In Vitro Determination of Thymidine-³H Labeling Index in Human Solid Tumors

A rapid and reproducible method for the preparation of autoradiographs from single-cell suspensions of human tumor cells, including melanoma and lung carcinoma, is described. Tritiated thymidine of high specific activity (6.0 Ci/mmole) was used so that autoradiographs could be developed and read at 24 hr after the sample was taken. Samples were prepared in duplicate, and a Hypaque-Ficoll density gradient was used to separate viable tumor cells from dead tumor cells, red blood cells, and tissue debris. Autoradiographs were prepared from single-cell suspensions of the viable tumor cells recovered from each split-sample fraction. The labeling index percentage (the number of labeled tumor cells per 100 tumor cells counted) was determined from the autoradiograph of each sample fraction, and the average of the two labeling indices was considered the labeling index percentage of the sample. The labeling index percentage was reproducible when compared with that of the other one-half. A twofold difference between labeling indices could be declared significant at p < 0.05. The values obtained for the labeling indices of a variety of tumors, including melanoma and oat cell carcinoma, were comparable to those reported using other methods, including in vivo labeling. Our data support previous observations that the labeling index varies little in simultaneous biopsy specimens from the same patient, at least in melanoma. The method described lends itself well to the serial study of the labeling index as a measure of the proliferative fraction of tumors obtained from patients with accessible disease.