Summary.-Freshly isolated clones of high cloning efficiency from a mouse fibrosarcoma were examined for DNA content, cell size, protein content, and malignant characteristics such as artificial lung-colony-forming ability, s.c. tumour take, host survival, and spontaneous metastatic ability.These malignant characteristics and other cell properties were heterogeneous among these clones; the malignant characteristics could vary and were not "all or none" in their nature. The higher the DNA content or the larger the cell volume, the higher the malignancy in terms of artificial lung-colony-forming efficiency, s.c. tumour take, and host survival.Despite variability of each parameter, the ratio of DNA content to cell size or protein content remained constant through these variations: the increased DNA paralleled increased protein and increased cell volume. The increased DNA was correlated with the more malignant characteristics of local growth and lung-colonyforming efficiency.Spontaneous metastasis to the lung was totally different from the local growth abilities; the small-cell clone produced more metastases.The graded nature of malignant properties and the differentiation between local growth and metastatic potential among the daughter clones indicate that malignancy reflects a complex moiety of cell properties.

SUMMARYA persistent infection of the human MCF-7 cell line (MCF-7RV) was established with the DBS strain of rubella virus at a low multiplicity of infection. Fluorescent antibody staining revealed that 100 % of the cells were positive for rubella antigens. The infected cells were refractory to superinfection with vesicular stomatitis virus (VSV) but were susceptible to herpes simplex virus type 2 (HSV-2). No interferon could be detected in infected cell culture fluid, and continuous passage in the presence of antibody did not lead to a decrease in the percentage of infected cells. Virus production in the persistently infected cells represented a 5-to 10-fold increase over primary infection. Plaque assays at 30 and 39 °C of the virus produced at 37 °C revealed the presence of temperature-sensitive (ts) mutants. If MCF-7RV cells were maintained at 30 °C, significant increases in virus production were observed, leading to cytopathic effect and destruction of the monolayer. If maintained at 39 °C, MCF-7RV cells produced less virus and demonstrated normal morphology.These data suggest that the naturally selected population of ts mutants being produced by these cells represents the mechanism by which persistence is maintained.

The colony-forming efficiency (CFE) of primary human tumor cells cultured in the adhesive-tumor-cell culture system (ATCCS) using Ham's F12 (F12) or Eagle's minimum essential medium, alpha modification (alphaMEM) and culture medium supplemented with either swine, equine or bovine sera were compared. AlphaMEM supplemented with equine serum provided the highest CFE of the combinations. The CFE increase due to the change from F12 to alphaMEM was approximately 5-fold, and the increase due to the change in serum from swine to equine was approximately 2-fold. Cytokeratin staining showed that this increase was not due to fibroblast growth. The high-average CFE with alphaMEM, approximately 3%, means that an inoculum of only 2 X 10(3) cells is needed to achieve formation of approximately 65 colonies in control cultures, thereby increasing the performance of this system when used in a chemosensitivity assay.

The kinetics of expression of radiation-induced micronuclei (MN) in synchronized Chinese hamster cells (CHO) was examined. The purpose of the study was to determine if the cell cycle distribution of a population significantly influences the levels of radiation induced MN, thereby obscuring the exact quantification of the radiation effect. Cells were synchronized by centrifugal elutriation, irradiated, and then different phases of the cell cycle were examined for: cell cycle progression, division probability, and temporal expression of MN. The results demonstrate that the time interval for maximal MN expression is long enough that the position of cells in the cell cycle and radiation induced division delays do not prevent the majority of cells from completing their first post-irradiation mitosis, therefore, expressing MN. By following the progression of synchronized cell populations by flow cytometry and also examining the time of division of individual cells for 24 hr after irradiation, we observed that the maximum number of cells from all phases of the cell cycle are in their first post-irradiation interphase at that time, thus explaining the MN results. Cancer Treat. Rep. 61. 425. and dose modifications in cultured cells (submitted for publication).JUNG. H. (1982) Postirradiation growth kinetics of viable and nonviable CHO cells. Radial. Res. 89.88. MEISTRICH. M.L.. MEYN. R.E. & BARLOGIE. B. (1977) Synchronization of mouse L-P59 cells by centrifugal MIDANDER J . (1 982a) Radiation-dose dependence of the formation of micronuclei in misonidazole treated cell MIDANDER. J . ( 1982b) Oxygen enhancement ratios for glutathione-deficient human fibroblasts determined from MIDANDER. J. & REVESZ, L. (1980) The frequency of micronuclei as a measure of cell survival in irradiated cell elutriation separation. Exp. Cell Res. 105. 169. cultures. Acta Radiol. Oncol. 21. 133. the frequency of radiation induced micronuclei. In(. J. Radiat. Biol. 42, 195. populations. In[. J. Radiat. B i d . 38, 231.

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