Correlation of the growth of Chinese hamster V279-17lb multicellular spheroids with cytokinetic parameters

The volumes of Chinese hamster V279-17lb multicellular spheroids grown in spinner culture were measured and correlated to DNA distribution and cell volume distribution information. Spheroid growth curves were obtained over a 21-day period using both electronic cell volume measurements and microscope micrometer sizing. The spheroids were collected from the aperture tube after electronic sizing, and single cell suspensions were prepared via trypsinization. Saline-washed cells were then stained with mithramycin and analyzed as unfixed or ethanol-fixed cells for both cell volume and DNA fluorescence measurements. Spheroid growth was determined to have an initial 8- to 9-day rapid growth phase, and a slow phase from 10 to 21 days. The single cell volume distributions of cells obtained from spheroids were dependent on spheroid volume and the relative proportions of small volume noncycling cells to large volume cycling cells comprising the spheroids were also dependent on spheroid volume. DNA distribution data and dual parameter DNA distribution cell volume contour patterns obtained for single cells dissociated from spheroids were closely related to the measured spheroid volume and revealed that the fraction of small, hypoxic G0-like cells was greatest in old, large spheroids.