An Improved Method for the Isolation of Rat Liver Nuclei by Density Centrifugation

Gill, D M

doi:10.1083/jcb.24.1.157

Cited by 16 publications

(4 citation statements)

References 11 publications

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“…They take advantage of the high density of nuclei, which is due to the large amount of nucleic acid contained therein. The nuclei are sedimented to the bottom of a tube in 2.2 ~ sucrose whereas the lighter components are floated upwards (68, 83,102). A pure preparation of thymus nuclei is shown in Figure 4.…”

Section: Nucleusmentioning

confidence: 99%

Lipids of subcellular particles

Fleischer

Rouser

1965

J Americ Oil Chem Soc

217

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Methods for isolation and characterization of subeellular particles as well as procedures for analysis of lipid class composition are discussed. The literature on distribution of lipids in subcellular particles is then reviewed. Pertinent new data from our laboratories are presented as well. The isolated particles are related to the organelles to which they correspond in the cell and are discussed with regard to heterogeneity and morphological integrity. Confusion can arise with regard to subcellular particles unless it is appreciated that: 1) preparation of particles of high purity generally requires more than the classical differential centrifugation scheme (both differential and gradient centrifugation may be required); 2) it is hazardous to apply exactly the same procedure for all tissues; 3) all subcellular fractions must be thoroughly characterized.The more recently devised DEAE cellulose column and thin-layer chromatographic procedures for analysis of lipid class composition are more reliable than the older hydrolytic or silieic acid column or paper chromatographic techniques. Endoplasmic Reticulum and Ribosomes (The Microsomal l~raction)

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Section: Nucleusmentioning

confidence: 99%

Lipids of subcellular particles

Fleischer

Rouser

1965

J Americ Oil Chem Soc

217

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“…Nuclei then sediment towards the bottom of the tube during centrifugation, but lighter components, such as cytoplasmic organelles, whole cells, erythrocytes, and nuclei with tabs of cytoplasm float to the surface. This principle was first applied in nonaqueous procedures (8)(9)(10)(11) and was subsequently adapted to aqueous methods by the use of 2.2 M sucrose (12)(13)(14)(15)(16)(17)(18)(19)(20). These methods appeared to give satisfactory preparations of nuclei, although the yields were often low (16,17) and the methods were difficult to adapt to the processing of large quantities of tissue.…”

Section: Introductionmentioning

confidence: 99%

Electron Microscopic and Biochemical Characteristics of Nuclei and Nucleoli Isolated From Rat Liver

Sadowski

Steiner

1968

The Journal of Cell Biology

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Rat liver nuclei were freed of cytoplasmic contamination by washing with Triton-X-100 and subsequent centrifugation through 2.2 M sucrose. Electron microscopic examination showed that the outer membranes of the nuclei had been removed, but that the nuclei otherwise resembled the nuclei of intact liver. Morphological studies, chemical estimations of DNA, RNA, and protein and the estimation of cytoplasmic "marker" enzymes suggested that contamination of nuclei by cytoplasmic components was limited. These nuclei were obtained in yields of about 70% and were suitable for the isolation of nucleoli. Nucleoli were isolated by the breaking of the nuclei by ultrasound and subsequent differential centrifugation. In ultrastructural appearance, the isolated nucleoli resembled nucleoli in intact tissue. However, at high magnifications the "granular" component of isolated nucleoli appeared to consist of tightly twisted fibers. The nucleoli could be obtained in yields of at least 30%, and the values for the chemical composition of the isolated nucleoli agreed with values previously reported.

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“…(b) Nuclei were prepared in 22 M sucrose pH 6 with or without addition of 3 mM CaCl 2 (Gill 1965). From the washed nuclei, albumin and globulins were removed with 014 M NaC1 and chromatine extracted in 2 M NaC1 repeatedly until the extract was no longer viscous.…”

Section: Methodsmentioning

confidence: 99%