A non-destructive X-ray fluorescence method of analysis of formalin fixed-paraffin embedded biopsied samples for biomarkers for breast and colon cancer

Abstract

In this work we present a methodology for the non-destructive elemental determination of formalin-fixed paraffin-embedded (FFPE) human tissue samples based on the Fundamental Parameters method for the quantification of micro Energy Dispersive X Ray Fluorescence (micro-EDXRF) area scans. This methodology intended to overcome two major constraints in the analysis of paraffin embedded tissue samples – retrieval of optimal region of analysis of the tissue within the paraffin block and the determination of the dark matrix composition of the biopsied sample. This way, an image treatment algorithm, based on R® tool to select the regions of the micro-EDXRF area scans was developed. Also, different dark matrix compositions were evaluated using varying combinations of H, C, N and O until the most accurate matrix was found: 8% H, 15% C, 1% N and 60% O for breast FFPE samples and 8% H, 23% C, 2% N and 55% O for colon. The developed methodology was applied to paired normal-tumour samples of breast and colon biopsied tissues in order to gauge potential elemental biomarkers for carcinogenesis in these tissues. The obtained results showed distinctive biomarkers for breast and for colon: there was a significant increase of P, S, K and Fe in both tissues, while a significant increase of Ca an Zn concentrations was also determined for breast tumour samples.