid and glycerin. The 

 subsequent calculations are all based upon the strength of the antitoxin powder. 

 In Ehrlich's first test serum i gram of the dry powder represented 1700 units. 

 Of the solution mentioned, i cc. represented 17 units; Ht cc, one unit. 



Having by dilution — 1 cc. of the first dilution in 17 of water — secured the 

 standard unit of antitoxin in a convenient bulk for the subsequent manipulations, 

 it is mixed with varying quantities of the toxic bouillon to be used for testing 

 the new serums, untU the least quantity is determined that will cause the death 

 of a 250 gram guinea-pig in exactly four days, when carefully injected beneath 

 the skin of the animal's abdomen. This quantity of toxin is the test dose or L + 

 dose. If the toxic bouillon was "normal" in constitution, it should represent ioq 

 of the least certainly fatal doses that formed the basis of the old method of test- 

 ing, but as toxic bouillons contain varying quantities of toxoids it may equal 

 anywhere from fifty to one hundred and fifty times that dose. 



The test dose of toxic bouillon, having been determined, remains invariable 

 throughout the test as before, the serum to be tested for comparison with the 

 standard being modified. The calculation is, however, different because the 

 guinea-pig is receiving, not ten times, but more nearly one hundred times the 

 least fat^ dose, and the quantity of the antitoxic serum that preserves life 

 beyond the fourth day is itself the unit. 



Example: The sample of serum issued as the standard contains 17 units per 

 cubic centimeter. Serum i cc. -j- water 16 cc. = i cc. is the unit, i cc. of 

 the dilution containing one antitoxic unit is mixed with o.oi, 0.025, o°5> °-°7Si 

 0.1 cc. of the toxic bouillon. All the animals receiving less than 0.1 cc. live. 

 A new series is started, and the guinea-pigs all weighing exactly 250 grams, 

 receive i imit of the antitoxin plus toxic bouUlon 0.08, 0.09, 0.095, 0.097, o.i, 

 o.ii, 0.12, etc. It is found that all receiving more than 0.097 die in four days, 

 but that the animal receiving that dose, though very ill, lives longer. The test 

 dose may then be assumed to be o.i, or it may be calculated more closely if 

 desired. 



To test the serum itself, guinea-pigs weighing exactly 250 grams are now all 

 given toxic bouillon o.r cc. plus varying quantities of the serum — Hoo, Hoo, 

 %00i etc. All live except those receiving less than Jioo> which die about or on 

 the fourth day. The serum can then be assumed to have 400 units per cubic 

 centimeter unless it be desired to test more closely. 



Standard test serums for making tests of antitoxic serums by the 

 EhrUch method was first shipped at small expense from the Kaiser- 

 liches Institut fiir Serum-Therapie at Hochst-on-the-Main. At 

 present the Hygienic Laboratory of the United States Public Health 

 Service has legal control of the manufacture of therapeutic serums 

 and kindred products in the United States, issuing licenses to those 

 engaged in legitimate manufacture, and furnishing a standard 

 test serum, similar to that of EhrUch, to those entitled to receive it. 



A full description of "The Immunity Unit for Standardizing 

 Diphtheria Antitoxin," by M. J. Rosenau, Director of the Hygienic 

 Laboratory, can be found in Bulletin No. 21 of the U. S. Public 

 Health and Marine Hospital Service, Washington, 1905. 



As the quantity to be injected at each dose diminishes according 

 to the number of units per cubic centimeter the serum contains, it 

 is of the highest importance that therapeutic serums be as strong 

 as possible. Various methods of concentration have been sug- 

 gested. Bujwid* and H. C. Ernstf found that when an antitoxic 



* "Centralbl. f. Bakt. u. Parasitenk.," Sept., 1897, Bd. xxii, Nos. 10 and 11, 

 p. 287. 

 t "Jour- Boston Soc. of Med. Sci.," May, 1898, vol. 11, No. 8, p. 137. 



