基础生物化学实验法.pdf

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1、基礎生物化學實驗法(1)IntroductiontotheBiochemistryLaboratory(2)GeneralLaboratoryProcedures(3)化學實驗操作法(4)免疫化學分析之各種方法(5)原子吸光法之基本原理及測定1IntroductiontotheBiochemistryLaboratory2GeneralLaboratoryProcedures345化學實驗操作法(2008.7.9)I基本操作1.粉碎2.混合3.溶解4.加熱5.冷却(1)液體之冷却(2)固體之冷却(3)冷却劑6.攪拌7.減壓8.蒸發9.乾燥(1)器具之乾燥(

2、2)乾燥劑(3)液體之乾燥(4)固體之乾燥10.蒸留II分離操作1.抽出(1)水及有機溶劑之相互溶解度(2)以分液漏斗抽出(3)以自働抽出器抽出2.鹽析3.過濾(1)普通過濾(2)減壓過濾6ExtractionandisolationofpurecompoundsExtraction:Itiscustomarytoseparatethecomponentsofapreviouslyunknowncompoundsbyextractionwithsolventsonincreasingpolarity.Withdriedmaterialthiscanbepr

3、oceedbysublimationandsteamdistillationonasample,followedbyextractionwiththeseries:lightpetroleum,ether,chloroform,ethanol(or80%aqueousethanol),andwater(successivelycold,warm,acidandalkaline).Thefurtherfractionationoftheextractobtainedisamatterwhichhastobeplannedaccordingtotherequi

4、rementsofagivingsituationbearinginmindthepropertiesofthecompoundstobeextracted.Exceptwhereisolationofessentiallyonecompoundbycrystallization,distillation,orsublimationispossible,thegreatvarietyofchromatographic,electrophoresis,andmultipledistributiontechniquesplayvitalpartinsubseq

5、uentoperations.7heptaneVI水(多価alcohol,…)(hydrocarbons,CS2,CCl4…)IVIICHCl3alcoholCHCl2(methanol,phenol…)CH3CHCl2,CH2Cl、CH2Cl…IIIether,ketonealdehyde…溶媒之分類完全混合不完全混合(少量互溶)不能混合8免疫組織化學方法研究胞外母質及其受體(Immunohistochemicaltechniquestostudytheextracellularmatrixanditsreceptors)基本方法書1.Review材料及

6、方法1.Antibodies(Methodsin1.Zn-bindingprotein抗體(Harlow&Lane,Enz.Vol.245,2.市售抗體1988)pp.316-346)CollagenI2.Usingantibodies,2.參考文獻CollagenIII1998)○1染色laminin,CollagenIV3.專書nidogen,及LamininImmunochemistrycollagentypeIV3.市售oftheextracellular○2collagensImmunohisto-matrix,Vol.I,○3fibroblas

7、t之chemicalkitVol.IImarkerthy-14.poly-lys4.MethodsinEnz.○4精子5.96孔盤6.螢光顯微鏡9免疫化學分析之各種方法1.染色方法(1)Immunohistochemical(IHC)methoda.horseradishperoxidase-labeledreagentsb.alkalinephosphate-labeledreagents(2)Immunofluororescence(IF)method2.immunoassay(ELISA)3.染色材料(1)stainingcells(2)staini

8、ngtissues(3)Immunoblotting10原子吸光法