{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T10:22:42Z","timestamp":1791886962281,"version":"4.3.4"},"reference-count":79,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2023,2,25]],"date-time":"2023-02-25T00:00:00Z","timestamp":1677283200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Institutes of Health\/National Institute of Allergy and Infectious Diseases","award":["R01 AI132638"],"award-info":[{"award-number":["R01 AI132638"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Cells"],"abstract":"<jats:p>Neutrophils are a vital component of the innate immune system and play an essential function in the recognition and clearance of bacterial and fungal pathogens. There is great interest in understanding mechanisms of neutrophil dysfunction in the setting of disease and deciphering potential side effects of immunomodulatory drugs on neutrophil function. We developed a high throughput flow cytometry-based assay for detecting changes to four canonical neutrophil functions following biological or chemical triggers. Our assay detects neutrophil phagocytosis, reactive oxygen species (ROS) generation, ectodomain shedding, and secondary granule release in a single reaction mixture. By selecting fluorescent markers with minimal spectral overlap, we merge four detection assays into one microtiter plate-based assay. We demonstrate the response to the fungal pathogen, Candida albicans and validate the assay\u2019s dynamic range using the inflammatory cytokines G-CSF, GM-CSF, TNF\u03b1, and IFN\u03b3. All four cytokines increased ectodomain shedding and phagocytosis to a similar degree while GM-CSF and TNF\u03b1 were more active in degranulation when compared to IFN\u03b3 and G-CSF. We further demonstrated the impact of small molecule inhibitors such as kinase inhibition downstream of Dectin-1, a critical lectin receptor responsible for fungal cell wall recognition. Bruton\u2019s tyrosine kinase (Btk), Spleen tyrosine kinase (Syk), and Src kinase inhibition suppressed all four measured neutrophil functions but all functions were restored with lipopolysaccharide co-stimulation. This new assay allows for multiple comparisons of effector functions and permits identification of distinct subpopulations of neutrophils with a spectrum of activity. Our assay also offers the potential for studying the intended and off-target effects of immunomodulatory drugs on neutrophil responses.<\/jats:p>","DOI":"10.3390\/cells12050743","type":"journal-article","created":{"date-parts":[[2023,2,27]],"date-time":"2023-02-27T04:13:16Z","timestamp":1677471196000},"page":"743","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":16,"title":["Multiparametric Profiling of Neutrophil Function via a High-Throughput Flow Cytometry-Based Assay"],"prefix":"10.3390","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4295-0007","authenticated-orcid":false,"given":"Kyle D.","family":"Timmer","sequence":"first","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3617-1947","authenticated-orcid":false,"given":"Daniel J.","family":"Floyd","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3735-3873","authenticated-orcid":false,"given":"Allison K.","family":"Scherer","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"},{"name":"Harvard Medical School, Boston, MA 20114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8572-6366","authenticated-orcid":false,"given":"Arianne J.","family":"Crossen","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0681-9087","authenticated-orcid":false,"given":"Johnny","family":"Atallah","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"},{"name":"Harvard Medical School, Boston, MA 20114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0271-7470","authenticated-orcid":false,"given":"Adam L.","family":"Viens","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David B.","family":"Sykes","sequence":"additional","affiliation":[{"name":"Harvard