ArticleCytometry. Part A : the journal of the International Society for Analytical Cytology2014
The effect of cell subset isolation method on gene expression in leukocytes.
Article in Cytometry. Part A : the journal of the International Society for Analytical Cytology, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 42 papers.
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Who cites it
42 citing papers in PubMed, 76 citations in OpenAlex.
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- MicroRNA Expression Differences in Blood-Derived CD19+ B Cells of Methotrexate Treated Rheumatoid Arthritis Patients.Frontiers in immunology · 2021Article
- Evaluating the Effects of Cell Sorting on Gene Expression.Journal of biomolecular techniques : JBT · 2020Article
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- Clinico-Biological Implications of Modified Levels of Cytokines in Chronic Lymphocytic Leukemia: A Possible Therapeutic Role.Cancers · 2020Review
- Longitudinal RNA-Seq Analysis of the Repeatability of Gene Expression and Splicing in Human Platelets Identifies a PlateletCirculation research · 2020Article
- Use of GapmeRs for gene expression knockdowns in human primary resting CD4+ T cells.Journal of immunological methods · 2020Article
- Transcriptional Profiling of CD8+ CMV-Specific T Cell Functional Subsets Obtained Using a Modified Method for Isolating High-Quality RNA From Fixed and Permeabilized Cells.Frontiers in immunology · 2020Article
- Monocyte isolation techniques significantly impact the phenotype of both isolated monocytes and derived macrophages in vitro.Immunology · 2020Article
- Alveolar Macrophage Transcriptional Programs Are Associated with Outcomes in Acute Respiratory Distress Syndrome.American journal of respiratory and critical care medicine · 2019Article
- The Unmixing Problem: A Guide to Applying Single-Cell RNA Sequencing to Bone.Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research · 2019Review
- Histone deacetylase inhibitors induce complex host responses that contribute to differential potencies of these compounds in HIV reactivation.The Journal of biological chemistry · 2019Article
Corrections and comments
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Authors and funding
9 authors at 4 institutions in 3 countries.
Funding
Abstract
Multiple scientific disciplines require the isolation of specific subsets of blood cells from patient samples for gene expression analysis by microarray or RNA-sequencing, preserving disease- or treatment-related signatures. However, little is known with respect to the impact of different cell isolation methods on gene expression and the effects of positive selection, negative selection, and fluorescence activated cell sorting (FACS) have not previously been assessed in parallel. To address this knowledge gap, CD4+ T cells, CD8+ T cells, B cells, and monocytes were isolated from blood samples from five independent donors using positive immunomagnetic selection, negative immunomagnetic selection, and FACS. We hypothesized that positive selection and FACS would yield higher purity but may have an impact on gene expression since both methods utilize antibodies that bind surface receptors of the cell type of interest. Moreover, FACS might upregulate stress response genes due to passage of the cells through the sorter. Microarray gene expression data were generated and subjected to unsupervised clustering and differential gene expression analysis. Surprisingly, these analyses revealed that gene expression signatures were more similar between cells isolated by negative selection and FACS compared to cells isolated by positive selection. Moreover, genes that are involved in the response to stress generally had the highest expression in cells isolated by negative or positive selection and not FACS. Thus, FACS is the recommended method for isolation of leukocyte subsets for gene expression studies since this method results in the purest subset populations and does not appear to induce a stress response.
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