Standardized Processing for Formalin-Fixed, ParaffinEmbedded Cell Pellet Immunohistochemistry Controls

Positive and negative controls with known expression of target proteins are essential for the development of immunohistochemistry (IHC) assays. While tissue controls are beneficial for well-characterized proteins with defined tissue and cellular expression patterns, they are less suitable for the initial development of IHC assays for novel, poorly characterized, or ubiquitously expressed proteins. Alternatively, due to their standardized nature, cell pellets, including cancer cell lines with defined protein or transcript expression levels (e.g., high, medium, and low expression), transfected over-expressing cell lines, or cell lines with genes deleted through cell engineering technologies like CRISPR, can serve as valuable controls, especially for the initial antibody characterization and selection. In order for these cell pellets to be used in the development of IHC assays for formalin-fixed, paraffin-embedded tissues, they need to be processed and embedded in a manner that recapitulates the procedures used for tissue processing. This protocol describes a process for creating and processing formalin-fixed, paraffin-embedded cell pellet controls that can be used for IHC method developments.

Automated summary

Positive and negative controls with known protein expression are crucial in developing immunohistochemistry assays. Tissue controls are suitable for well-characterized proteins, while cell pellets, including cancer cell lines with defined protein or transcript expression levels, can serve as valuable controls for novel or poorly characterized proteins. Processed and embedded in a way that mimics tissue processing, these formalin-fixed, paraffin-embedded cell pellet controls are useful for the development of IHC methods.