The Human Cell Atlas is a large international collaborative effort to map all cell types of the human body. Single-cell RNA sequencing can generate high-quality data for the delivery of such an atlas. However, delays between fresh sample collection and processing may lead to poor data and difficulties in experimental design. This study assesses the effect of cold storage on fresh healthy spleen, esophagus, and lung from ≥ 5 donors over 72 h. We collect 240,000 high-quality single-cell transcriptomes with detailed cell type annotations and whole genome sequences of donors, enabling future eQTL studies. Our data provide a valuable resource for the study of these 3 organs and will allow cross-organ comparison of cell types. We see little effect of cold ischemic time on cell yield, total number of reads per cell, and other quality control metrics in any of the tissues within the first 24 h. However, we observe a decrease in the proportions of lung T cells at 72 h, higher percentage of mitochondrial reads, and increased contamination by background ambient RNA reads in the 72-h samples in the spleen, which is cell type specific. In conclusion, we present robust protocols for tissue preservation for up to 24 h prior to scRNA-seq analysis. This greatly facilitates the logistics of sample collection for Human Cell Atlas or clinical studies since it increases the time frames for sample processing.

中文翻译：

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scRNA-seq 评估人肺、脾和食道组织冷保存后的稳定性

人类细胞图谱是一项大型国际合作成果，旨在绘制人体所有细胞类型的图谱。单细胞 RNA 测序可以生成用于交付此类图谱的高质量数据。然而，新鲜样品采集和处理之间的延迟可能会导致数据不佳和实验设计困难。本研究评估了 72 小时内冷藏对来自 ≥ 5 名捐赠者的新鲜健​​康脾脏、食道和肺的影响。我们收集了 240,000 个高质量的单细胞转录组，其中包含详细的细胞类型注释和供体的全基因组序列，为未来的 eQTL 研究提供了可能。我们的数据为这 3 个器官的研究提供了宝贵的资源，并将允许细胞类型的跨器官比较。我们发现冷缺血时间对前 24 小时内任何组织中的细胞产量、每个细胞的读取总数以及其他质量控制指标影响不大。然而，我们观察到 72 小时时肺 T 细胞的比例下降，线粒体读数百分比升高，并且脾脏 72 小时样本中背景环境 RNA 读数的污染增加，这是细胞类型特异性的。总之，我们提出了在 scRNA-seq 分析之前组织保存长达 24 小时的稳健方案。这极大地促进了人类细胞图谱或临床研究的样品收集的物流，因为它增加了样品处理的时间范围。