Transcription of Immunohistochemistry (IHC)
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Immunohistochemistry (IHC)
Learn more | more | (IHC) Handbook Variables Influencing Experimental Design VariablesFactors*TissueSpecies, Type, Section Size and ThicknessTargetAbundance and Subcellular LocalizationEpitopeConformation Dependent Availability and Post-translational ModificationsSample PreparationParaffin-embedded or FrozenFixation MethodPerfusion or ImmersionFixativeAldehydes (Crosslinking), Alcohols (Precipitating), or AcetoneBlocking BufferNormal Serum, BSA, Casein, Various Detergents, and Salt ConcentrationsAntigen Retrieval MethodNone, Heat-induced Epitope Retrieval (HIER), and Protease-induced Epitope Retrieval (PIER)PermeabilizationTriton-X 100, Tween-20, or SaponinDetection MethodDirect, Indirect, or Signal Amplification MethodsPrimary AntibodyMonoclonal or PolyclonalSecondary AntibodySpecies Reactivity and LabelMultiplexSimultaneous or Sequential Antibody Addition, and Antibody SpeciesImaging MethodFluorescence or ChromogenicLabelFluorochromes (numerous options)Chromogenic Reagents: Enzymes (HRP, AP) and Substrates (DAB, AEC, NBT/BCIP)CounterstainFluorescence: DAPI, Hoechst 33342 Chromogenic: Hematoxylin, Fast Green FCF* The above table is not intended to be exhaustive but rather summarizes common factors influencing IHC experimental design at to IHCS ample Preparation and Fixation Key Differences of Sample Preparation Standard Fixatives What is the Difference Between Paraformaldehyde, Formaldehy
Multiplex Simultaneous or Sequential Antibody Addition, and Antibody Species Imaging Method Fluorescence or Chromogenic ... distribution, and localization. Tissues are sectioned from fixed embedded (e.g. IHC-Paraffin or plastic) or ... Use a microtome to cut the paraffin tissue blocks into 4-10 µm thick sections and transfer them to a 37°C water
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