A method is described for the microanalysis of protein, obtained from isolated tissue elements, in the range of 500 µµg-500 mµg. The method entails solubilization of cellular protein with phosphoric acid and heat after extraction of acid-soluble compounds, lipids, and RNA. A procedure for the extraction and recovery of cellular RNA by the use of 40% trichloroacetic acid is presented. The solubilized protein, in the form of a microdroplet, is photomicrographed with monochromatic light at 230 mµ. Total density in the microdroplet is determined from calibrated photographic plates by microdensitometry, and is converted to protein mass by using an experimentally determined average specific absorbance value. A solubilized protein labeled with tritium can be recovered after photomicrography, combusted, and reduced to generate tritiated gas for high-efficiency tritium radiometry. Total protein was analyzed in (a) nerve cells of three different sizes from Deiters' nucleus of the rabbit; and the whole rod cell and rod cell nucleus of the rabbit retina.
Article| September 01 1968
A METHOD FOR DETERMINING TOTAL PROTEIN OF ISOLATED CELLULAR ELEMENTS AND CORRESPONDING TRITIUM RADIOACTIVITY
From the Department of Physiology and Neurosensory Laboratory, State University of New York at Buffalo School of Medicine, Buffalo, New York 14214
Received: February 16 1968
Revision Received: April 17 1968
Online Issn: 1540-8140
Print Issn: 0021-9525
Copyright © 1968 by The Rockefeller University Press.
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Edward Koenig; A METHOD FOR DETERMINING TOTAL PROTEIN OF ISOLATED CELLULAR ELEMENTS AND CORRESPONDING TRITIUM RADIOACTIVITY . J Cell Biol 1 September 1968; 38 (3): 562–573. doi: https://doi.org/10.1083/jcb.38.3.562
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