INCREASE OF PRECISION AND ACCURACY OF DNA CYTOMETRY BY CORRECTING DIFFRACTION AND GLARE ERRORS

In recent TV-based image cytometers considerable disproportionalities exist between the IOD values of reference cells, as well as diploid, t etraploid, and octoploid analysis cells compared with their theoretica l IOD ratios. An important source of these deviations is the limited s patial resolution of the microscopic objectives, based on the effects of diffraction. Compared to these influences the glare is less importa nt. A correcting method is given for reducing both effects on the DNA measurements, which considers a narrow region along the nuclear contou r to be optically disturbed. The correction of this mean optical densi ty (MOD)- and size-dependent geometric resolution error is applicable to any cell type. The method was tested on 25 rat liver imprints and 2 9 fine needle aspirates from breast cancers. The resulting stemline ra tios are close to the theoretical ones. A further improvement was then reached by a glare correction, The slide-by-slide variations of steml ine ratios, remaining after the corrections, were considered in a test statistic for defining the aneuploidy of a given stemline. Statistica lly based clear-cut decision rules were obtained for DNA histogram int erpretation.