{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,11]],"date-time":"2026-03-11T04:26:33Z","timestamp":1773203193070,"version":"3.50.1"},"reference-count":21,"publisher":"Springer Science and Business Media LLC","issue":"1","license":[{"start":{"date-parts":[[2021,7,5]],"date-time":"2021-07-05T00:00:00Z","timestamp":1625443200000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2021,7,5]],"date-time":"2021-07-05T00:00:00Z","timestamp":1625443200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Allergy Asthma Clin Immunol"],"published-print":{"date-parts":[[2021,12]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:sec>\n                    <jats:title>Rationale<\/jats:title>\n                    <jats:p>On Wright-stained sputum cytospins, eosinophil differential of\u2009\u2265\u20091.2% is considered abnormal, and\u2009\u2265\u20092.3% identifies an eosinophilic endotype. We hypothesized that failure to consider free eosinophil granules (FEG), and the re-emergence (unmasking) of eosinophilia due to various reasons underestimate the prevalence of the eosinophilic endotype.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>This is a retrospective analysis of our Institutional Review Board-approved clinical sputum database. Of the 24,176 examinations of sputa from patients with various airway diseases, 17,693 were viable cell counts from 9570 patients (6604 on a single occasion, 2967 from multiple occasions). The prevalence of intact eosinophil % at 1.2 and 2.3% thresholds was first examined. Then, additional evidence of eosinophilia was assessed by semi-quantitative enumeration of FEGs. In those patients whose sputa were examined on multiple occasions (at the time of an exacerbation or after corticosteroid dose was reduced), re-emergence (unmasking) of eosinophilia was assessed<\/jats:p>\n                    <jats:p>.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>Using the threshold of eosinophilia\u2009\u2265\u20091.2%, 6289\/17693 (35.6%) of sputa were classified as eosinophilic. This increased to 7850\/17693 (44.4%) when the presence of FEGs was considered. Using the threshold of eosinophilia\u2009\u2265\u20092.3%, 4647\/17693 (26.3%) of sputa were classified as eosinophilic. This increased to 5435\/17693 (30.7%) when the presence of FEG were considered. Extrapolating from the prevalence of re-emergence observed in the 2967 patients who had sputa examined on multiple occasions to the whole sample, we estimated that eosinophilia at 1.2% threshold would be observed in at least 60% of the samples, and a clinically relevant eosinophilia at 2.3% threshold would be observed in at least 48.5% of the samples.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Conclusions<\/jats:title>\n                    <jats:p>Using a large sputum cytometry clinical database (17,693 viable cell counts), we demonstrate that a single\u00a0time point intact cell count underestimates the prevalence of eosinophilia in a variety of airway diseases. The prevalence of eosinophilia increases from 35.6 to 60% (40% underestimation) at the 1.2% threshold, and from 26.3 to 48.5% (45% underestimation) at the 2.3% clinically relevant threshold, when free granules and a second examination are considered. This has important implications to identify the eosinophilic and Th2 high endotype both for clinical trials of anti-eosinophil therapies, and to select patients who may respond well to glucocorticosteroids and anti-IL5 therapies.