The Case for Static Microdensitometry

It is often difficult to obtain unequivocal data for genome size in cell systems where the available sample size is too small to yield a reliable estimate based on biochemical assays and counts of cell numbers. It is particularly difficult to resolve DNA levels per cell when tissue samples contain mixtures of diploid cells at various stages of the cell cycle and in tissues consisting of cells with polyploid or polytene chromosomes. Similar problems can often plague analysis of such complex tissue samples by flow cytometry using DNA-specific fluorescent probes in cell systems where the available sample size is too small and/or heterogeneous to yield reliable estimates of DNA amounts per cell.

In these cases, it has been productive to resort to static cytophotometry, and most recently to static, image analysis microdensitometry (12), employing the DNA-Feulgen reaction with a scanning and integrating cytophotometer of tested accuracy and precision (38-41) to determine integrated optical density (IOD), which can also be expressed as relative integrated absorbance (RIA), of the Feulgen-DNA dye complex bound to individual nuclei. From such data, it is feasible to obtain reliable estimates of genome size or C values for a diverse array of eukaryotic organisms (11,12,16).

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