7 Color Multiplex Immunofluorescence Protocol for Spatial Biology with a Digital Slide Scanner

New protocol for efficient generation of seven-color, whole slide imaging with ZEISS Axioscan

Tissue and tumor sections have long been studied using immuno-detection techniques, such as immunohistochemistry (IHC). Depending on the imaging platform available, labeling of these samples was often limited either to one chromogen or three to four fluorophores, restricting researchers and drug developers in their experimental designs. Multiplex immunofluorescence (mIF) has emerged as a process that allows a higher number of fluorescent probes on a single tissue section, permitting in-depth comprehensive studies to identify specific proteins, cell types, spatial biology studies, and more.

Dr. Caroline Bouzin is the facility manager of the 2IP Imaging Platform at the Institute of Experimental and Clinical Research at the UCLouvain, Brussels, Belgium. She has worked with oncologists and pulmonologists to fine-tune a protocol for customized mIF stainings on formalin-fixed paraffin-embedded (FFPE) tissue samples followed by data collection with the ZEISS Axioscan digital slide scanner. A combination of signal amplification (TSA) with carefully selected filtersets matched to a panel of seven fluorophores results in the fast generation of ready-to-analyze digital images without the need for image post-processing, such as spectral unmixing or tissue alignment. This workflow is equipping researchers in her facility for high-throughput, spatial biology analyses, such as the immune microenvironment of tumors.