ISOLATION AND SOME CHARACTERISTICS OF CELL NUCLEI FROM BRAIN CORTEX OF ADULT CAT
Open accessAbstract
A rapid detergent method for the isolation of nuclei from cat brain cortex is described. It involves the homogenization of the tissue in buffered 0.34 M sucrose with the addition of the non-ionic detergent Cemulsol NPT 12 and the subsequent low speed centrifugal sieving of the nuclei through two layers of sucrose (0.68 M and 1.0 M). The final purification is achieved by high speed centrifugation (40,000 g) of the nuclear suspension layered over 1.8 M sucrose. Observations by light microscopy indicate that highly purified and well preserved nuclei are obtained from neurons and glial cells. Electron microscopy reveals some microsomal contaminants adhering to the nuclear membrane. According to an analysis of the nuclear size distribution, a considerable loss of smaller nuclei (10 to 20micro(2)), mainly from glial cells, occurs during the purification procedure. The action of different detergents is compared, the best results being obtained with Cemulsol NPT 12 or Triton X-100. Chemical analyses of the purified nuclear fraction give the following content expressed in picograms per nucleus: DNA, 6.54; RNA, 2.94; cholesterol, 1.50; and protein, 97.5. The sucrose density gradient centrifugation of nuclei isolated from cat brain cortex shows that their density is equal to or higher than that of 2.2 M sucrose and is thus similar to the density of nuclei from other tissues. The observation of a varying influence of different suspending media on the density of brain cell nuclei resolves the conflicting data in the literature on the density of these nuclei.
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Protein yield after FANS is somewhat lower than expected based on the known average weight of protein per nucleus from mammalian brain (~100 pg).
“This is consistent with a somewhat lower protein yield after FANS than expected based on the known average weight of protein per nucleus from mammalian brain (~100 pg)38.”
From Neuron Enriched Nuclear Proteome Isolated from Human Brain (2013) · cites this paper as [38]
The Chemical Characteristics of Purified Nuclei section (p. 387, Table II) reports 97.5 pg of protein per nucleus, supporting the approximately 100 pg benchmark. The measurement is for purified nuclei from adult cat brain cortex, not an established average across mammalian brains. The sentence's comparison with FANS yield is the citing authors' own conclusion. (LLM, conf=0.84)
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- Authors
- Hadjiolov, A. A., Tencheva, Z. S., & Bojadjieva-Mikhailova, A. G.
- Affiliations
- Bulgarian Academy of Sciences (BG)
- Venue
- The Journal of Cell Biology
- Publisher
- Rockefeller University Press
- Published
- 1965-08-01
- Volume
- 26
- Issue
- 2
- Pages
- 383-393
- ISSN
- 0021-9525 (ISSN-L)
- 1540-8140
- PMC
- PMC2106732
- OpenAlex
- W2064213936
- Cited by
- 34
- References
- 11
- Citations by year
- 2023: 1 · 2019: 1 · 2018: 1 · 2016: 1 · 2015: 2 · 2013: 2
- FWCI
- 1.47
- Type
- journal article
- Language
- en
- Open access
- green
- www.ncbi.nlm.nih.gov
- Licence
- cc-by-nc-sa
- Locations
- The Journal of Cell Biology (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion, cc-by-nc-sa)
- PubMed Central (publishedVersion, cc-by-nc-sa)
- PubMed (publishedVersion)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Nuclear Structure and Function
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Chromatin Dynamics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
- Keywords
- cerebral cortex, cats, detergent extraction, Triton X-100, sucrose density gradient centrifugation, nuclear density, glial cells, neurons
- MeSH terms
- Animals, Cats, Cell Nucleus, Centrifugation, Cerebral Cortex, Chemistry, Cholesterol, Detergents, DNA, Microscopy, Electron, Microscopy, Phase-Contrast, Neurons, Proteins, RNA, Chemical Phenomena, In Vitro Techniques
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