GLANDERS 129 
country both the concentrated mallein and a 5 to 7% solution of the 
precipitated mallein (Mallein Siccum Foth) are used. The ordinary 
mallein used for subcutaneous test cannot be relied upon. 
This test should not be made in the presence of a conjunctivitis. 
It is also important, where there is a reaction, that the purulent dis- 
charge be not wiped out of the eye thereby leaving one in doubt. Like 
the subcutaneous method, there is no definite relation between the 
extent of the reaction and the amount of pathological changes that 
exist. 
The cutaneous and intradermal application of mallein do not give 
uniformly satisfactory results. 
The agglutination method or serum diagnosis. Rabieaux found that 
the difference which exists between the agglutinating power of a 
serum from a glandered and from a healthy horse may be used as the 
basis of a method for diagnosing glanders. He collected the serum as 
pure as possible, diluted it with sterile, distilled water to 1 in 10, or to 
1 in 500. The diluted serum was then mixed in a small sterile tube 
with an equal volume of a 24 to 72 hour culture of Bact. mallei in 
peptonized bouillon (without glycerin). The mixture was placed in 
an incubator at a temperature of 35° to 37° C. and examined at 
variable times under the microscope. In dilutions of from 1 in 10 
to 1 in 50 the agglutination occurred in 20 minutes to 3 hours. In 
serum of a non-glandered horse from 2 to 6 hours were required to 
produce the agglutination. In weaker dilutions the differences were 
more marked. The development of the method can be followed from 
the writings of M’Fadyean, Bourget and Méry, Arpad, Fedorowsky, 
Reinecke, Bonome, Schiitz and Miessner, Schntirer and Moore, 
Taylor and Giltner. 
The method consists in the preparation of a test fluid from a suitable 
culture of Bact. mallet to which is added the diluted serum. 
The “‘test-fluid” is prepared by washing the growth from a 72 hour 
acid-agar culture by the aid of a sterile wire loop into distilled water 
containing 0.85 per cent. sodium chloride and 0.5 per cent. carbolic 
acid crystals. This suspension is then placed in a thermostat at 60° C 
for two hours, which kills the bacteria. Three cubic centimeters of 
the “‘test-fluid”’ are placed in each of several small test-tubes. Witha 
sterile pipette, the diluted serum is added to the tubes of test-fluid 
and thoroughly mixed. In making the different dilutions, the amount 
of diluted serum to be used is readily ascertained by the following 
table: : 
