Systematic variables affecting sv 40 induced t antigen expression and transformation in human cells

Kaplan, M. M.; Lubiniecki, A. S.; Blattner, W. A.; Mason, T.; Giard, D. J.; Gunnell, M.; Triantafellu, N.; Fraumeni, J. F. J.

SV40 infection of human skin fibroblasts and tumor cells resulted in the expression of T-antigen and transformed foci. By examining various conditions of input virus multiplicity and initial cell density, the systematic variation of T-antigen determination was minimized. The most uniform results were obtained at multiplicities of about 275 plaque-forming units/cells. Within limits (5 .times. 104-2 .times. 105 cells/dish), initial cell density had little effect on T-antigen expression. Volume of virus inoculum was critical for some cell lines, but not for others. Cell passage level had no general effect on T-antigen expression, although specific cell lines demonstrated increased or decreased levels of T-antigen expression with serial passage for no apparent reason. T-antigen expression correlated with virus induced cell transformation (focus formation) at 2 multiplicities. T-antigen assays at 3 days gave consistently more reproducible results than transformation assays at 21 days in 7 cell lines tested at 2 multiplicities of infection. These results defined input multiplicity as the major source of systematic variability and will permit development of a more reproducible tool in the evaluation of individuals at high risk of cancer.