QUANTIFICATION OF RELATIVE AREA OF PS2 IMMUNOHISTOCHEMICAL STAINING AND EPITHELIAL PERCENTAGE IN BREAST CARCINOMAS - THE EFFECT OF THE LATTER ON THE INTERPRETATION OF A CYTOSOLIC PS2 ASSAY

In immunochemical assays of specific cell constituents in cytosols from tumors, the relationship between epithelial and stromal fractions is not taken into account. This may influence the outcome of the measurements and result in incorrect categorization as negative or positive. In a setting addressing pS2 (only detectable in epithelial cells) in breast carcinomas, we investigated three possibilities that may overcome this problem using histologic sections of breast carcinomas of 50 patients: (a) visual estimation of area percentage of immunohistochemical staining of the cell constituent of interest performed by three independent individuals, (b) quantification of area percentage of the immunohistochemical results by true color image analysis, and (c) quantification of the epithelial and stromal compartments of the tumors in Heidenhain's-azan-stained tissue sections, using the true color image analysis system, to assess the epithelial percentage in the tumors. This percentage was used as a correction factor for data on pS2 obtained by cytosolic determinations. Visual estimation appeared to be subject to interobserver variation and, subsequently, becomes less applicable in the absence of strict scoring rules. Based on tests for correlation, image analysis system quantification seemed reproducible in both quantification procedures. However, due to the high magnification necessary to visualize the immunohistochemical staining product, the effect of field selection caused systematic differences between repeat measurements (Friedman test). As a result of the contrasting colors of the azan staining, the calculation of the epithelial percentage could be performed at a low magnification. Consequently, here the effect of field selection was not present. Correction of cytosolic values for epithelial percentage resulted in 8% of the cases changing category. In conclusion, the combined results of the two disciplines (histology and biochemistry) may provide more information than either of them alone.