PO-281 Automated multiparametric tissue imaging platform using existing microscope hardware for the detection of spatially resolved single-cell resolution data

Abstract

IntroductionCharacterising complexities of the tumour microenvironment is elemental to understanding disease mechanisms. The spatial relationships between infiltrating immune cells and the remodelling of the cellular matrix is widely recognised as a key component to defining tumour heterogeneity. Current methodologies for analysing the spatial dimension in tissues, like traditional immunofluorescence (IF) and immunohistochemistry (IHC), are limited to a few parameters at a time, restricting the scope of identifiable cells. Conversely, single-cell technologies like mass cytometry and NGS-based tools provide multiplexing capabilities, but at the expense of the associated spatial information. Here we present a novel multiplexed imaging technology, termed CODEX, (CO-Detection by indEXing) that combines the high-parameter space of single-cell methodologies with the spatial analysis.Material and methodsThe CODEX technology involves labelling antibodies with oligonucleotide-based tags followed by a single staining step. Over 50 parameters are simultaneously measured in a single specimen in Akoya’s novel, fully automated process. Unlike other cyclic IF approaches involving multiple antibody staining and stripping steps, the CODEX platform involves a single initial staining step and subsequent gentle manipulation of the tissue thereafter. This provides a faster workflow and prevents tissue degradation due to the harsh conditions used to remove adhered antibody. Other multiplexed imaging technologies, including imaging cytometry and MIBI, require expensive equipment precluding their routine use across various labs. The CODEX technology developed by Akoya is comprised of a fluidics instrument that interfaces with existing microscope hardware as well as a suite of associated specialised reagent offerings.Results and discussionsHere we present the results of an analysis of human tissue samples, including both fresh-frozen (FF) and formalin-fixed paraffin embedded (FFPE) specimens using the CODEX platform. Panels of antibodies were validated against a variety of markers and used to stain human tonsil, lymph node and tumour tissues. These data were processed using a custom analysis pipeline from image registration through cell segmentation. Single-cell data was analysed using clustering algorithms to define cell types.ConclusionThis CODEX study showcases the impact of high-parameter spatially resolved analysis of clinically relevant tissues enables the detection of unique cell types and cell niches within the tumour microenvironment.