Probing the Feasibility of Single-Cell Fixed RNA Sequencing from FFPE Tissue

Area of Science:

• Genomics
• Molecular Biology
• Bioinformatics

Background:

• Single-cell RNA sequencing (scRNA-seq) offers deep insights into cellular heterogeneity but is limited by the need for fresh or frozen samples.
• Formalin-fixed paraffin-embedded (FFPE) tissues are widely available clinical archives, but their use in scRNA-seq has been challenging.
• Chromium Fixed RNA Profiling (scFFPE-seq) was developed to enable single-nucleus RNA sequencing from FFPE samples.

Purpose of the Study:

• To evaluate the efficacy of scFFPE-seq by comparing its performance against fresh tissue samples.
• To assess the ability of scFFPE-seq to recover unique cell types and capture transcriptomic information from FFPE tissues.
• To validate scFFPE-seq findings using complementary methods like Hematoxylin and Eosin (H&E) staining.

Main Methods:

• Isolation of nuclei from FFPE tissue blocks (colon, ileum, skin).
• Performance of single-nucleus RNA sequencing using the scFFPE-seq technique.
• Comparison of scFFPE-seq data with data from fresh tissue samples.
• Validation of results using H&E images.

Main Results:

• scFFPE-seq successfully recovered unique cell types from challenging FFPE tissues.
• Transcriptomic data from scFFPE-seq showed comparable cell abundance and cell type annotation to fresh tissues.
• Pathway characterization from scFFPE-seq data aligned with findings from fresh samples.
• scFFPE-seq findings were validated using H&E images.

Conclusions:

• scFFPE-seq is a viable method for generating high-quality single-nucleus RNA sequencing data from FFPE tissues.
• The technique provides comparable results to fresh tissue analysis, demonstrating its potential for scientific research.
• scFFPE-seq unlocks the potential of vast FFPE sample archives for retrospective genomic studies.