Medical School, Boston, MA 20114, USA"},{"name":"Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA"},{"name":"Harvard Stem Cell Institute, Cambridge, MA 02114, USA"},{"name":"Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Michael K.","family":"Mansour","sequence":"additional","affiliation":[{"name":"Department of Medicine, Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA"},{"name":"Harvard Medical School, Boston, MA 20114, USA"},{"name":"Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2023,2,25]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"441","DOI":"10.1111\/bjh.13113","article-title":"Mucosal Barrier Injury, Fever and Infection in Neutropenic Patients with Cancer: Introducing the Paradigm Febrile Mucositis","volume":"167","author":"Herbers","year":"2014","journal-title":"Br. J. Haematol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"17032","DOI":"10.1038\/nrdp.2017.32","article-title":"Severe Congenital Neutropenias","volume":"3","author":"Skokowa","year":"2017","journal-title":"Nat. Rev. Dis. Primers"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"5","DOI":"10.1007\/s15010-013-0525-9","article-title":"The Current Spectrum of Infection in Cancer Patients with Chemotherapy Related Neutropenia","volume":"42","author":"Nesher","year":"2014","journal-title":"Infection"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"513","DOI":"10.2337\/dc17-2131","article-title":"Risk of Infection in Type 1 and Type 2 Diabetes Compared With the General Population: A Matched Cohort Study","volume":"41","author":"Carey","year":"2017","journal-title":"Diabetes Care"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"4409","DOI":"10.1128\/IAI.00787-12","article-title":"Insulin Treatment Directly Restores Neutrophil Phagocytosis and Bactericidal Activity in Diabetic Mice and Thereby Improves Surgical Site Staphylococcus Aureus Infection","volume":"80","author":"Hidekazu","year":"2012","journal-title":"Infect. Immun."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"224","DOI":"10.1038\/cmi.2014.2","article-title":"The Role of Neutrophils in the Development of Liver Diseases","volume":"11","author":"Xu","year":"2014","journal-title":"Cell Mol. Immunol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1002\/hep4.1645","article-title":"Loss of Coordinated Neutrophil Responses to the Human Fungal Pathogen, Candida Albicans, in Patients with Cirrhosis","volume":"5","author":"Knooihuizen","year":"2021","journal-title":"Hepatol. Commun."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"614","DOI":"10.1016\/j.humimm.2020.05.009","article-title":"Improving the Function of Neutrophils from Chronic Granulomatous Disease Patients Using Mesenchymal Stem Cells\u2019 Exosomes","volume":"81","author":"Mahmoudi","year":"2020","journal-title":"Hum. Immunol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"101","DOI":"10.1007\/s12016-020-08800-x","article-title":"Chronic Granulomatous Disease: A Comprehensive Review","volume":"61","author":"Yu","year":"2021","journal-title":"Clin. Rev. Allerg. Immunol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"894","DOI":"10.1093\/infdis\/jiab009","article-title":"Cytokine Augmentation Reverses Transplant Recipient Neutrophil Dysfunction Against the Human Fungal Pathogen Candida Albicans","volume":"224","author":"Barros","year":"2020","journal-title":"J. Infect. Dis."