IHC Embedding Optimization

Paraffin embedded vs. frozen tissue

Once your tissue sample is fixed using the appropriate method and fixative, it is ready for sectioning and mounting. In order to preserve tissue morphology during sectioning, the tissue must be embedded in paraffin. If you are performing IHC on frozen sections, fixation occurs after sectioning. The decision to use frozen or formalin fixed paraffin embedded (FFPE) samples is dependent on which downstream applications you intend to use.

Praffin embedded Frozen
Fixation step Pre-embedding Post-sectioning
Sectioning equipment Microtome Cryostat
Stroage stability Years at room temperature One year at -20C
Advantages Tissue morphology preservation Enzyme activity preservation
Disadvantages Causes more epitope masking Ice crystals may form and disrupt tissue structure
Downstream applications DNA/RNA PCR amplification Free nuclei for FISH or cell cycle analysis
Preacautions Temperature must be monitored to avoid melting wax before deparaffinization and mounting. Tissues must be frozen and warmed slowly to avoid ice crystals and shattering, respectively.

Keywords: IHC, optimization, ICH-P, IHC-F, FFPE, frozen section, paraffin embedding, paraffin embedded vs frozen

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