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科免疫組織化學染色介紹AnIntroductionofImmunocytochemistry歡迎出席本課程

明白免疫染色的原理瞭解免疫染色的種類知道免疫染色的應用口腔病理科免疫組織化學染色介紹AnIntroducImmunohistochemistrycytochemistryAntibodyAntigenAntibodyAntigenDetectionSystemAntigenAntibody釣魚作為比喻Antibody魚餌魚Antigen浮標LabellingSystem釣竿MethodMethodFixation/FreshLabellingSystemPolyclonal-amixofIg(rabbit)Monoclonal-asingleclassofIg(mouse)Synthetic\CommercialEnzyme:HorseradishPeroxidase(HP),AlkalinePhosphatase(AP);FITC,TRITC(tetramethylrhadamineisothiocyanate);gold;isotope

Chromagen:Diaminobenzidine(DAB)(Autolysis)24hExcessiveEnzyme(trypsin)MicrowaveAutoclave免疫染色的原理ImmunohistochemistryAntibodyAnMeritsSimpleQuickCheapDrawbackHighbackgroundLowerintensityDirectMethodAntigenCytoplasmAntigenEndothelialcellNucleusLabelling(Onlyone)LabelledPrimaryAntibodyHumanRabbit/mouseanti-human2.

免疫染色的種類MeritsDrawbackLowerintensityDHigherintensityMeritsSimpleQuickCheapDrawbackHighbackgroundIndirectMethodCytoplasmAntigenAntigenEndothelialcellNucleusUnlabelledPrimaryAntibodyFirstlabellingSecondlabellingHumanRabbitanti-humanHorseanti-rabbitLabelledSecondaryAntibodyHigherintensityMeritsDrawbackPAPMethodAntigenCytoplasmAntigenEndothelialcellNucleusUnlabelledSecondary(Bridging)AntibodyPeroxidaseAnti-Peroxidase(PAP)UnlabelledPrimaryAntibodyFirstlabellingSecondlabellingThirdlabellingHigherintensityDrawback

ExpensiveTimeconsumingHumanRabbitanti-humanHorseanti-rabbitRabbitPAPPAPMethodAntigenCytoplasmAntABCMethodAntigenCytoplasmAntigenEndothelialcellNucleusUnlabelledPrimaryAntibodyHigherintensityDrawback

ExpensiveHumanRabbitanti-humanHorseanti-rabbitBiotinylatedSecondary(Bridging)AntibodyAvidin-BiotinylatedPeroxidaseComplex(ABC)ABCMethodAntigenCytoplasmAntCytoplasmNucleusAntigenAntigenEndothelialcellEliminationofEndogenousPeroxidaseLabelledPrimaryAntibodyEndogenousperoxidaseH2O2H2O2H2O2H2O2EliminateendogeousperoxidaseLabellingsystem:PeroxidaseDrawback

HighbackgroundCytoplasmNucleusAntigenAntigenCytoplasmNucleusEliminationofNon-SpecificStainingAntigenAntigenEndothelialcellAntigenAntigenEndothelialcellLabelledPrimaryAntibodyLabelledPrimaryAntibodyNormalSerum(samespeciesof2ndAb)EndothelialcellAntigenCollagenEndothelialcellLabelledPrimaryAntibodyLabelledPrimaryAntibodySpecificbindingNon-specificbindingCytoplasmNucleusEliminationofAvidin-Biotin

Peroxidase

Complex

(ABC)

methodH2O2Normalserum1oAbBiotinylated2oAbABCreagentDABstainHsuSM,1981Avidin-BiotinPeroxidaseComplPAPMethodAntigenAntigenLymphoidcellNucleusUnlabelledPrimaryAntibodyHumanRabbitanti-humanHorseanti-rabbitRabbitAPAAPUnlabelledSecondary(Bridging)Antibody(1)DoubleStainingCytoplasmUnlabelledSecondary(BridgingAntibody(2)Rabbit

PAPAPAAPPAPAlkalinePhosphasteAntiAlkalinePhosphastePAPMethodAntigenAntigenLymphComparisonofdifferentmethodsofIHCMethodsMeritsDrawbacksDirectmethodSimpleQuick

CheapHighbackgroundIndirectmethodSimpleQuickCheapHighbackgroundPAPHighintensityExpensiveTimeconsumingABCHighintensityExpensiveComparisonofdifferentmethod3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源A:CK(AE1/AE3)showsdiffusecytoplasmicstainingpatternatfullthicknessofthesquamousmucosa.

