LEADER 05015nam 2200565 a 450 001 9910455468303321 005 20200520144314.0 010 $a1-283-66474-7 010 $a0-19-159161-0 010 $a0-585-48392-2 035 $a(CKB)111087026789928 035 $a(EBL)1043143 035 $a(OCoLC)815671912 035 $a(MiAaPQ)EBC1043143 035 $a(Au-PeEL)EBL1043143 035 $a(CaPaEBR)ebr10612575 035 $a(EXLCZ)99111087026789928 100 $a19970311d1997 uy 0 101 0 $aeng 135 $aur|n|---||||| 200 00$aSubcellular fractionation$b[electronic resource] $ea practical approach /$fedited by J.M. Graham and D. Rickwood 210 $aOxford ;$aNew York $cIRL Press at Oxford University Press$dc1997 215 $a1 online resource (360 p.) 225 1 $aThe practical approach series 300 $aDescription based upon print version of record. 311 $a0-19-963494-7 320 $aIncludes bibliographical references and index. 327 $aCover; Contents; List of contributors; Abbreviations; 1. Homogenization of tissues and cells; 1. Introduction; 2. Aims of the homogenization procedure; 3. Influence of sample type; 4. Homogenization media; 5. Methods of homogenization; Type 1 homogenizers; Type 2 homogenizers; 6. Homogenization of tissues and cells; Mammalian liver; Brain; Muscle; Mammalian tissue culture cells; Plant organelles; Yeast; Other fungi and algae; Trypanosomes; Bacteria; References; 2. Isolation of subcellular fractions; 1. Introduction; 2. Composition of a tissue homogenate; 3. Properties of cell organelles 327 $aFactors affecting organelle density and size4. Centrifugal methods for the separation of organelles; Separation by size; Separation by density; Density perturbation; 5. Non-centrifugal procedures; Immunoisolation; Separation by electrophoresis; 6. Identification of separated material; Marker enzymes; Introduced markers for endocytic and exocytic pathways; Characteristic non-enzymatic proteins; 7. Assessment of the purity of fractions; Purity and purification; Problems from cell heterogeneity within tissues; Problems arising from organelle fragmentation 327 $aMissorting in the exocytic and endocytic pathways8. Fractionation problems; No separation; Aggregation following resuspension of a fraction; Poor recovery of markers; Damage to cell structures; 9. A systematic approach to cell fractionation; Preliminary studies; Determination of the properties of components of the homogenate; Method development; Simplification of the separation; References; 3. Isolation and characterization of nuclei and nuclear subfractions; 1. Introduction; 2. Methods of preparing purified nuclei; Types of cells and tissue samples; Homogenization media 327 $aHomogenization methodsCentrifugation conditions; Assays of nuclear purity; 3. Methods for purifying metaphase chromosomes; 4. Isolation of nuclear subfractions; Preparation of nucleoli; Preparation of nuclear membranes; Isolation of nuclear matrix; Preparation of nucleoids; 5. Isolation of nucleoprotein complexes; Isolation of polynucleosomes of chromatin; Ribonucleoproteins; 6. Isolation of nuclear macromolecules; Isolation of nuclear proteins; Isolation of nuclear RNA; Isolation of DNA; 7. Functional assays of nuclei; Analysis of DNA-binding proteins; Transcription assays; References 327 $a4. Subcellular fractionation of mitochondria1. Introduction; 2. Purification of mitochondria from various eukaryotic sources; Introduction; Protocols for purification of mitochondria from several eukaryotic sources; Further purification of mitochondrial fractions; 3. Determination of mitochondrial purity; Introduction; Use of the oxygen electrode to determine mitochondrial integrity; Determination of the integrity of the mitochondrial outer membrane; Glucose hexokinase trap method for estimation of P:O ratios; 4. Subfractionation of mitochondria 327 $aPreparation of submitochondrial particles by sonication 330 $aMany investigations into the structure and function of cells and tissues require the isolation of a particular membrane or subcellular component (organelle). This book covers all the necessary aspects, from breaking up the cells (homogenization), via a variety of separation techniques (the isolation and fractionation chapters), to characterization of the separated organelles. 410 0$aPractical approach series. 606 $aSubcellular fractionation$vHandbooks, manuals, etc 606 $aSubcellular fractionation$vLaboratory manuals 608 $aElectronic books. 615 0$aSubcellular fractionation 615 0$aSubcellular fractionation 676 $a571.6/5 701 $aGraham$b J. 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