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"678771","DOI":"10.3389\/fimmu.2021.678771","article-title":"A Bittersweet Response to Infection in Diabetes; Targeting Neutrophils to Modify Inflammation and Improve Host Immunity","volume":"12","author":"Dowey","year":"2021","journal-title":"Front. Immunol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1955","DOI":"10.1182\/blood-2017-11-818286","article-title":"Early-Onset Invasive Aspergillosis and Other Fungal Infections in Patients Treated with Ibrutinib","volume":"131","author":"Ghez","year":"2018","journal-title":"Blood"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"478","DOI":"10.3324\/haematol.2019.219220","article-title":"Ibrutinib Induces Multiple Functional Defects in the Neutrophil Response against Aspergillus Fumigatus","volume":"105","author":"Blez","year":"2020","journal-title":"Haematologica"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Rein\u00e9, J., Rylance, J., Ferreira, D.M., Pennington, S.H., Welters, I.D., Parker, R., and Morton, B. (2020). The Whole Blood Phagocytosis Assay: A Clinically Relevant Test of Neutrophil Function and Dysfunction in Community-Acquired Pneumonia. BMC Res. Notes, 13.","DOI":"10.1186\/s13104-020-05034-0"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"1222","DOI":"10.3389\/fimmu.2017.01222","article-title":"Functional Analysis of Phagocyte Activity in Whole Blood from HIV\/Tuberculosis-Infected Individuals Using a Novel Flow Cytometry-Based Assay","volume":"8","author":"Tembo","year":"2017","journal-title":"Front. Immunol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1186\/s40635-020-0299-1","article-title":"New Automated Analysis to Monitor Neutrophil Function Point-of-Care in the Intensive Care Unit after Trauma","volume":"8","author":"Hesselink","year":"2020","journal-title":"Intensive Care Med. Exp."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"46","DOI":"10.1016\/j.jim.2019.05.006","article-title":"A Versatile High-Throughput Assay to Characterize Antibody-Mediated Neutrophil Phagocytosis","volume":"471","author":"Karsten","year":"2019","journal-title":"J. Immunol. Methods"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"e00790-18","DOI":"10.1128\/IAI.00790-18","article-title":"Novel Assay to Characterize Neutrophil Responses to Oral Biofilms","volume":"87","author":"Morvarid","year":"2019","journal-title":"Infect. Immun."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1785","DOI":"10.1091\/mbc.e08-11-1135","article-title":"ADAMs 10 and 17 Represent Differentially Regulated Components of a General Shedding Machinery for Membrane Proteins Such as Transforming Growth Factor \u03b1, L-Selectin, and Tumor Necrosis Factor \u03b1","volume":"20","author":"Gall","year":"2009","journal-title":"Mol. Biol. Cell"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1068","DOI":"10.3389\/fimmu.2019.01068","article-title":"L-Selectin: A Major Regulator of Leukocyte Adhesion, Migration and Signaling","volume":"10","author":"Ivetic","year":"2019","journal-title":"Front. Immunol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"680","DOI":"10.1016\/j.bbamcr.2012.11.027","article-title":"ADAM17 Cleaves CD16b (Fc\u03b3RIIIb) in Human Neutrophils","volume":"1833","author":"Wang","year":"2013","journal-title":"Biochim. Et Biophys. Acta BBA\u2014Mol. Cell Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1189\/jlb.0307193","article-title":"Role of ADAM17 in the Ectodomain Shedding of TNF-\u03b1 and Its Receptors by Neutrophils and Macrophages","volume":"82","author":"Bell","year":"2007","journal-title":"J. Leukoc. Biol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2686","DOI":"10.4049\/jimmunol.179.5.2686","article-title":"Cutting Edge: TNF-\u03b1-Converting Enzyme (TACE\/ADAM17) Inactivation in Mouse Myeloid Cells Prevents Lethality from Endotoxin Shock","volume":"179","author":"Horiuchi","year":"2007","journal-title":"J. Immunol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1093\/intimm\/dxl119","article-title":"Effects of TREM-1 Activation in Human Neutrophils: Activation of Signaling Pathways, Recruitment into Lipid Rafts and Association with TLR4","volume":"19","author":"Fortin","year":"2007","journal-title":"Int. Immunol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1021","DOI":"10.1038\/mi.2016.104","article-title":"Reversal of TREM-1 Ectodomain Shedding and Improved