Easily and efficiently isolate highly purified human neutrophils from fresh human peripheral blood leukocyte samples by immunomagnetic negative selection, with the EasySep™ Human Neutrophil Isolation Kit. Widely used in published research for more than 20 years, EasySep™ combines the specificity of monoclonal antibodies with the simplicity of a column-free magnetic system.
In this EasySep™ negative selection procedure, unwanted cells are labeled with antibody complexes and magnetic particles. The magnetically labeled cells are then separated from the untouched desired neutrophils by using an EasySep™ magnet and simply pouring or pipetting the cells into a new tube. Following magnetic cell isolation in as little as 14 minutes, the desired neutrophils are ready for downstream applications such as flow cytometry, culture, or DNA/RNA extraction. The isolation kit is compatible with cells prepared using HetaSep™ (Catalog #07906) sedimentation or red blood cell lysis.
This product replaces the EasySep™ Human Neutrophil Enrichment Kit (Catalog #19257), for even faster cell isolations.
Learn more about how immunomagnetic EasySep™ technology works or how to fully automate immunomagnetic cell isolation with RoboSep™. Alternatively, choose ready-to-use, ethically sourced, primary Human Peripheral Blood Neutrophils, Frozen isolated with EasySep™ Human Neutrophil Isolation Kit. Explore additional products optimized for your workflow, including culture media, supplements, antibodies, and more.
Figure 1. Typical EasySep™ Human Neutrophil Isolation Profile
Starting with whole blood prepared using HetaSep™ or Lymphoprep™ with RBC lysis, the neutrophil content (CD45+CD16+CD66b+) of the isolated fraction typically ranges from 98.7 ± 0.9% (mean ± SD). In the above example, the purities of the start and final isolated fractions are 52.0% and 99.0%, respectively.
This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.
Dysfunction of low-density neutrophils in peripheral circulation in patients with sepsis. R. Sun et al. Scientific reports 2022 jan
Abstract
Low-density neutrophils (LDNs) have been described in tumors and various autoimmune diseases, where they exhibit immune dysfunction and alter disease progression. Nevertheless, LDNs have been rarely reported in sepsis. We studied sepsis patients admitted to the intensive care unit. Wright-Giemsa stain assay and Transmission electron microscopy were performed to detect the morphology of neutrophils. Flow cytometry was used to analyze the number and function of LDNs. Concentration of cytokines was measured using ELISA. Neutrophil chemotaxis was examined using an under-agarose chemotaxis model. We found that LDNs were significantly elevated in patients with sepsis. Phenotypes and morphological characteristics suggest that LDNs may be formed by mixtures of neutrophils at various maturation stages. In vitro experiments showed that LDN formation was closely associated with neutrophil degranulation. We preliminarily discussed changes in immune function in LDNs. Compared with high-density neutrophils, expression levels of CXC chemokine receptor 4 on LDN surfaces were increased, phagocytotic capacity was decreased, and life span was prolonged. The chemotactic ability of LDNs was significantly reduced, possibly related to the increased expression of P2X1. These data suggest that LDNs are essential components of neutrophils in sepsis. To clarify the source and dysfunction mechanism of LDN in sepsis may be helpful for the diagnosis and treatment of sepsis in the future.
Exosomal transfer of activated neutrophil-derived lncRNA CRNDE promotes proliferation and migration of airway smooth muscle cells in asthma. X.-Y. Zhang et al. Human molecular genetics 2022 feb
Abstract
Activated neutrophil-derived exosomes reportedly contribute to the proliferation of airway smooth muscle cells (ASMCs), thereby aggravating the airway wall remodeling during asthma; however, the specific mechanism remains unclear. Lipopolysaccharide (LPS)-EXO and si-CRNDE-EXO were extracted from the media of human neutrophils treated with LPS and LPS??+??si-CRNDE (a siRNA targets long non-coding RNA CRNDE), respectively. Human ASMCs were co-cultured with LPS-EXO or si-CRNDE-EXO, and cell viability, proliferation and migration were measured. The interplay of colorectal neoplasia differentially expressed (CRNDE), inhibitor of nuclear factor kappa B kinase subunit beta (IKK$\beta$) and nuclear receptor subfamily 2 group C member 2 (TAK1) was explored using RNA immunoprecipitation (RIP) and Co-IP assays. A mouse model of asthma was induced using ovalbumin. CRNDE was upregulated in LPS-EXO and successfully transferred from LPS-treated neutrophils to ASMCs through exosome. Mechanically, CRNDE loaded in LPS-EXO reinforced TAK1-mediated IKK$\beta$ phosphorylation, thereby activating the nuclear factor kappa B (NF-$\kappa$B) pathway. Functionally, silencing CRNDE in LPS-EXO, an IKK$\beta$ inhibitor, and an NF-$\kappa$B inhibitor all removed the upregulation of cell viability, proliferation and migration induced by LPS-EXO in ASMCs. In the end, the in vivo experiment demonstrated that CRNDE knockdown in neutrophils effectively reduced the thickness of bronchial smooth muscle in a mouse model for asthma. Activated neutrophils-derived CRNDE was transferred to ASMCs through exosomes and activated the NF-$\kappa$B pathway by enhancing IKK$\beta$ phosphorylation. The latter promoted the proliferation and migration of ASMCs and then contributed to airway remodeling in asthma.
Sputum from patients with primary ciliary dyskinesia contains high numbers of dysfunctional neutrophils and inhibits efferocytosis. M. Blanter et al. Respiratory research 2022 dec
Abstract
BACKGROUND Primary ciliary dyskinesia (PCD) is a genetic disorder characterized by recurrent airway infection and inflammation. There is no cure for PCD and to date there are no specific treatments available. Neutrophils are a crucial part of the immune system and are known to be dysfunctional in many inflammatory diseases. So far, the role of the neutrophils in PCD airways is largely unknown. The purpose of this study was to investigate the phenotype and function of airway neutrophils in PCD, and compare them to blood neutrophils. METHODS Paired peripheral blood and spontaneously expectorated sputum samples from patients with PCD (n??=??32) and a control group of patients with non-PCD, non-cystic fibrosis bronchiectasis (n??=??5) were collected. The expression of neutrophil-specific surface receptors was determined by flow cytometry. Neutrophil function was assessed by measuring the extent of actin polymerization, production of reactive oxygen species (ROS) and release of neutrophil extracellular traps (NETs) in response to activating stimuli. RESULTS Sputum neutrophils displayed a highly activated phenotype and were unresponsive to stimuli that would normally induce ROS production, actin polymerization and the expulsion of NETs. In addition, PCD sputum displayed high activity of neutrophil elastase, and impaired the efferocytosis by healthy donor macrophages. CONCLUSIONS Sputum neutrophils in PCD are dysfunctional and likely contribute to ongoing inflammation in PCD airways. Further research should focus on anti-inflammatory therapies and stimulation of efferocytosis as a strategy to treat PCD.
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