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EXTRACTION OF DNA/RNA FROM FORMALIN-FIXED, PARAFFIN-EMBEDDED TISSUE BASED ON THE ANTIGEN RETRIEVAL PRINCIPLE... 

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Figure 5.2 Comparison of immunohistochemical staining results among variable periods, 6 h to 30 days, of formalin-fixed, paraffin-embedded human breast cancer tissue (A-N), and cell line MCF-7 sections (O-Bl). All four markers, estrogen receptor (ER) (A-G), CK (cytokeratin cocktail, H-N), Her2/neu (O-U), and MIB-1 (V-Bl), showed comparable positive immunostaining results at +++ level after antigen retrieval. Original magnification x 200. Bar = 50 tm. Reproduced with permission from Shi et al., I Histochem. Cytochem. 2007 55 105-109. See color insert.

If a monoclonal antibody was generated by immunization with a full-length native protein rather than a peptide, then the immunized mouse will generate antibodies that recognize both linear and conformationally dependent epitopes. Only a small subset of these monoclonal antibodies will likely be useful for clinical use on formalin-fixed, paraffin-embedded tissue (FFPE) samples. Those that are useful tend to have epitopes that are linear the epitopes are not dependent on the protein s three-dimensional conformation (see Chapter 16). Therefore, for antibodies generated in response to immunization with full-length proteins, the peptides that serve as positive controls will be linear stretches of amino acids derived from the native protein sequence, as listed in protein databases. 

Part of contents pertaining to the protein-coated beads is reproduced with permission from our article entitled Protein-embedding technique a potential approach to standardization of immunohistochemistry for formalin-fixed, paraffin-embedded tissue sections published in J. Histochem. Cytochem. 2005,53(9) 1167-1170. 

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