SEMINAR

Scalable single nucleus RNA sequencing of heavily-fixed or FFPE tissues by fixative exchange (FX)-seq

Chang Ho Sohn

Yonsei University

Recent advances in single cell analyses by next-generation sequencing have broaden our understanding about how complex biological systems work at single cell level. Yet, current single cell RNA sequencing methods require fresh or freshly frozen samples for analysis. Excluding recently released 10x fixed cell kit and visium kit, most commercial platforms are unable to detect transcripts from cells and tissues heavily fixed by paraformaldehyde, which conventionally requires for sample preservation. Tremendous clinical samples which contain rich patient disease information are available in the form of FFPE but it is mostly incompatible with the current single cell sequencing methodologies for transcript detection. Here we report a new method via fixative exchange-seq, allowing us to directly detect mRNAs from heavily PFA fixed samples and FFPE slides or blocks by successful in situ synthesis of cDNAs. Using FX-seq, we demonstrated its utility and versatility by analyzing a total of 320k nuclei from from PFA-fixed and FFPE-treated mouse brain and human cancer tissue. Overall, FX-seq offers the potential to overcome the limitations of current tools by analyzing single-nucleus transcriptomes in archived clinical FFPE samples