Abstract
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.
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Selected References
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- EDSTROM J. E., KAWIAK J. Microchemical deoxyribonucleic acid determination in individual cells. J Biophys Biochem Cytol. 1961 Mar;9:619–626. doi: 10.1083/jcb.9.3.619. [DOI] [PMC free article] [PubMed] [Google Scholar]
- EDSTROM J. E. Quantitative determination of ribonucleic acid in the micromicrogram range. J Neurochem. 1958 Oct;3(1):100–106. doi: 10.1111/j.1471-4159.1958.tb12614.x. [DOI] [PubMed] [Google Scholar]
- HYDEN H., EGYHAZI E. Changes in the base composition of nuclear ribonucleic acid of neurons during a short period of enhanced protein production. J Cell Biol. 1962 Oct;15:37–44. doi: 10.1083/jcb.15.1.37. [DOI] [PMC free article] [PubMed] [Google Scholar]
- HYDEN H. Quantitative assay of compounds in isolated, fresh nerve cells and glial cells from control and stimulated animals. Nature. 1959 Aug 8;184:433–435. doi: 10.1038/184433a0. [DOI] [PubMed] [Google Scholar]
- KOENIG E. A QUANTITATIVE MICRO METHOD FOR DETERMINATION OF SPECIFIC RADIOACTIVITIES OF H3-PURINES AND H3-PYRIMIDINES. Anal Biochem. 1963 Nov;6:424–434. doi: 10.1016/0003-2697(63)90095-2. [DOI] [PubMed] [Google Scholar]
- KOENIG E. SYNTHETIC MECHANISMS IN THE AXON. II. RNA IN MYELIN-FREE AXONS OF THE CAT. J Neurochem. 1965 May;12:357–361. doi: 10.1111/j.1471-4159.1965.tb04236.x. [DOI] [PubMed] [Google Scholar]
- Koenig E. Observations on selected isolated retinal elements and an analysis of rod cell RNA of the rabbit. J Cell Biol. 1967 Jul;34(1):265–274. doi: 10.1083/jcb.34.1.265. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Koenig E. Synthetic mechanisms in the axon. IV. In vitro incorporation of [3H]precursors into axonal protein and RNA. J Neurochem. 1967 Apr;14(4):437–446. doi: 10.1111/j.1471-4159.1967.tb09542.x. [DOI] [PubMed] [Google Scholar]