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Published on: July 28, 2023
Protocol for isolating cells from a single cryopreserved gastrointestinal endoscopic biopsy for single-cell RNA
Daniel Horowitz1, Joshua Rusbuldt1, Lynn Tomsho1
1Johnson & Johnson, Immunology, Translational Sciences & Medicine, Spring House, PA, USA.
STAR Protocols
|May 21, 2026
Summary
This study introduces a new protocol for isolating viable cells from cryopreserved gastrointestinal endoscopic biopsies. This method enables single-cell RNA sequencing (scRNA-seq) analysis of previously inaccessible tissue samples.
Area of Science:
- Gastroenterology
- Molecular Biology
- Bioinformatics
Background:
- Existing methods struggle to isolate viable cells from cryopreserved intestinal tissues.
- There is a need for protocols to analyze cellular material from preserved endoscopic biopsies.
Purpose of the Study:
- To present a detailed protocol for isolating single cells from cryopreserved gastrointestinal endoscopic biopsies.
- To enable downstream single-cell RNA sequencing (scRNA-seq) applications.
Main Methods:
- Protocol involves thawing and washing cryopreserved biopsies.
- Enzymatic digestion and cell purification steps are detailed.
- Includes dead cell removal, cell recovery quantification, and 10X Genomics (GEM) library preparation.
Main Results:
- Successful isolation of viable cells from cryopreserved duodenal and colonic endoscopic biopsies.
- The protocol is optimized for single-cell RNA sequencing.
Conclusions:
- This protocol provides a reliable method for obtaining single cells from cryopreserved gastrointestinal biopsies.
- Facilitates comprehensive cellular and molecular analysis of preserved intestinal tissues.
