191 Pre-analytic variables and multiplex immunofluorescence assays

Pre-analytic variables in formalin-fixed paraffin embedded (FFPE) specimens are a large barrier to assay reliability, and the characterization potential correction of such represent unmet need dermatopathology. Multiplex immunofluorescence (mIF) is an emerging modality, which tissue-sparing also allows for assessment marker co-expression intensity. The purpose this study was two-fold: 1) characterize FFPE block age as pre-analytic variable, 2) determine whether novel pan-membrane (PM) could be used normalization. We tissue microarrays (TMAs) constructed from archival 125 patients with melanoma collected over 20-year period. TMAs were stained mIF panel (FoxP3, CD8, PDL1, PD1, Tumor, CD163), PM cocktail, DAPI nuclei, imaged using Vectra Polaris, analyzed HALO Multi-IF module evaluate impact on cell density Cell densities did not vary significantly 5/6 markers, but FoxP3+ decreased (-1.5%/yr, p<0.001). Marker intensities showed several small shifts age: CD163 (+0.5%/yr), tumor (+1.3%/yr), (+0.82%/yr) increased (p<0.001), while PD-L1 (-0.26%/yr, p<0.01). Overall, these data suggest that most there changes respect age; however, more rare types FoxP3, may become noticeable age. next considered beyond evaluated stain might serve proxy immunoreactivity. Linear regressions performed intensity vs All markers except positive regression coefficients (p<0.001). This suggests our useful surrogate immunoreactivity allow normalization between individual specimens, correcting variation.