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Updated: Aug 8, 2025
Isolation of Human Neutrophils from Whole Blood and Buffy Coats
Published on: September 17, 2021
Protocol for bulk RNA sequencing of enriched human neutrophils from whole blood and estimation of sample purity
Anna L K Gonye1, Thomas J LaSalle2, Samuel S Freeman3
1Center for Cancer Research, Department of Medicine, Massachusetts General Hospital, Boston, MA, USA; Broad Institute of MIT and Harvard, Cambridge, MA, USA; Cellular and Molecular Medicine Graduate Program, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Abstract:
Although neutrophils are the most abundant leukocyte in healthy individuals and impact outcomes of diseases ranging from sepsis to cancer, they remain understudied due to technical constraints of isolation, preservation, and sequencing. We present a modified Smart-Seq2 protocol for bulk RNA sequencing of neutrophils enriched from whole blood. We describe steps for neutrophil isolation, cDNA generation, library preparation, and sample purity estimation via a bioinformatic approach. Our approach permits the collection of large cohorts and enables detection of neutrophil transcriptomic subtypes. For complete details on the use and execution of this protocol, please refer to LaSalle et al. (2022)1 and Boribong et al. (2022).2.
