Visible and near-infrared hyperspectral imaging techniques allow the reliable quantification of prognostic markers in lymphomas: a pilot study using the Ki67 proliferation index as an example.

ABSTRACT In this study, we examined the suitability of visible and infrared (Vis-NIR) hyperspectral imaging (HSI) for the quantification of prognostic markers in Non-Hodgkin Lymphoma (NHL) on the example of the Ki67 proliferation index. Ki67 quantification was done on six follicular lymphomas (FL) and 12 diffuse large B-cell lymphomas (DLBCL) by applying classical immunohistochemistry. The Ki67 index was comparatively assessed visually, using HSI-based quantification and a digital imaging analysis (DIA) platform. There was no significant difference between visual assessment (VA), DIA, and HSI in FL. For DLBCL, VA resulted in significantly higher Ki67 values than HSI (p = 0.023) and DIA (p = 0.006). No such difference was seen comparing analysis by HSI and DIA (p = 0.724). Cohens Kappa showed a “substantial correlation” of Ki67 values for HSI and DIA in FL and DLBCL (Kappa: 0.667 and 0.657). Here we provide the first evidence that, comparably to traditional DIA, HSI can be used reliably to quantify protein expression, as exemplified by the Ki67 proliferation index. By covering the near-infrared-spectrum, HSI might offer additional information about the biochemical composition of pathological specimens, although our study could not show that HSI is clearly superior to conventional DIA. However, the analysis of multiplex immunohistochemistry might benefit from such an approach, especially if overlapping immunohistochemical reactions were possible. Further studies are needed to explore the impact of this method on the analysis and quantification of multiple marker expression in pathological specimens.