ANALYSIS OF CELL NUCLEI WITH INTERFERENCE-MICROSCOPE AND CYTOPHOTOMETRY .I. DETERMINATION OF GLOBULINS

The globulinfraction of cell nuclei was determined by dry mass determinations in comparison with cytophotometric measurements. Lymphocytes from the thymus gland, and liver nuclei from the rat and kidney nuclei from the pig, isolated with Behrens' technique, were extracted in smears, up to 30 hrs, in physiological saline and tyrode solution. The decrease of nuclear dry mass was measured with the interference microscope, and the content of DNA, histones and total proteins was determined by cytophotometry (Feulgen-reaction, staining with gallocyanin chrome alum, fast green at pH 8 and pH 2, naphtol yellow S). Using glycerol as embedding medium, after extraction with 0,14 M saline at 3-5° C the dry weight of liver nuclei decreased by 18% after 2 hrs, by 29% after 10 hrs and by 34% after 30 hrs. Nuclear proteins were extracted up to 2 hrs, nucleo-histones were removed after 10 hrs. Since the loss of proteins is in good agreement with the globulin content of liver nuclei, obviously the nuclear globulins have been extracted until 2 hrs. After extraction with tyrode solution at room temperatue, the loss of dry matter was in the same range (2 hrs: 12%, 10 hrs: 20%, 30 hrs: 29%). Using tyrode solution as embedding medium, the dry weight of kidney nuclei remained unchanged for 2 hrs when extracted with 0,14 M saline, since the globulin-fraction is lost while nuclei are measured by interferometry. With thymocytes only a loss of 18% of nuclear dry weight was found after 24 hrs due to removal of DNA and histones. Suspending the thymus gland in 0,14 M saline, the nuclear dry weight increased by 40% in 4 hrs, probably due to adsorption of proteins. The results were discussed in the light of the errors of the measurements and compared to biochemical and histochemical datas. © 1968 Springer-Verlag.