Heuston Elisabeth F, Doumatey Ayo P, Naz Faiza, Islam Shamima, Anderson Stacie, Kirby Martha R, Wincovitch Stephen, Dell'Orso Stefania, Rotimi Charles N, Adeyemo Adebowale A
Center for Research on Genomics and Global Health, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
Genomic Technology Section, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD, USA.
Commun Biol. 2025 Jan 4;8(1):10. doi: 10.1038/s42003-024-07445-2.
Single cell studies have transformed our understanding of cellular heterogeneity in disease but the need for fresh starting material can be an obstacle, especially in the context of international multicenter studies and archived tissue. We developed a protocol to obtain high-quality cells and nuclei from dissected human skeletal muscle archived in the preservative Allprotect® Tissue Reagent. After fluorescent imaging microscopy confirmed intact nuclei, we performed four protocol variations that compared sequencing metrics between cells and nuclei enriched by either filtering or flow cytometry sorting. Cells and nuclei (either sorted or filtered) produced statistically identical transcriptional profiles and recapitulated 8 cell types present in skeletal muscle. Flow cytometry sorting successfully enriched for higher-quality cells and nuclei but resulted in an overall decrease in input material. Our protocol provides an important resource for obtaining high-quality single cell genomic material from archived tissue and to streamline global collaborative efforts.
单细胞研究改变了我们对疾病中细胞异质性的理解，但对新鲜起始材料的需求可能成为障碍，特别是在国际多中心研究和存档组织的背景下。我们开发了一种方案，用于从保存在Allprotect®组织试剂中的解剖人类骨骼肌中获取高质量的细胞和细胞核。在荧光成像显微镜确认细胞核完整后，我们进行了四种方案变体，比较了通过过滤或流式细胞术分选富集的细胞和细胞核之间的测序指标。细胞和细胞核（分选或过滤的）产生了统计学上相同的转录谱，并概括了骨骼肌中存在的8种细胞类型。流式细胞术分选成功富集了更高质量的细胞和细胞核，但导致输入材料总体减少。我们的方案为从存档组织中获取高质量单细胞基因组材料和简化全球合作努力提供了重要资源。
