Chapter 2 58 ◀ Figure S1. Standardized identification of PD-1T TILs by flowcytometry and digital IHC analysis. (A) PD-1T expression assessed by flow cytometry in total lymphocytes. Shown is the PD-1 expression on T cells from a reference sample derived from healthy donor peripheral blood (left) and from two representative NSCLC samples with either high or low PD-1T infiltration (right). The gate for PD-1+ T cells is shown in grey and for PD-1T TILs in red. (B) Immunohistochemical (IHC) analysis of PD-1T TILs. Example of a PD-1 IHC with digital markup; the black square shows PD-1T TILs localized in a tertiary lymphoid structure (TLS). (C) Correlation of flow cytometry and digital IHC algorithm-based quantification of PD-1T TILs in 16 NSCLC tumor samples. Shown are different OD thresholds with corresponding R2 values. An OD of 0.25 was chosen as cut-off with the highest correlation, R2 = 0.615, P***<0.001. R2 and P-values were calculated using linear regression analysis. (D) Correlation curve showing normalized PD-1T values for flowcytometry (FACS) and digital IHC analysis using a cut-off of 0.25 OD.
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