Digital quantification of somatostatin receptor subtype 2a immunostaining: a validation study.

Objective: The aim of this study was to develop an open-source and reproducible digital quantitative analysis (DIA) of somatostatin receptor subtype 2a (SST₂) staining in formalin-fixed paraffin-embedded tissues of pancreatic neuroendocrine tumors (panNETs) and growth hormone (GH)-secreting pituitary adenomas (GHomas). Design: SST₂ immunostaining of 18 panNETs and 39 GHomas was assessed using a novel DIA protocol and compared with a widely used semi-quantitative immunoreactivity score (IRS). Methods: The DIA software calculates the staining intensity/area and the percentage of positive cells (%PC). Four representative images were selected for each sample by two independent selectors (S1 and S2), with the analysis performed by two independent analyzers (A1 and A2). Agreement between observers was calculated using the concordance correlation coefficient (CCC). Results: In panNETs, the CCC ranged 0.935-0.977 for intensity/area and 0.942-0.983 for %PC. In GHomas, the CCC ranged 0.963-0.997 for intensity/area and 0.979-0.990 for %PC. In both panNETs and GHomas, the DIA staining intensity was strongly correlated with the IRS (Spearman rho: 0.916-0.969, P< 0.001), as well as the DIA %PC with the IRS %PC (Spearman rh: 0.826-0.881, P< 0.001). In GHomas, the biochemical response to somatostatin receptor ligands correlated with SST₂ expression, evaluated both as DIA intensity/area (Spearman rho: -0.448 to -0.527, P = 0.007-0.004) and DIA %PC (Spearman rho: -0.558 to -0.644, P < 0.001). Conclusions: The DIA has an excellent inter-observer agreement and showed a strong correlation with the widely used semi-quantitative IRS. The DIA protocol is an open-source, highly reproducible tool and provides a reliable quantitative evaluation of SST₂ immunohistochemistry. [ABSTRACT FROM AUTHOR]