Abstract
The presence of complement-fixing antibodies has been described in the serum of individuals infected with any of a number of central nervous system viruses, 1 as well as a method for their determination that eliminates all non-specific reactions. A recent human epidemic of encephalitis was promptly diagnosed as due to Western equine encephalomyelitis virus by means of the complement-fixation reaction. 2
The antigens prepared for the complement-fixation test as previously described were virulent. In order to diminish the risk involved in the general use of such antigens, attempts were made to render them avirulent without materially destroying their antigenicity. This result has been accomplished by exposing the virus preparations to ultraviolet light for suitable length of time, as we shall describe.
The antigens were prepared in the following manner: Batches of 4-weeks-old W-Swiss mice were injected intracerebrally with one of the viruses; when prostrate these animals were sacrificed, their brains removed, weighed, and emulsified in 0.85% saline containing 2% normal guinea pig serum heated at 56°C for 20 minutes, the proportion being 1 g of infected brain to 10 cc of diluent. This emulsion was kept in the icebox for 20 hours and then centrifuged in a horizontal centrifuge at 2500 rpm for a half hour; the supernatant was frozen and thawed 5 times in a dry ice-alcohol mixture, and finally centrifuged in an angle head centrifuge 3 at 5000 rpm for 1 hour. The supernatant, following addition of merthiolate in a dilution of 1/10,000, constituted the antigen.
Antigens thus prepared with the viruses of rabies, Eastern and Western equine encephalomyelitis, lymphocytic choriomeningitis, and St. Louis encephalitis have been tested for virulence within 1 to 3 days of preparation by intracerebral injection into Swiss mice and found virulent in dilutions of 10-4 for Western equine encephalomyelitis, 10-3 for Eastern equine encephalomyelitis, 10-2 for rabies, lymphocytic choriomeningitis, and St. Louis encephalitis viruses respectively.
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