B:p63showsnuclearstainingpatternatbasaltointermediatelayerofsquamousmucosa.C:EMAshowsmembranousstainingpatternofepithelialcells.D:Vimentinshowsdiffusecytoplasmicstainingpatternonsubepithelialstromalcells.TheepitheliumisnegativeforvimentinOralmucosa(100X)3.免疫染色的應用了解細胞來源A:CK(AE1/AE33.

免疫染色的應用了解細胞來源C(SmoothMuscleActin,SMA)stainontumorcellswithmyoepithelialdifferentiation.Pleomorphicadenomaofparotidgland(400X)A:CK(AE1/AE3)stainsontumorcellswithductaldifferentiation.B(S-100protein)ontumorcellswithmyoepithelialdifferentiation.3.免疫染色的應用了解細胞來源C(SmoothMusc3.

免疫染色的應用了解細胞來源D:Vimentinshowscytoplasmicstainingpatternonsarcomatoidcarcinoma.Neuroendocrinecarcinomaofbreast(400X).E(chromogranin)andF(synaptophysin)showgranularcytoplasmicstainingpatternontumorcells.3.免疫染色的應用了解細胞來源D:Vimentinsh3.

免疫染色的應用了解細胞來源Metastaticovariancolonictypemucinouscarcinoma(400X).Carcinoembronicantigen(CEA)stainsonapicalsurfaceofadenocarcinoma.Esophagealmoderatelydifferentiatedsquamouscellcarcinoma(400X).p16showsstrongfullthicknessoverexpressiononthecytoplasmandnucleusoftumorcells.Theadjacentmucousglandisnegativeforp16.3.免疫染色的應用了解細胞來源Metastaticova3.

免疫染色的應用了解細胞來源Ovarianintestinaltypemucinousadenocarcinoma(400X).I(CK7)andJ(CK20)coexpressontumorwithheterogenouscytoplasmicstainingpattern.3.免疫染色的應用了解細胞來源Ovarianintest3.

免疫染色的應用了解細胞來源K:Colonicadenocarcinoma(400X).CK20showsdiffuseandstrongcytoplasmicstainingpatternoftumorcells.L:Malignantmelanoma(400X).HMB-45showscytoplasmicstainingpatternonmelanomacells.3.免疫染色的應用了解細胞來源K:Colonicade3.

免疫染色的應用了解細胞來源N(CD15)andO(CD30)coexpressoncellmembraneandparanuclearspaceofRScellsNodularsclerosingtypeofclassicalHodgkin’slymphomaM:LCAstainsonalllymphoidcellsincludinglargemalignantRScellsandsurroundreactivesmalllymphocytes.3.免疫染色的應用了解細胞來源N(CD15)andO3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源免疫染色的应用了解细胞来源课件Table1Antibodiesused,sourcesandtheirspecificityAntibodiesSources(dilution)SpecificityTable1Antibodiesused,sour免疫染色的应用了解细胞来源课件Conclusion-Table1Antibodiesused,theirspecificityandresultsAntibodiesSources(dilution)SpecificityResultsConclusion-Table1AntibodiesDoyouhaveanyqueries?Doyouhaveanyqueries?口

腔

病

理

科免疫組織化學染色介紹AnIntroductionofImmunocytochemistry歡迎出席本課程

明白免疫染色的原理瞭解免疫染色的種類知道免疫染色的應用口腔病理科免疫組織化學染色介紹AnIntroducImmunohistochemistrycytochemistryAntibodyAntigenAntibodyAntigenDetectionSystemAntigenAntibody釣魚作為比喻Antibody魚餌魚Antigen浮標LabellingSystem釣竿MethodMethodFixation/FreshLabellingSystemPolyclonal-amixofIg(rabbit)Monoclonal-asingleclassofIg(mouse)Synthetic\CommercialEnzyme:HorseradishPeroxidase(HP),AlkalinePhosphatase(AP);FITC,TRITC(tetramethylrhadamineisothiocyanate);gold;isotope

Chromagen:Diaminobenzidine(DAB)(Autolysis)24hExcessiveEnzyme(trypsin)MicrowaveAutoclave免疫染色的原理ImmunohistochemistryAntibodyAnMeritsSimpleQuickCheapDrawbackHighbackgroundLowerintensityDirectMethodAntigenCytoplasmAntigenEndothelialcellNucleusLabelling(Onlyone)LabelledPrimaryAntibodyHumanRabbit/mouseanti-human2.