Bacterial Clearance by Intranasal Metalloproteinase Inhibitors","volume":"10","author":"Weiss","year":"2017","journal-title":"Mucosal. Immunol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1238","DOI":"10.1126\/science.2551036","article-title":"Neutrophil Mac-1 and MEL-14 Adhesion Proteins Inversely Regulated by Chemotactic Factors","volume":"245","author":"Kishimoto","year":"1989","journal-title":"Science"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1204","DOI":"10.1111\/j.1365-2222.2005.02320.x","article-title":"L-Selectin Expression on Neutrophils from Allergic Patients","volume":"35","author":"Vega","year":"2005","journal-title":"Clin. Exp. Allergy"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"138","DOI":"10.3389\/fcimb.2017.00138","article-title":"Ectodomain Shedding by ADAM17: Its Role in Neutrophil Recruitment and the Impairment of This Process during Sepsis","volume":"7","author":"Mishra","year":"2017","journal-title":"Front. Cell Infect. Microbiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"666732","DOI":"10.3389\/fphar.2021.666732","article-title":"Phagocytosis, Degranulation and Extracellular Traps Release by Neutrophils\u2014The Current Knowledge, Pharmacological Modulation and Future Prospects","volume":"12","author":"Gierlikowska","year":"2021","journal-title":"Front. Pharmacol."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"340","DOI":"10.1016\/j.it.2007.06.002","article-title":"Neutrophil Granules: A Library of Innate Immunity Proteins","volume":"28","author":"Borregaard","year":"2007","journal-title":"Trends Immunol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"855","DOI":"10.4049\/jimmunol.124.2.855","article-title":"The Sequential Release of Granule Constitutents from Human Neutrophils","volume":"124","author":"Bentwood","year":"1980","journal-title":"J. Immunol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1535","DOI":"10.4049\/jimmunol.150.4.1535","article-title":"Control of Exocytosis in Early Neutrophil Activation","volume":"150","author":"Kjeldsen","year":"1993","journal-title":"J. Immunol."},{"key":"ref_33","doi-asserted-by":"crossref","unstructured":"Hopke, A., Nicke, N., Hidu, E.E., Degani, G., Popolo, L., and Wheeler, R.T. (2016). Neutrophil Attack Triggers Extracellular Trap-Dependent Candida Cell Wall Remodeling and Altered Immune Recognition. PLoS Pathog., 12.","DOI":"10.1371\/journal.ppat.1005644"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"794","DOI":"10.1093\/rheumatology\/40.7.794","article-title":"Inhibition of Neutrophil Responses by Cyclosporin A. An Insight into Molecular Mechanisms","volume":"40","author":"Spisani","year":"2001","journal-title":"Rheumatology"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"e334","DOI":"10.1002\/ctm2.334","article-title":"Cyclosporine Modulates Neutrophil Functions via the SIRT6\u2013HIF-1\u03b1\u2013Glycolysis Axis to Alleviate Severe Ulcerative Colitis","volume":"11","author":"Lu","year":"2021","journal-title":"Clin. Transl. Med."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"2644","DOI":"10.1128\/AAC.01618-07","article-title":"Impact of Mycophenolic Acid on the Functionality of Human Polymorphonuclear Neutrophils and Dendritic Cells during Interaction with Aspergillus Fumigatus","volume":"52","author":"Markus","year":"2008","journal-title":"Antimicrob. Agents Chemother."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1664","DOI":"10.4049\/jimmunol.2100599","article-title":"TLR Signaling Rescues Fungicidal Activity in Syk-Deficient Neutrophils","volume":"208","author":"Viens","year":"2022","journal-title":"J. Immunol."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"633","DOI":"10.1016\/0891-5849(94)00181-I","article-title":"Modification of Neutrophil Oxidant Production with Diphenyleneiodonium and Its Effect on Bacterial Killing","volume":"18","author":"Hampton","year":"1995","journal-title":"Free Radic. Bio Med."