Automated Immune Cell Quantification in EMPD

Eighteen (86%) of the 21 cases of EMPD were subjected to evaluation of CD3, CD8, and PD-1 in the immune infiltrate by automated image analysis. Three cases were excluded from automated image analysis because of metastatic disease in a lymph node (n = 1), a scant immune infiltrate (n = 1), or a small biopsy specimen (n = 1). The slides chosen were scanned at 20× magnification (Aperio ScanScope AT Turbo; Leica Biosystems). With the help of image analysis software (Aperio ImageScope), three 1 mm² squares were designated in the areas of highest density of immune cells positive for the IHC marker in question. The number of cells positive for the marker in each of the 3 squares was then counted using a modified nuclear algorithm, as described in our previous study [18 (link)]. For each tumor, the mean number of cells positive per square was then calculated to obtain a final number of positive cells per mm² for each case.