A NEW METHOD TO DISTINGUISH SEPARATE COMPONENTS IN CYTOCHEMICALLY STAINED MACROMOLECULES BY ANALYSIS OF THE KINETICS OF THE STAINING AND DESTAINING PROCESSES

A new method making it possible to analyze properties of the end products of cytochemical staining reactions in macromolecular substances not expressed in the absorption spectra is described. Mathematical analysis of the course of changes in the absorbance during staining or destaining of the macromolecular substance provides information about the reaction velocities of these processes for the individual components present as well as about the concentrations of the components thus detected. This analysis is performed by a procedure of computerized curve-fitting between experimentally determined and theoretically expected absorbance curves. The theoretical time course of absorbance is derived for a number of pseudo first-order reaction schemes, and a description is given of the way in which the most appropriate reaction scheme can be recognized and the parameters are estimated, both on the basis of the experimental absorbance curve. Possible complications due to dye impurities, depolymerization and diffusion processes are discussed.