Comparison of DNA histograms by standard flow cytometry and image cytometry on sections in Barrett's adenocarcinoma
Several mathematical algorithms have been published so far to correct histograms obtained from DNA-measurements on thin sections. All these methods require knowledge of the thickness of the section currently under investigation. As the problem of determining the actual section thickness has recently been solved, the different methods for recalculation can be compared under strictly controlled conditions. For histogram recalculation, the algorithms published by Bins, McCready, Bacus, Rigaut and Haroske are applied on a series of rat liver tissue sections with increasing section thicknesses. The histograms are compared to that of measurements of a single cell preparation of the same tissue block. Our results suggest that DNA-measurements on thin paraffin sections are possible if the actual section thickness is known. Correct ploidy equivalents throughout the whole spectrum of section thicknesses investigated are calculated using the formulas of McCready and Haroske. This is also the case using the correction method of Bins in very thin sections. The algorithms of Bacus and Rigaut do not lead to reliable results. Recalculated histograms show some differences to those obtained by DNA-measurements of single cell preparations. The histograms show broader CVs of the peaks and lymphocytes as internal ploidy standard are not useful in the majority of the algorithms. Nevertheless the promising results of this study legitimize efforts to establish standards for the interpretation of recalculated histograms in the near future. The broader applicability to tumour sections remains unproven. Therefore, further investigation are necessary using more heterogeneous tissue including aneuploid tumour cell populations.