The pathogenesis of lymphocytic choriomeningitis (LCM) virus infection of mice has been subjected to intensive study by virus titration and histopathologic techniques (1-5). There are a number of unresolved questions relating to the pathogenesis of the various types of infection which require study of the distribution of virus at the cellular, rather than at the whole organ level. This report describes a direct immunofluorescent antibody test for LCM antigen, and its application to study of cellular distribution of infection in acutely and congenitally infected mice. Materials and MethodsMice.--Weanling mice of the HA/ICR strain were purchased from Charles River Mouse Farms, Brookline, Massachusetts. Latent LCM infection was not detected in these mice.LCM Virus.--The CA 1371 (mouse brain-passaged) and WCP (viscerotropic) strains of LCM virus, and the procedures for preparation of virus stocks have been described previously (6). Swiss mice congenitally infected with LCM virus were from the colony of carrier mice initiated by Haas (7); these mice were in approximately the 41st generation as carriers.Virus Titrations.--Virus suspensions (clarified by centrifugation at 1800 Rmt for 10 minutes) and serial 10-fold dilutions were made in medium 199 with 3 per cent inactivated calf serum. Materials were held in an ice bath, and inoculations made as soon as possible after preparation of suspensions. Weanling mice were inoculated intracerebrally with 0.03 ml, using 6 mice per dilution. Survivors were rechallenged at 14 days postinoculation with approximately 1000 LD60 of strain CA 1371 LCM virus, and held an additional 14 days. In. fectivity titers, based on typical death or immunity to challenge as the endpoint, were calculated according to the method of Reed and Muench, and are expressed as the logt0 of the number of IDs0 per 0.03 mi of 10 per cent suspension. Because of the high degree of pathogenicity of the CA 1371 strain following intracerebral inoculation, immunity tests were not considered necessary to evaluate infectivity titration endpoints with this strain.Tissue Cultures.--Primary cultures of trypsinized kidney ceils of monkey (rhesus and Cercopitkecus), rabbit, and Syrian hamster were used, as well as WI-26 cells and primary cultures of HA/ICR mouse embryo. All cells were grown on 9 x 35 mm coverslips in Leighton tubes. 829

The immunological properties of the purified 15,000-dalton protein of Rauscher murine type-C virus were analyzed by radioimmunoassay. The majority of the antigenic determinants of this protein were found to be remarkably specific to Rauscher and Friend virus and to a lesser extent to Molonev virus. Determinants reactive with other murine viruses (group-specific) or type-C viruses of other species (interspecies) were also demonstrated but were minor components of the total antigenic specificities of the protein. The results provide evidence that the antigenic properties of this protein specify the Friend-Moloney-Rauscher subgroup of type-C viruses.

MATERIALS AND METHODS Media. Cells were grown in Dulbecco's modification of Eagle medium supplemented with 10% calf serum (Colorado Serum Co.) in 50-mm petri dishes (Falcon Plastics, Los Angeles). Cells and viruses. The control cell lines used included BALB/3T3 (3) and normal rat kidney (NRK) cells (8). The Schmidt-Ruppin strain of Rous sarcoma virus (SR-RSV) subgroup D (5) was used to transform BALB/3T3 and NRK. Two weeks after infection with this avian tumor virus, focal areas of transformed cells were visible; these were selected by means of cloning cylinders. The resulting mixed cultures of normal and transformed cells were cloned in microtiter plates (34). The SR-RSV-transformed lines obtained were designated SR-BALB and SR-NRK. A B77 avian sarcoma virus (36) transformant of BALB/3T3, B77-BALB, was provided by P. Vogt. This line was also cloned prior to use. The Kirsten (Ki-) and Rauscher (R-) strains of murine leukemia virus (MuLV) were propagated as previously described (34). The Carr-Zilber strain of RSV and Rousassociated viruses, RAV-1 and RAV-2, were obtained from the Resources and Logistics Segment, National Cancer Institute (NCI). Purification of avian gs antigen. Chicken serum containing high-titered avian myeloblastosis virus (AMV) was provided by J. W. Beard, Duke University, through the Resources and Logistics Segment, NCI. After clarification by centrifugation at 10,000 rpm for 15 min, the virus was pelleted at 30,000 rpm for 90 min, banded in a 15 to 60% sucrose gradient, and then repelleted. After resuspension and dialysis overnight against 0.01 M Tris (pH 7.8)-0.1 M 893