},{"key":"ref_39","first-page":"139","article-title":"Mycophenolate Mofetil: Safety and Efficacy in the Prophylaxis of Acute Kidney Transplantation Rejection","volume":"5","author":"Dalal","year":"2009","journal-title":"Ther. Clin. Risk Manag."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"205","DOI":"10.1038\/sj.bmt.1703349","article-title":"CsA-Based Post-Graft Immunosuppression: The Main Factor for Improving Outcome of Allografted Patients with Acquired Aplastic Anemia. A Retrospective Survey by the Spanish Group of Hematopoietic Transplantation","volume":"29","author":"Arranz","year":"2002","journal-title":"Bone Marrow Transpl."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"277","DOI":"10.1159\/000345992","article-title":"Exocytosis of Neutrophil Granule Subsets and Activation of Prolyl Isomerase 1 Are Required for Respiratory Burst Priming","volume":"5","author":"McLeish","year":"2013","journal-title":"J. Innate Immun."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1038\/ni1408","article-title":"Dectin-1 Is Required for \u03b2-Glucan Recognition and Control of Fungal Infection","volume":"8","author":"Taylor","year":"2007","journal-title":"Nat. Immunol."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"e02043-19","DOI":"10.1128\/mBio.02043-19","article-title":"Spleen Tyrosine Kinase Is a Critical Regulator of Neutrophil Responses to Candida Species","volume":"11","author":"Negoro","year":"2020","journal-title":"mBio"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2978","DOI":"10.4049\/jimmunol.1002268","article-title":"Class IA Phosphoinositide 3-Kinase \u03b2 and \u03b4 Regulate Neutrophil Oxidase Activation in Response to Aspergillus Fumigatus Hyphae","volume":"186","author":"Boyle","year":"2011","journal-title":"J. Immunol."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"2409","DOI":"10.1080\/10428194.2020.1753043","article-title":"The Effect of Ibrutinib on Neutrophil and \u0393\u03b4 T Cell Functions","volume":"61","author":"Risnik","year":"2020","journal-title":"Leuk. Lymphoma"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"1119","DOI":"10.1038\/s41590-020-0736-z","article-title":"Single-Cell Transcriptome Profiling Reveals Neutrophil Heterogeneity in Homeostasis and Infection","volume":"21","author":"Xie","year":"2020","journal-title":"Nat. Immunol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"792122","DOI":"10.3389\/fimmu.2021.792122","article-title":"Single-Cell Transcriptome Profiling Reveals Neutrophil Heterogeneity and Functional Multiplicity in the Early Stage of Severe Burn Patients","volume":"12","author":"Huang","year":"2022","journal-title":"Front. Immunol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"508","DOI":"10.1111\/j.1365-2141.2005.05794.x","article-title":"Circulating CD10\u2212\/CD16low Neutrophils Provide a Quantitative Index of Active Bone Marrow Neutrophil Release","volume":"131","author":"Orr","year":"2005","journal-title":"Brit. J. Haematol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1343","DOI":"10.1182\/blood-2016-04-713206","article-title":"Mature CD10+ and Immature CD10\u2212 Neutrophils Present in G-CSF\u2013Treated Donors Display Opposite Effects on T Cells","volume":"129","author":"Marini","year":"2017","journal-title":"Blood"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"668","DOI":"10.1111\/j.1462-5822.2005.00659.x","article-title":"Neutrophil Extracellular Traps Capture and Kill Candida Albicans Yeast and Hyphal Forms","volume":"8","author":"Urban","year":"2006","journal-title":"Cell. Microbiol."