A standard tissue as a control for histochemical and immunohistochemical staining

Abstract

The variable quality of histochemical and immunohistochemical staining of tissues may be attributed to pre-analytical and analytical variables. Both categories of variables frequently are undefined or inadequately controlled during specimen collection and preparation. Pre-analytical variables may alter the molecular composition of tissues, which results in variable staining; such variations may cause problems when different tissues are used as staining controls. We developed a standard tissue for use as a staining control. Our standard tissue contains five components: 1) nine combined human cell lines mixed with stroma from human spleen; 2) a squamous cancer cell line, A431; 3) fungus; 4) transverse sections of the mosquitofish and 5) normal human spleen. The first three components were embedded in HistoGel^™ and all components were processed to paraffin and used to construct a single standard paraffin block. The muscles of mosquitofish and arteries of the spleen are positive controls for eosin staining, while other tissues are useful for assessing hematoxylin staining. The mosquitofish tissues also are excellent controls for the Masson trichrome stain and all mucin-related histochemical stains that we tested. The goblet cells of the intestine and skin stained strongly with Alcian blue, pH 2.5 (AB-2.5), mucicarmine, colloidal iron, periodic acid Schiff (PAS) or PAS-hematoxylin (PASH) and combination stains such as colloidal iron-PASH. Cell lines were not useful for evaluating histochemical stains except for PASH. The splenic stroma was a useful control for AB-2.5; however, eosin and mucin stains stained cell lines poorly, probably due to their rapid growth and associated loss of some differentiated characteristics such as production of mucins. Nevertheless, the cell lines were a critical control for immunohistochemical stains. Immunostaining of specific cell lines was consistent with the presence of markers, e.g., EGFr in DU145 cells. The cell lines expressed a wide range of markers, so they were useful controls for immunohistochemical staining including EGFr, HER2, E-cadherin, cytokeratins, Ki67, PCNA, estrogen receptor, progesterone receptor, CD3, CD20 and CD45, activated (cleaved) caspase 3 and Bcl-2. The cell lines also were a control for the TUNEL stain.

Key words::

• analytical
• variables
• histochemistry
• histology
• immunohistochemistry
• pre-analytical variables
• standard tissue

Acknowledgments

This manuscript is based upon a presentation by WEG at the Biological Stain Commission 2014 Annual Meeting. It is supported in part by the following grants: the Cooperative Human Tissue Network (1UM1CA183728), the UAB Pancreatic (2P50CA101955) and Breast (5P50CA089019) SPORES, the DOD Grant (W81XWH-10-1-0543), the UAB Comprehensive Cancer Center Core Support Grant (P30CA13148),the U54 MSM/TU/UAB Comprehensive Cancer Center Partnership (2U54CA118948) and NCI Institutional National Research Service Award (T32) (5T32CA183926-02).

Declaration of interest: The authors report no conflict of interest. The authors alone are responsible for the content and writing of this paper.