Computational detection and quantification of human and mouse neutrophil extracellular traps in flow cytometry and confocal microscopy

Ginley, Brandon; Emmons, Tiffany R.; Lutnick, Brendon; Urban, Constantin F.; Segal, Brahm H.; Sarder, Pinaki

doi:10.1038/s41598-017-18099-y

Cited by 25 publications

(31 citation statements)

References 26 publications

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“…In contrast to this study, two previous studies using imaging flow cytometry failed to identify NETs in solution, although they were able to identify potential precursors to NETs, that is, cells with decondensed nuclei and morphological changes thought to be related NET formation . An additional study used imaging flow cytometry to identify what look to be DNA strands attached to whole cells . Unfortunately, due to the lack of a brightfield image and NET protein markers, it is difficult to confirm if these are truly NETs or some kind of artifact produced during cell preparation.…”

Section: Discussioncontrasting

confidence: 61%

Rapid Quantification of NETs In Vitro and in Whole Blood Samples by Imaging Flow Cytometry

et al. 2019

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Neutrophil extracellular trap (NET) formation involves the release of DNA outside the cell to neutralize pathogens. Techniques such as live microscopy, flow cytometry, and intravital imaging allow the characterization of NETs, but these either cannot be applied in vivo, lack specificity or require invasive procedures. We developed an automated analysis method to rapidly acquire and characterize cells as NETs or NET precursors, as opposed to cells undergoing other forms of cell death, using imaging flow cytometry. NETs were maintained in solution using a novel three‐dimensional cell culture system in which cells are suspended at the interface of two liquids of different density. Critically, we identify NETs using an image analysis algorithm based on morphological data showing the extrusion of DNA beyond the cell boundaries. In vitro, we used this technique to demonstrate different requirements for NET formation in human and mouse neutrophils. We also measured NETs in whole blood during infection of mice with the malaria parasite Plasmodium yoelii. We expect this technique will provide a valuable approach to better understand the process of NET formation and its importance in disease. © 2019 International Society for Advancement of Cytometry

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Section: Discussioncontrasting

confidence: 61%

Rapid Quantification of NETs In Vitro and in Whole Blood Samples by Imaging Flow Cytometry

et al. 2019

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“…Immunocytochemistry and immunohistochemistry are among the most common methods used to detect NETs. 28 Several methods permit NET detection real time in vivo, including live cell imaging, [29][30][31][32] intravital imaging, 10,33 and the use of DNA-intercalating dyes. 34,35 NETs can also be detected ex vivo in fluid secretions, serum, and tissue sections.…”

Section: Neutrophil Extracellular Trap Detectionmentioning

confidence: 99%

Neutrophil Extracellular Traps and Liver Disease

Hilscher

Shah

2019

Semin Liver Dis

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Neutrophil extracellular traps, or NETs, are heterogenous, filamentous structures which consist of extracellular DNA, granular proteins, and histones. NETs are extruded by a neutrophil in response to various stimuli. Although NETs were initially implicated in immune defense, subsequent studies have implicated NETs in a spectrum of disease processes, including autoimmune disease, thrombosis, and cancer. NETs also contribute to the pathogenesis of several common liver diseases, including alcohol-associated liver disease and portal hypertension. Although there is much interest in the therapeutic potential of NET inhibition, future clinical applications must be balanced against potential increased risk of infection.

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“…2). The use of different methods of detection and quantification of NETs in vitro, in serum and in tissue [48][49][50][51][52][53][54][55][56] also impedes interpretation and/or comparison.…”

Section: Introductionmentioning

confidence: 99%

To NET or not to NET:current opinions and state of the science regarding the formation of neutrophil extracellular traps

et al. 2019

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Since the discovery and definition of neutrophil extracellular traps (NETs) 14 years ago, numerous characteristics and physiological functions of NETs have been uncovered. Nowadays, the field continues to expand and novel mechanisms that orchestrate formation of NETs, their previously unknown properties, and novel implications in disease continue to emerge. The abundance of available data has also led to some confusion in the NET research community due to contradictory results and divergent scientific concepts, such as pro-and anti-inflammatory roles in pathologic conditions, demarcation from other forms of cell death, or the origin of the DNA that forms the NET scaffold. Here, we present prevailing concepts and state of the science in NET-related research and elaborate on open questions and areas of dispute.

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Computational detection and quantification of human and mouse neutrophil extracellular traps in flow cytometry and confocal microscopy

Cited by 25 publications

References 26 publications

Rapid Quantification of NETs In Vitro and in Whole Blood Samples by Imaging Flow Cytometry

Rapid Quantification of NETs In Vitro and in Whole Blood Samples by Imaging Flow Cytometry

Neutrophil Extracellular Traps and Liver Disease

To NET or not to NET:current opinions and state of the science regarding the formation of neutrophil extracellular traps

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