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1532","DOI":"10.1126\/science.1092385","article-title":"Neutrophil Extracellular Traps Kill Bacteria","volume":"303","author":"Brinkmann","year":"2004","journal-title":"Science"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"822","DOI":"10.1002\/cyto.a.23169","article-title":"Measurement of NET Formation in Vitro and in Vivo by Flow Cytometry","volume":"91","author":"Masuda","year":"2017","journal-title":"Cytom. Part A"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"1155","DOI":"10.1002\/ajh.24185","article-title":"Flow Cytometric Assay for Direct Quantification of Neutrophil Extracellular Traps in Blood Samples","volume":"90","author":"Gavillet","year":"2015","journal-title":"Am. J. Hematol."},{"key":"ref_54","doi-asserted-by":"crossref","unstructured":"Saffarzadeh, M., Juenemann, C., Queisser, M.A., Lochnit, G., Barreto, G., Galuska, S.P., Lohmeyer, J., and Preissner, K.T. (2012). Neutrophil Extracellular Traps Directly Induce Epithelial and Endothelial Cell Death: A Predominant Role of Histones. PLoS ONE, 7.","DOI":"10.1371\/journal.pone.0032366"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"1322","DOI":"10.1038\/s41467-019-09160-7","article-title":"Neutrophil Activation and NETosis Are the Major Drivers of Thrombosis in Heparin-Induced Thrombocytopenia","volume":"10","author":"Perdomo","year":"2019","journal-title":"Nat. Commun."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"15880","DOI":"10.1073\/pnas.1005743107","article-title":"Extracellular DNA Traps Promote Thrombosis","volume":"107","author":"Fuchs","year":"2010","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_57","doi-asserted-by":"crossref","first-page":"1105","DOI":"10.1182\/blood.V78.4.1105.1105","article-title":"Membrane Surface Antigen Expression on Neutrophils: A Reappraisal of the Use of Surface Markers for Neutrophil Activation","volume":"78","author":"Kuijpers","year":"1991","journal-title":"Blood"},{"key":"ref_58","doi-asserted-by":"crossref","first-page":"2831","DOI":"10.4049\/jimmunol.177.5.2831","article-title":"Combinatorial SNARE Complexes Modulate the Secretion of Cytoplasmic Granules in Human Neutrophils","volume":"177","author":"Mollinedo","year":"2006","journal-title":"J. Immunol."},{"key":"ref_59","doi-asserted-by":"crossref","unstructured":"Borysowski, J., Mi\u0119dzybrodzki, R., Wierzbicki, P., K\u0142osowska, D., Korczak-Kowalska, G., Weber-D\u0105browska, B., and G\u00f3rski, A. (2017). A3R Phage and Staphylococcus Aureus Lysate Do Not Induce Neutrophil Degranulation. Viruses, 9.","DOI":"10.3390\/v9020036"},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"e02759-19","DOI":"10.1128\/mBio.02759-19","article-title":"Inhibition of Neutrophil Primary Granule Release during Yersinia Pestis Pulmonary Infection","volume":"10","author":"Eichelberger","year":"2019","journal-title":"mBio"},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"293","DOI":"10.2119\/molmed.2010.00138","article-title":"Contribution of Neutrophils to Acute Lung Injury","volume":"17","author":"Grommes","year":"2011","journal-title":"Mol. Med."},{"key":"ref_62","doi-asserted-by":"crossref","unstructured":"Miralda, I., Klaes, C.K., Graham, J.E., and Uriarte, S.M. (2020). Human Neutrophil Granule Exocytosis in Response to Mycobacterium Smegmatis. Pathogens, 9.","DOI":"10.3390\/pathogens9020123"},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.cyto.2014.02.004","article-title":"Human Neutrophils Produce IL-12, IL-10, PGE2 and LTB4 in Response to Paracoccidioides Brasiliensis. Involvement of TLR2, Mannose Receptor and Dectin-1","volume":"67","author":"Balderramas","year":"2014","journal-title":"Cytokine"},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"508","DOI":"10.3389\/fimmu.2014.00508","article-title":"Neutrophil-Derived Cytokines: Facts Beyond Expression","volume":"5","author":"Tecchio","year":"2014","journal-title":"Front. Immunol."},{"key":"ref_65","doi-asserted-by":"crossref","unstructured":"Barbier, M., and Damron, F.H. (2016). Rainbow Vectors for Broad-Range Bacterial Fluorescence Labeling. