Figure 1
Ab-oligo validation of representative biomarker from five phenotypic categories: tumor, immune, stromal, cell signaling, and functional. (A) Formalin-fixed paraffin embedded (FFPE) 5 µm tissue sections, and (B) a model of peripheral blood (cancer cell line mixed with peripheral blood mononuclear cells, PBMCs). Antibody or control staining pattern (green) and DAPI nuclear stain (blue) shown in all panels. Antigen detection was by indirect immunofluorescence (primary + secondary antibody, 2°), Ab-oligo plus secondary antibody, and Ab-oligo plus imaging strand (IS). Control settings included UV-cleavage after Ab-oligo + IS staining, and IS only. (B) White arrowheads designate staining on cells with antibody, or no staining on controls. (C) Graph of signal to background ratio for tissue and peripheral blood model. Ab-oligo + IS, UV-cleaved, and IS only were normalized to the IS only control in stained tumor and to the negative cell population within each validation condition for the blood model. The signal to background ratio was above 1 for all markers. Scale bar = 20 µm. αSMA, alpha smooth muscle actin; Ab-oligo, oligo conjugated antibody; CK cytokeratin, EGFR epidermal growth factor receptor, IS imaging strand, Sig:Bkgd signal to background ratio, UV ultraviolet.
