Multiplex Immunochromogenic Tissue Staining Employing Primary Antibodies from the Same Species

Chromogenic immunohistochemistry (IHC) serves as an essential assay for the diagnoses of many diseases including cancer. Single-marker IHC detection is the standard used for clinical diagnostic assays. A technology to stain multiple biomarkers chromogenically on a single tissue will also yield contextual biomarker information. Methods to chromogenically stain multiple biomarkers simultaneously employing antibodies from the same species are limited and require complex protocols. Here we describe both manual and automated protocols using the UltraPlex™ mxIHC technology that allows simultaneous detection of up to three biomarkers on a single tissue using a single heat-induced antigen retrieval step in formaldehyde-fixed paraffin-embedded (FFPE) tissue and using primary antibodies from any species.