免疫染色的種類MeritsDrawbackLowerintensityDHigherintensityMeritsSimpleQuickCheapDrawbackHighbackgroundIndirectMethodCytoplasmAntigenAntigenEndothelialcellNucleusUnlabelledPrimaryAntibodyFirstlabellingSecondlabellingHumanRabbitanti-humanHorseanti-rabbitLabelledSecondaryAntibodyHigherintensityMeritsDrawbackPAPMethodAntigenCytoplasmAntigenEndothelialcellNucleusUnlabelledSecondary(Bridging)AntibodyPeroxidaseAnti-Peroxidase(PAP)UnlabelledPrimaryAntibodyFirstlabellingSecondlabellingThirdlabellingHigherintensityDrawback

ExpensiveTimeconsumingHumanRabbitanti-humanHorseanti-rabbitRabbitPAPPAPMethodAntigenCytoplasmAntABCMethodAntigenCytoplasmAntigenEndothelialcellNucleusUnlabelledPrimaryAntibodyHigherintensityDrawback

ExpensiveHumanRabbitanti-humanHorseanti-rabbitBiotinylatedSecondary(Bridging)AntibodyAvidin-BiotinylatedPeroxidaseComplex(ABC)ABCMethodAntigenCytoplasmAntCytoplasmNucleusAntigenAntigenEndothelialcellEliminationofEndogenousPeroxidaseLabelledPrimaryAntibodyEndogenousperoxidaseH2O2H2O2H2O2H2O2EliminateendogeousperoxidaseLabellingsystem:PeroxidaseDrawback

HighbackgroundCytoplasmNucleusAntigenAntigenCytoplasmNucleusEliminationofNon-SpecificStainingAntigenAntigenEndothelialcellAntigenAntigenEndothelialcellLabelledPrimaryAntibodyLabelledPrimaryAntibodyNormalSerum(samespeciesof2ndAb)EndothelialcellAntigenCollagenEndothelialcellLabelledPrimaryAntibodyLabelledPrimaryAntibodySpecificbindingNon-specificbindingCytoplasmNucleusEliminationofAvidin-Biotin

Peroxidase

Complex

(ABC)

methodH2O2Normalserum1oAbBiotinylated2oAbABCreagentDABstainHsuSM,1981Avidin-BiotinPeroxidaseComplPAPMethodAntigenAntigenLymphoidcellNucleusUnlabelledPrimaryAntibodyHumanRabbitanti-humanHorseanti-rabbitRabbitAPAAPUnlabelledSecondary(Bridging)Antibody(1)DoubleStainingCytoplasmUnlabelledSecondary(BridgingAntibody(2)Rabbit

PAPAPAAPPAPAlkalinePhosphasteAntiAlkalinePhosphastePAPMethodAntigenAntigenLymphComparisonofdifferentmethodsofIHCMethodsMeritsDrawbacksDirectmethodSimpleQuick

CheapHighbackgroundIndirectmethodSimpleQuickCheapHighbackgroundPAPHighintensityExpensiveTimeconsumingABCHighintensityExpensiveComparisonofdifferentmethod3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源3.免疫染色的應用了解細胞來源3.

免疫染色的應用了解細胞來源A:CK(AE1/AE3)showsdiffusecytoplasmicstainingpatternatfullthicknessofthesquamousmucosa.

B:p63showsnuclearstainingpatternatbasaltointermediatelayerofsquamousmucosa.C:EMAshowsmembranousstainingpatternofepithelialcells.D:Vimentinshowsdiffusecytoplasmicstainingpatternonsubepithelialstromalcells.TheepitheliumisnegativeforvimentinOralmucosa(100X)3.免疫染色的應用了解細胞來源A:CK(AE1/AE33.

免疫染色的應用了解細胞來源C(SmoothMuscleActin,SMA)stainontumorcellswithmyoepithelialdifferentiation.Pleomorphicadenomaofparotidgland(400X)A:CK(AE1/AE3)stainsontumorcellswithductaldifferentiation.B(S-100protein)ontumorcellswithmyoepithelialdifferentiation.3.免疫染色的應用了解細胞來源C(SmoothMusc3.

免疫染色的應用了解細胞來源D:Vimentinshowscytoplasmicstainingpatternonsarcomatoidcarcinoma.Neuroendocrinecarcinomaofbreast(400X).E(chromogranin)andF(synaptophysin)showgranularcytoplasmicstainingpatternontumorcells.3.免疫染色的應用了解細胞來源D:Vimentinsh3.

免疫染色的應用了解細胞來源Metastaticovariancolonictypemucinouscarcinoma(400X).Carcinoembronicantigen(CEA)stainsonapicalsurfaceofadenocarcinoma.Esophagealmoderatelydifferentiatedsquamouscellcarcinoma(400X).p16showsstrongfullthicknessoverexpressiononthecy

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