Phytohemagglutinin, an extract of the kidney bean, Phaseolus vulgaris, induces in human leukocyte cultures an inhibitor of the cytopathic effects of Sindbis virus. The physicochemical and biological properties of this virus-inhibitor are similar to those of interferon induced by Newcastle disease virus, except for an instability at pH 2 and 10 and at 56 degrees C.
Subcutaneous implantation of DBA/2-derived L5178Y cells into DBA/2 mice followed 10 d later by nodule excision protected 100% of mice from the rapid outgrowth of an intraperitoneal challenge of L5178Y cells given 7 d postexcision. Challenged mice remained clinically normal for 48--250 d before onset of an ultimately fatal tumor outgrowth. The numbers of L5178Y cells in the peritoneal cavity increased logarithmically for 4 d after challenge and then declined to low but detectable levels which persisted throughout the clinically normal period. Cells active in 18-h in vitro cytolytic assays against 51Cr-labeled L5178Y target cells were found in the peritoneal cavity. The effector cells were determined to be Thy1.2 positive. Their activity was tumor specific and reached peak levels 4 d after tumor challenge and then gradually declined to undectable levels during the following 70 d. Tumor emergence occurred most frequently during the period when CMC activity was no longer demonstrable in the remaining clinically normal mice. A transient peak of low level cytophilic antitumor antibody was detected about 30 d after tumor cell challenge. The temporal associations between the numbers of tumor cells and the levels of cell-mediated lysis against L5178Y cells indicate the importance of the cell-mediated cytolysis response in limiting initial tumor outgrowth and suggest its role as one of the factors responsible for long-term tumor suppression during tumor dormancy.
The replication of vesicular stomatitis virus in human leukocyte cultures shows that virus yields can be enhanced 6-to 180-fold by treating the leukocyte cultures with phytohemagglutinin prior to virus inoculation. Data suggest that a substance is produced in phytohemagglutinin-treated leukocyte cultures which is capable, on transfer to fresh leukocytes, of inducing blast cell formation and enhancing virus replication.
The present communication reports a new procedure for measurement of infective virus, based on the counting of individual infected cells which are identified by staining of viral antigen with fluorescent antibody. In this procedure, complete monolayers of susceptible cells are used and the very cells which become primarily infected by the inoculated virus are visualized and counted.In recent years the plaque technique has been widely adopted for the measurement of infective virus (1, 2). However, this technique, and certain modifications thereof (3, 4), are not applicable to virus-cell systems in which the spread of virus from cells infected initially to neighboring cells does not readily occur.Some work has been reported endeavoring to base infectivity titrations on recognizable changes in single cells without requiring successive cycles of cell infection. Deibel and Hotchin (5) determined the proportion of influenza virus-infected cells in a smear of a cell suspension by means of the fluorescent antibody technique, and were thus able to calculate the infectivity titer of the virus seed. Marcus and Puck (6) used a quantitative plating technique for mammalian cells to measure Newcastle disease virus (NDV). Their virus assay procedure is based on the fact that NDVinfected HeLa ceils are unable to produce macroscopic colonies.The new quantitative procedure for measurement of infective NDV in HeLa ceils is characterized by two features: the time required for assay of infective virus is short, and the experimental arrangement used is suitable for both virus assay and conduct of experiments involving observations on cells.In this communication, the linear relationship between the concentration of virus in the inoculum and the number of fluorescent cells in the first cycle of multiplication is described. The precision and sensitivity of the procedure are also described. Results of studies on microepidemiology of NDV in HeLa cells are reported, and brought to bear on the counting procedure. The significant advantages of the new procedure are discussed.
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