Application of X-ray microanalysis to the study of histochemical staining reactions

The application of X-ray microanalysis to the study of histochemical staining reactions is reviewed. Advantages of the technique are the ability to examine colourless intermediate reaction products, and to study the efficiency of their conversion to the final reaction product. The number of X-rays emitted by an element is, under appropriate conditions, proportional to the amount of that element. Errors such as glare and deviations from Beer's Law in microdensitometry and quenching of fluorescence in microfluorimetry, are avoided. The possibility of analysing several elements simultaneously is a further advantage, as the amount of reaction product can be related to the quantity of some other cellular constituent. The application of X-ray microanalysis to the study of the binding of the fluorochrome quinacrine to nuclei is described. The extinction of this fluorochrome is too low for it to be measured by microdensitometry. X-ray microanalysis shows that its fluorescence is not proportional to the amount of quinacrine bound, and thus fluorescence cannot be used for this study. Quinacrine was shown to be strongly bound to nuclei, and the strength of its affinity is consistent with intercalative binding.