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 Substrat
The label, either fluorescent or enzymatic, is used to visualize the antigen-antibody complex. In this multi-step application, staining may be influenced by multiple variables and requires the careful optimization of new assays for each target antigen. For example, when investigating a high abundance protein in formaldehyde-fixed tissue, the IHC
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