Springer.A.Guide.to.Methods.Biomedical.Sciences.Corley.Springer.2005

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1、AGUIDETOMETHODSINTHEBIOMEDICALSCIENCESThispageintentionallyleftblankAGUIDETOMETHODSINTHEBIOMEDICALSCIENCESRONALDB.CORLEY,Ph.D.BostonUniversitySchoolofMedicineBoston,MA,USASpringereBookISBN:0-387-22845-4PrintISBN:0-387-22844-6©2005SpringerScience+BusinessMedia,Inc.Print©20

2、05SpringerScience+BusinessMedia,Inc.BostonAllrightsreservedNopartofthiseBookmaybereproducedortransmittedinanyformorbyanymeans,electronic,mechanical,recording,orotherwise,withoutwrittenconsentfromthePublisherCreatedintheUnitedStatesofAmericaVisitSpringer'seBookstoreat:http

3、://ebooks.springerlink.comandtheSpringerGlobalWebsiteOnlineat:http://www.springeronline.comThankstoTLOMLGOMDandtoallofourchildren,towhomIdedicatethis.ThispageintentionallyleftblankAcknowledgementsTherearemanypeopletothankfortheirhelpinmakingthisbookpossi-ble.Atruedebtofgr

4、atitudegoestothemanystudentsandtechnicianswhohaveworkedinmylaboratory,andmadebringingnewmethodstothelabsomuchfun.Ihavelearnedmorefromyouthanyoudidfromme.Youwouldneverhaveknownthatsomeofyourworkwouldendupinabooklikethis!ThanksalsotoAlexforhelpwithresearch,Kylleforallthepro

5、ofreading,Kateriforhelpwiththefigures,andKathyforbailingmeoutasneeded.ThispageintentionallyleftblankContents1DETECTIONANDANALYSISOFPROTEINS1A.Introduction1B.Basicmethodsforproteinanalysis3Determinationofproteincontent3Gelsandgradientsforseparationofmacromolecules4Sediment

6、ationequilibrium4Sucrosedensitygradients5Sedimentationvelocity5Subcellularfractionation5Electrophoresis6Polyacrylamidegelelectrophoresis(PAGE)6SDS-PAGE7Reducingandnon-reducingSDS-PAGE8Native(non-denaturing)PAGEgels8Isoelectricfocusing(IEF)8Two-dimensional(2D)gelelectropho

7、resis9Detectionofproteinsingels10Identificationofproteinsingelsbystaining11Coomassiebrilliantblue11PonceauS(Ponceaured)11Silverstaining11Zincstaining12Identificationofproteinsingelsusingmetaboliclabeling12Pulsechaseanalysis12Westernblotanalysis(Immunoblotting)13Chromatogr

8、aphy13Gelfiltration(or“sizeexclusion”)Chromatography14IonexchangeChromatography14Adsorptionandpa