Nuclear Isolation from Cryopreserved In Vitro Derived Blood Cells

Area of Science:

• Stem cell biology
• Hematopoiesis
• Genomics and epigenomics

Background:

• Induced pluripotent stem cells (iPSCs) are valuable for studying blood development and creating cell therapeutics.
• Multiomics technologies like single nucleus RNA sequencing (snRNAseq) and snATACseq hold promise for advancing developmental biology.
• Isolating intact nuclei from diverse cell types, especially cryopreserved samples, presents significant technical hurdles for multiomics studies.

Purpose of the Study:

• To provide a detailed, robust protocol for isolating high-quality nuclei from iPSC-derived hematopoietic cells at various developmental stages.
• To address the technical challenges associated with nuclear isolation from different iPSC-derived cell types (adherent stromal/endothelial and non-adherent hematopoietic progenitors).
• To facilitate the use of these isolated nuclei in downstream single-nucleus multiomics workflows.

Main Methods:

• Development and optimization of a nuclear isolation protocol specifically for iPSC-derived cells.
• Focus on isolating nuclei from both adherent stromal/endothelial and non-adherent hematopoietic progenitor cells.
• Incorporation of troubleshooting steps to minimize nuclear clumping and debris.

Main Results:

• Successful isolation of high-quality nuclei from iPSC-derived hematopoietic cells across different developmental stages.
• The protocol effectively handles distinct cell types, including challenging adherent and non-adherent populations.
• Minimized nuclear clumping and debris, ensuring sufficient quantity and quality for downstream multiomics analyses.

Conclusions:

• The presented method provides a reliable approach for obtaining high-quality nuclei from iPSC-derived hematopoietic cells for multiomics.
• This protocol overcomes common technical barriers, enhancing data quality and accessibility for researchers.
• The adaptable method holds potential for isolating nuclei from other cryopreserved cell types for diverse research applications.