Simultaneous Feulgen densitometry and autoradiographic grain counting with the Quantimet 720D image-analysis system. II. Automated grain counting.

A Quantimet 720 Image-Analysis System has been configured and programmed to enumerate silver grains over labeled nuclei in Feulgen-stained autoradiographs. The accuracy and reproducibility of the estimates have been documented in 3H-thymidine (3HTdR)-labeled rat kidney cell cultures. The Quantimet and visual grain count estimates showed excellent correlation over a wide range of counts and was independent of grain density and clustering pattern. One advantage of this approach is that the grain counts can be related to specific cellular structures. The simultaneous measurement of the mean optical density of the grain-free nucleus provides a way of evaluating 3HTdR-absorption effects for valid grain count comparisons. Using a light pen, about 800 cells are measured per hour. The grain count estimates may be made simultaneously with estimates of Feulgen-stained DNA content in the labeled nuclei. This automated technology opens new avenues for critical applications to cell cycle analysis and related problems.