PDF(Pubmed)
Abstract：
OBJECTIVE: Transcriptional profiling of immune cells is an indispensable tool in biomedical research; however, heterogenous sample types routinely used in transcriptomic studies may mask important cell type-specific transcriptional differences. Techniques to isolate desired cell types are used to overcome this limitation. We sought to evaluate the use of immunomagnetic B cell isolation on RNA quality and transcriptional output. Additionally, we aimed to develop a B cell gene signature representative of a freshly isolated B cell population to be used as a tool to verify isolation efficacy and to provide a transcriptional standard for evaluating maintenance or deviation from traditional B cell identity.
RESULTS: We found RNA quality and RNA-sequencing output to be comparable between donor-matched PBMC, whole blood, and B cells following negative selection by immunomagnetic B cell isolation. Transcriptional analysis enabled the development of an 85 gene B cell signature. This signature effectively clustered isolated B cells from heterogeneous sample types in our study and naïve and memory B cells when applied to transcriptional data from a published source. Additionally, by identifying B cell signature genes whose functional role in B cells is currently unknown, our gene signature has uncovered areas for future investigation.
RESULTS: We found RNA quality and RNA-sequencing output to be comparable between donor-matched PBMC, whole blood, and B cells following negative selection by immunomagnetic B cell isolation. Transcriptional analysis enabled the development of an 85 gene B cell signature. This signature effectively clustered isolated B cells from heterogeneous sample types in our study and naïve and memory B cells when applied to transcriptional data from a published source. Additionally, by identifying B cell signature genes whose functional role in B cells is currently unknown, our gene signature has uncovered areas for future investigation.
摘要：
目的：免疫细胞转录谱是生物医学研究中不可或缺的工具；然而，转录组学研究中常规使用的异源样品类型可能掩盖重要的细胞类型特异性转录差异。使用分离所需细胞类型的技术来克服该限制。我们试图评估免疫磁性B细胞分离对RNA质量和转录输出的使用。此外,我们的目的是开发一种代表新鲜分离的B细胞群体的B细胞基因标签,作为验证分离效果的工具,并提供一种转录标准,用于评估维持状态或与传统B细胞身份的偏离.
结果：我们发现与供体匹配的PBMC之间的RNA质量和RNA测序输出相当，全血,和通过免疫磁性B细胞分离阴性选择后的B细胞。转录分析能够开发85基因B细胞特征。当应用于来自已发表来源的转录数据时，该特征有效地聚集了我们研究中来自异质样品类型的分离的B细胞，以及初始和记忆B细胞。此外,通过鉴定B细胞特征基因，其在B细胞中的功能作用目前尚不清楚，我们的基因签名揭示了未来调查的领域。
结果：我们发现与供体匹配的PBMC之间的RNA质量和RNA测序输出相当，全血,和通过免疫磁性B细胞分离阴性选择后的B细胞。转录分析能够开发85基因B细胞特征。当应用于来自已发表来源的转录数据时，该特征有效地聚集了我们研究中来自异质样品类型的分离的B细胞，以及初始和记忆B细胞。此外,通过鉴定B细胞特征基因，其在B细胞中的功能作用目前尚不清楚，我们的基因签名揭示了未来调查的领域。
