Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation
Summary: High-quality single-cell RNA-sequencing data of liver cells, especially hepatocytes, are challenging due to cell death associated with hepatocyte isolation using fluorescence-activated cell sorting (FACS). Here, we present a protocol to obtain viable hepatocytes and nonparenchymal liver cel...
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| Format: | Article |
| Language: | English |
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Elsevier
2022-12-01
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| Series: | STAR Protocols |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666166722007043 |
| _version_ | 1798006911748014080 |
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| author | Simeng Wang |
| author_facet | Simeng Wang |
| author_sort | Simeng Wang |
| collection | DOAJ |
| description | Summary: High-quality single-cell RNA-sequencing data of liver cells, especially hepatocytes, are challenging due to cell death associated with hepatocyte isolation using fluorescence-activated cell sorting (FACS). Here, we present a protocol to obtain viable hepatocytes and nonparenchymal liver cells for scRNA-seq, using centrifugation. We detail steps for liver wash and enzyme perfusion, followed by in vitro dissociation of liver cells and gradient centrifugation. We further describe hepatocytes harvesting for subsequent viability check and scRNA-seq.For complete details on the use and execution of this protocol, please refer to Wang et al. (2021) and Mederacke et al. (2015). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. |
| first_indexed | 2024-04-11T13:02:01Z |
| format | Article |
| id | doaj.art-c43e6358f9104fe0bc2941cb4aec4ce4 |
| institution | Directory Open Access Journal |
| issn | 2666-1667 |
| language | English |
| last_indexed | 2024-04-11T13:02:01Z |
| publishDate | 2022-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | STAR Protocols |
| spelling | doaj.art-c43e6358f9104fe0bc2941cb4aec4ce42022-12-22T04:22:54ZengElsevierSTAR Protocols2666-16672022-12-0134101824Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugationSimeng Wang0Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390-9046, USA; Corresponding authorSummary: High-quality single-cell RNA-sequencing data of liver cells, especially hepatocytes, are challenging due to cell death associated with hepatocyte isolation using fluorescence-activated cell sorting (FACS). Here, we present a protocol to obtain viable hepatocytes and nonparenchymal liver cells for scRNA-seq, using centrifugation. We detail steps for liver wash and enzyme perfusion, followed by in vitro dissociation of liver cells and gradient centrifugation. We further describe hepatocytes harvesting for subsequent viability check and scRNA-seq.For complete details on the use and execution of this protocol, please refer to Wang et al. (2021) and Mederacke et al. (2015). : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.http://www.sciencedirect.com/science/article/pii/S2666166722007043GenomicsRNAseqMetabolismModel OrganismsMolecular Biology |
| spellingShingle | Simeng Wang Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation STAR Protocols Genomics RNAseq Metabolism Model Organisms Molecular Biology |
| title | Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation |
| title_full | Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation |
| title_fullStr | Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation |
| title_full_unstemmed | Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation |
| title_short | Protocol to obtain high-quality single-cell RNA-sequencing data from mouse liver cells using centrifugation |
| title_sort | protocol to obtain high quality single cell rna sequencing data from mouse liver cells using centrifugation |
| topic | Genomics RNAseq Metabolism Model Organisms Molecular Biology |
| url | http://www.sciencedirect.com/science/article/pii/S2666166722007043 |
| work_keys_str_mv | AT simengwang protocoltoobtainhighqualitysinglecellrnasequencingdatafrommouselivercellsusingcentrifugation |