Single Nuclei Isolation
Single Nuclei Isolation - Related Videos
05:59
Isolation of Nuclei from Human Intermuscular Adipose Tissue and Downstream Single-Nuclei RNA Sequencing
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Cited by 2 •
2024
The biology of intermuscular adipose tissue (IMAT) is largely unexplored due to the limited accessibility of human tissue. Here, we present a detailed protocol for nuclei isolation and library preparation of frozen human IMAT for single nuclei RNA sequencing to identify the cellular composition of this unique adipose depot.
09:09
Isolation of Nuclei from Flash-Frozen Liver Tissue for Single-Cell Multiomics
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Cited by 6 •
2022
Here, we present a protocol for isolating nuclei from flash-frozen, archived liver tissues for single-nucleus RNA-seq, ATAC-seq, and joint multiomics (RNA-seq and ATAC-seq).
06:00
Nuclei Isolation from Adult Mouse Kidney for Single-Nucleus RNA-Sequencing
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Cited by 10 •
2021
Here, we present a protocol to isolate high-quality nuclei from frozen mouse kidneys that improve the representation of medullary kidney cell types and avoids the gene expression artifacts from enzymatic tissue dissociation.
07:12
A Simple, Quick, and Partially Automated Protocol for the Isolation of Single Nuclei from Frozen Mammalian Tissues for Single Nucleus Sequencing
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Cited by 2 •
2023
The study describes a simple, quick, and partially automated protocol to isolate high-quality nuclei from frozen mammalian tissues for downstream single nuclei RNA sequencing.
10:03
Nuclei Isolation from Mouse Cardiac Progenitor Cells for Epigenome and Gene Expression Profiling at Single-Cell Resolution
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Cited by 5 •
2023
Here, we present a protocol describing cell nuclei preparation. After microdissection and enzymatic dissociation of cardiac tissue into single cells, the progenitor cells were frozen, followed by isolation of pure viable cells, which were used for single-nucleus RNA sequencing and the single-nucleus assay for transposase-accessible chromatin with high-throughput sequencing analyses.
FAQs
Mechanical disruption is intended to break the plasma membrane while leaving the nucleus intact. This distinction allows nuclear contents to be recovered without requiring complete preservation of the original cell. The released nuclei can then enter a stabilizing buffer, creating a suspension suitable for downstream genetic and molecular analyses rather than losing fragile cellular material during whole-cell dissociation.
These steps improve the quality of the nuclear suspension in different ways. Stabilizing buffer receives the nuclei after release, filtration removes larger debris, and density-based centrifugation can enrich the nuclear fraction. Together, they reduce unwanted material in the preparation and produce a more suitable input for molecular profiling, including analyses of DNA, chromatin, or RNA.
The approach reduces dependence on fresh tissue and can be applied to frozen or archived material, according to the overview. It is also useful when tissue structure or cell fragility makes whole-cell dissociation problematic. By analyzing nuclei rather than requiring intact cells, the method can reduce effects associated with dissociation and preserve access to cell-type-resolved genetic information.
An isolated nuclear suspension can support several molecular readouts, including DNA, chromatin, and RNA profiling. These measurements can be used for single-nucleus sequencing and for examining gene expression, epigenetic states, and genomic variation. In genetics, the major value is the ability to connect these features with individual cellular or nuclear populations rather than interpreting the tissue only as a bulk sample.
A typical workflow begins with mechanical disruption of the tissue or cells to break the plasma membrane. The released nuclei are transferred into a stabilizing buffer, passed through a filter to remove debris, and may then undergo density-based centrifugation for enrichment. The resulting suspension becomes the preparation for downstream DNA, chromatin, RNA, or single-nucleus sequencing analyses.
The core components described are the starting tissue or cells, a mechanical disruption step, stabilizing buffer, a filter, and equipment for density-based centrifugation when enrichment is needed. Their sequence matters: nuclei must be released before filtration and possible density separation. The resulting preparation is intended to limit debris and provide a usable input for molecular and genetic assays.
It is particularly valuable when researchers need cell-type-resolved information from frozen, archived, or structurally complex samples, or when fragile cells are poorly suited to whole-cell dissociation. Single-nucleus analyses can reveal differences in gene expression, epigenetic states, and genomic variation across nuclear populations, extending genetic profiling to samples that are difficult to process as intact cells.