Wilna A. Woods

Effect of levamisole on the in vitro immune response of spleen lymphocytes.

Stimulation of the immune system by chemical agents such as poly I:poly C (1-3), polyanions (4), tilorone (5), pyran copolymer (6), and levamisole (7-9) have been demonstrated by in vivo methods. The chemicals poly I:poly C (10) and levamisole (11) are active in vitro stimulators of spleen cell cultures, and levamisole has been shown to be a stimulator of peripheral blood lymphocytes (12). We have used the in vitro mixed lymphocyte reaction to demonstrate a direct stimulatory action of levamisole on cells involved in allogeneic recognition reactions and report here that low concentrations of levamisole induce blastogenic stimulation of spleen cell cultures as well as enhance allogeneic stimulation in mixed lymphocyte cultures. Methods and Materials. Inbred mouse strains. C57B1/6 and DBA/2 male mice, 6-8 wk of age, were used throughout the study. Levamisole. L-2, 3, 5, 6-tetrahydro-6-phenylimidazole [2, 1-b] thiazole hydro-chloride (LMS, obtained from Janssen Pharmaceuticals, Beerse, Belgium) was dissolved in distilled water at a concentration of 1 mg/ml. Appropriate dilutions were made in RPMI-1640 medium containing 20% fetal bovine serum, 100 units/ml penicillin and 1 μg/ml streptomycin (RPMI-FBS). The drug was added to cultures in 100 /xl volume at the initiation of the cultures. Mixed lymphocyte culture. The one way mixed lymphocyte reaction (MLR) was employed, using mitomycin C (75 Mg/ml final concentration) as target cell blocking agent (13). Spleens were removed and minced finely in RPMI-1640; single cell suspensions were prepared by forcing the pieces through a 16-gauge needle and passing the suspension through gauze pads. The cells were washed twice in 50 ml RPMI-1640 with centrifugation at 980 g for 5 min in a PR2 refrigerated centrifuge. Target cells were inactivated by treating an aliquot of the cell suspension with mitomycin C (Cal. Biochem., San Diego, CA) for 30 min at 37° followed by two 50 ml washes of RPMI-1640.

Effect of Actinomycin D on Growth of Rubella Virus in Tissue Cultures

Summary Multiplication of rubella virus in primary and continuous-line African green-monkey kidney cells was delayed by addition of low doses of actinomycin D to the culture medium before or at the time of infection. Addition of the drug 2 hr after infection was less effective. However, virus replication became normal after 3 to 8 days, although cellular RNA synthesis in drug-treated control cultures continued to be inhibited.

Comparison of Neutralization Rate and Agar Diffusion Methods for Intratypic Sero-Differentiation of Polioviruses.

Discussion and summary Although these data are derived from examination of only 5 strains of type 1 poliovirus they are representative of the results of a larger number of similar observations(6). It is evident that when antigenic differences were detected by the agar diffusion method they were confirmed by the neutralization rate test. Thus, the results obtained with these 2 methods of determining intratypic antigenic differences would appear to be highly comparable. When these findings are considered with those of Nakano and Gelfand(4), who showed that the neutralization rate test and their modification of the method of Wecker gave similar results, it can be concluded that in spite of the potential differences, the 3 technics are in remarkable agreement. Our results are at variance with those of McBride(2) in that a clear difference could be demonstrated between the Mahoney and LSc-2ab strains, the latter being derived from the former, whereas he found no such distinction, but employed only antiserum prepared against Mahoney. Nakano and Gelfand(4) did find the 2 viruses to be antigenically distinct, but this was most evident in tests with the antiLSc serum. The discrepancy in results in the 3 laboratories might be explained by differences in the viruses tested or more likely by differences in specificity of the antisera employed. That the latter is the probable explanation is demonstrated by the results of tests in our laboratory with anti-Mahoney serum kindly supplied by Dr. Gelfand, which were in agreement with those of Nakano and Gelfand(4) and failed to differentiate clearly LSc and Mahoney viruses. However, with suitable technics not only can the related Mahoney and LSc strains be distinguished one from the other, but alterations which occur during multiplication in the human intestine can be detected. Changes during passage in the human intestine are shown by the few data given here and further documented by more extensive observations in our laboratory(6).† Nakano and Gelfand(4) indicate that they too have demonstrated antigenic differences between the viruses excreted by vaccinated persons and the vaccine virus. Similar observations have been made by Wassermann and Fox(7). Thus, it is apparent that the antigenicity of polioviruses is not such a stable trait as had been hoped.

An immunofluorescent focus assay for gross leukemia virus.

Summary An immunofluorescent focus assay for GLV is described. GLV was demonstrated in tissues and fluids of AKR mice, as well as in tumor cells from a spontaneous lymphoma of AKR mice. A sensitive 50% focus reduction assay for neutralizing antibody to GLV is described.