Innovative paraffin embedding to unlock archival blocks for highly demanding genomic analyses

Abstract DNA analysis from formalin-fixed paraffin-embedded (FFPE) tissues is frequently compromised by fragmentation and fixation-induced artifacts, limiting advanced genomic applications. While fixation chemistry has been extensively studied, the contribution of paraffin embedding to DNA damage remains poorly defined. Here, starting from a direct experimental observation of embedding-associated DNA degradation, we identify paraffin embedding as an unrecognized source of oxidative DNA damage that can be mitigated by supplementing paraffin with lipophilic antioxidants. We analysed 190 samples spanning pre-clinical and clinical settings, including 126 murine specimens and 64 human tumors processed using four fixation and embedding workflows. Using more than 180 sequencing assays — including targeted sequencing, whole-exome sequencing, shallow whole-genome sequencing, and long-read Oxford Nanopore whole-genome sequencing — we show that antioxidant-supplemented paraffin consistently preserves DNA integrity, increasing DNA integrity number (DIN) values and enriching for DNA fragments exceeding 10 kb in length. In human tumors, antioxidant-supplemented paraffin enabled robust performance across all tested genomic applications, reducing fixation-related artifacts and improving compatibility with both short- and long-read sequencing. Collectively, these findings identify paraffin embedding as a key pre-analytical determinant of molecular preservation and provide a simple, scalable strategy for enabling genomic analysis from archival pathology specimens.