Publications

Spatial Analysis: Framework for Interrogating the Immune Landscape in Prostate Cancer

Recently, our laboratory validated a multiplex immunohistochemistry (IHC) approach. We evaluated two main methods, multiplex immunofluorescence, and multiplex iterative chromogenic IHC and have standardized on the iterative IHC approach. The advantages of this latter system are: (1) the ability to use any primary antibody; (2) the ability to use relatively inexpensive commercial whole slide brightfield scanners; (3) the ability to perform q/c for each round of staining using brightfield whole slide images; (4) the lack of a requirement for fluorescent imaging; (5) the lack of need to spectrally separate potentially overlapping fluorescent colors, and (6) the ability to use off-the-shelf commercial software for the analysis (HALO by Indica Labs). In this study Busra Ozbek, M.D., employed a 7-antibody 8-pseudo-color assay to characterize several T cell phenotypes including classical T helper (CD3+CD4+ cells), classical cytotoxic T cells (CD3+CD8+ cells) and for each of these whether they expressed the marker PD1, which is a key drug target. She also characterized regulatory T cells (CD3+CD4+FOXP3+). This overall approach is modular can apply to any IHC‐validated antibody combinations and sets the groundwork for more detailed spatial analyses.