International Journal of Cell Cloning, Vol 10, 344-351, Copyright © 1992 by AlphaMed Press
An in vitro thymidine incorporation assay for human cancers: technical details revisited
K Sugihara, LA Collins, HD Homesley and CE Welander
Bowman Gray School of Medicine, Winston-Salem, North Carolina.
Among the choices of laboratory methods for in vitro testing of human tumor
chemosensitivity, a thymidine incorporation assay (TIA) has been described.
Advantages of a TIA over a colony-forming assay include greater probability
of tumor growth, requirement for fewer tumor cells, less dependence on an
absolute single-cell suspension, shorter time to completion, and the
ability to measure up to three logs cell-kill. Specific technical details
which are reported in the literature for performing a TIA are incomplete
and/or conflicting between one laboratory and another. This paper reports
details of methods designed to remove unbound 3[H]-thymidine from the
agarose, standardize background counts, and determine optimal cell-plating
densities, labeling time, optimal concentration of 3[H]-thymidine, and the
best day after seeding cultures to add the 3[H]-thymidine. Care has been
taken to compare the end point of drug sensitivity determinations following
these methodologic changes, both among serial TIAs as well as with
colony-forming assays.