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0146827"},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"635825","DOI":"10.3389\/fimmu.2021.635825","article-title":"Use of Flow Cytometry to Evaluate Phagocytosis of Staphylococcus Aureus by Human Neutrophils","volume":"12","author":"Boero","year":"2021","journal-title":"Front. Immunol."},{"key":"ref_67","first-page":"9","article-title":"Measurement of Phagocytosis and of the Phagosomal Environment in Polymorphonuclear Phagocytes by Flow Cytometry","volume":"51","author":"Simons","year":"2010","journal-title":"Curr. Protoc. Cytom."},{"key":"ref_68","doi-asserted-by":"crossref","first-page":"220","DOI":"10.1039\/C2IB00089J","article-title":"Use of Fungal Derived Polysaccharide -Conjugated Particles to Probe Dectin-1 Responses in Innate Immunity","volume":"4","author":"Tam","year":"2011","journal-title":"Integr. Biol."},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"448","DOI":"10.1097\/MEG.0000000000000801","article-title":"Granulocyte Colony-Stimulating Factor Improves Survival of Patients with Decompensated Cirrhosis: A Randomized-Controlled Trial","volume":"29","author":"Prajapati","year":"2016","journal-title":"Eur. J. Gastroen. Hepat."},{"key":"ref_70","doi-asserted-by":"crossref","first-page":"986","DOI":"10.1016\/j.jaip.2020.08.024","article-title":"Neutrophil Functional Profiling and Cytokine Augmentation for Patients with Multiple Recurrent Infections: A Case Study","volume":"9","author":"Alexander","year":"2021","journal-title":"J. Allergy Clin. Immunol. Pract."},{"key":"ref_71","doi-asserted-by":"crossref","first-page":"12937","DOI":"10.1038\/s41598-019-49419-z","article-title":"G-CSF and GM-CSF Modify Neutrophil Functions at Concentrations Found in Cystic Fibrosis","volume":"9","author":"Castellani","year":"2019","journal-title":"Sci. Rep."},{"key":"ref_72","first-page":"983698","article-title":"Neutrophils in Cancer: Two Sides of the Same Coin","volume":"2015","author":"Rosales","year":"2015","journal-title":"J. Immunol. Res."},{"key":"ref_73","doi-asserted-by":"crossref","first-page":"86","DOI":"10.1007\/s12281-019-00343-9","article-title":"Fungal Infections with Ibrutinib and Other Small-Molecule Kinase Inhibitors","volume":"13","author":"Zarakas","year":"2019","journal-title":"Curr Fungal Infect Reports"},{"key":"ref_74","doi-asserted-by":"crossref","first-page":"16","DOI":"10.1016\/0049-0172(92)90009-3","article-title":"Mechanisms of Action of Cyclosporine and Effects on Connective Tissues","volume":"21","author":"Russell","year":"1992","journal-title":"Semin. Arthritis Rheu."},{"key":"ref_75","doi-asserted-by":"crossref","first-page":"2","DOI":"10.1191\/0961203305LU2109OA","article-title":"Mechanisms of Action of Mycophenolate Mofetil","volume":"14","author":"Allison","year":"2005","journal-title":"Lupus"},{"key":"ref_76","doi-asserted-by":"crossref","first-page":"657","DOI":"10.1189\/jlb.1012544","article-title":"ROS Production in Phagocytes: Why, When, and Where?","volume":"94","author":"Erard","year":"2013","journal-title":"J. Leukoc. Biol."},{"key":"ref_77","doi-asserted-by":"crossref","first-page":"373","DOI":"10.3389\/fcimb.2017.00373","article-title":"Neutrophils to the ROScue: Mechanisms of NADPH Oxidase Activation and Bacterial Resistance","volume":"7","author":"Nguyen","year":"2017","journal-title":"Front. Cell Infect. Microbiol."},{"key":"ref_78","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1189\/jlb.1210701","article-title":"Stable Accumulation of P67phox at the Phagosomal Membrane and ROS Production within the Phagosome","volume":"91","author":"Tlili","year":"2012","journal-title":"J. Leukoc. Biol."},{"key":"ref_79","doi-asserted-by":"crossref","first-page":"945749","DOI":"10.3389\/fcell.2022.945749","article-title":"Regulation of Neutrophil NADPH Oxidase, NOX2: A Crucial Effector in Neutrophil Phenotype and Function","volume":"10","author":"Paclet","year":"2022","journal-title":"Front. Cell Dev. 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