Methods of quantitative autoradiography using incident light microphotometry.

Abstract

Several different approaches of automated grain counting of microautoradiographic grain densities have been reported in the literature. Application of grain counters to cell biology is limited, however, primarily due to shortage of methods allowing the interpretation of grain counts on a molecular basis. Two suitable methods of quantitative autoradiography at the cellular level are reviewed, developed for the isotopes 14C and 125 I. They permit evaluation of absolute radioactivity in autoradiographs and, thus, determination of synthesis processes such as deoxyribonucleic acid synthesis, and of antigen densities on cell surfaces. In this approach towards quantitative autoradiography, grain densities are compared photometrically over labeled cells and over a standard source on the same autoradiograph. Allowance has to be made for the specific geometric factors of the isotope used. This can be advantageously done with an integrating type of measurement using incident light bright-field. With this type of recording, there is an exponential dependence of the photometric values on the radioactive dose. As an example of application, results are presented of the deoxyribonucleic acid synthesis rate of human myelocytes in aplastic anemia and of the immunoglobulin G density on lymphocyte membranes in the normal state and in chronic lymphocytic leukemia.