<\/jats:p>\n                  <\/jats:sec>","DOI":"10.1186\/s13223-021-00567-w","type":"journal-article","created":{"date-parts":[[2021,7,5]],"date-time":"2021-07-05T06:04:33Z","timestamp":1625465073000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":19,"title":["Underestimation of airway luminal eosinophilia by quantitative sputum cytometry"],"prefix":"10.1186","volume":"17","author":[{"given":"Melanie","family":"Kjarsgaard","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Adil","family":"Adatia","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Anurag","family":"Bhalla","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Nicola","family":"LaVigne","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Katherine","family":"Radford","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chynna","family":"Huang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Manali","family":"Mukherjee","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1041-9492","authenticated-orcid":false,"given":"Parameswaran","family":"Nair","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2021,7,5]]},"reference":[{"key":"567_CR1","doi-asserted-by":"publisher","first-page":"144","DOI":"10.1155\/2011\/430317","volume":"18","author":"L D'Silva","year":"2011","unstructured":"D\u2019Silva L, Hassan N, Wang HY, Kjarsgaard M, Efthimiadis A, Hargreave FE, Nair P. Heterogeneity of bronchitis in airway diseases in tertiary care clinical practice. Can Respir J. 2011;18:144\u20138.","journal-title":"Can Respir J"},{"key":"567_CR2","doi-asserted-by":"publisher","first-page":"475","DOI":"10.1164\/ajrccm.161.2.9903097","volume":"161","author":"J Belda","year":"2000","unstructured":"Belda J, Leigh R, Parameswaran K, O\u2019Byrne PM, Sears MR, Hargreave FE. Induced sputum cell counts in healthy adults. Am J Respir Crit Care Med. 2000;161:475\u20138.","journal-title":"Am J Respir Crit Care Med"},{"key":"567_CR3","doi-asserted-by":"publisher","first-page":"81","DOI":"10.1016\/j.jaci.2013.05.007","volume":"132","author":"P Nair","year":"2013","unstructured":"Nair P. What is an \u201ceosinophilic phenotype\u201d of asthma? J Allergy Clin Immunol. 2013;132:81\u20133.","journal-title":"J Allergy Clin Immunol"},{"key":"567_CR4","doi-asserted-by":"publisher","first-page":"341","DOI":"10.1136\/thorax.56.5.341","volume":"56","author":"JS Erjefalt","year":"2001","unstructured":"Erjefalt JS, Greiff L, Andersson M, Adelroth E, Jeffery PK, Persson CG. Degranulation patterns of eosinophil granulocytes as determinants of eosinophil driven disease. Thorax. 2001;56:341\u20134.","journal-title":"Thorax"},{"key":"567_CR5","doi-asserted-by":"crossref","first-page":"1177","DOI":"10.1111\/all.12206","volume":"68","author":"P Nair","year":"2013","unstructured":"Nair P, Ochkur SI, Protheroe C, Radford K, Efthimiadis A, Lee NA, Lee JJ. Eosinophil peroxidase in sputum represents a unique biomarker of airway eosinophilia. Allergy. 2013;68:1177\u201384.","journal-title":"Allergy"},{"key":"567_CR6","doi-asserted-by":"publisher","first-page":"281","DOI":"10.1155\/2007\/190618","volume":"14","author":"L D'Silva","year":"2007","unstructured":"D\u2019Silva L, Allen CJ, Hargreave FE, Parameswaran K. Sputum neutrophilia can mask eosinophilic bronchitis during exacerbations. Can Respir J. 2007;14:281\u20134.","journal-title":"Can Respir J"},{"key":"567_CR7","doi-asserted-by":"publisher","first-page":"2217","DOI":"10.1016\/j.rmed.2007.05.010","volume":"101","author":"L D'Silva","year":"2007","unstructured":"D\u2019Silva L, Cook RJ, Allen CJ, Hargreave FE, Parameswaran K. Changing pattern of sputum cell counts during successive exacerbations of airway disease. Respir Med. 2007;101:2217\u201320.","journal-title":"Respir Med"},{"key":"567_CR8","doi-asserted-by":"publisher","first-page":"384","DOI":"10.1136\/thx.2009.126722","volume":"65","author":"DC Cowan","year":"2010","unstructured":"Cowan DC, Cowan JO, Palmay R, Williamson A, Taylor DR. Effects of steroid therapy on inflammatory cell subtypes in asthma. Thorax. 2010;65:384\u201390.","journal-title":"Thorax"},{"key":"567_CR9","doi-asserted-by":"publisher","first-page":"1174","DOI":"10.1183\/09031936.96.09061174","volume":"9","author":"E Pizzichini","year":"1996","unstructured":"Pizzichini E, Pizzichini MM, Efthimiadis A, Hargreave FE, Dolovich J. Measurement of inflammatory indices in induced sputum: effects of selection of sputum to minimize salivary contamination. Eur Respir J. 1996;9:1174\u201380.","journal-title":"Eur Respir J"},{"key":"567_CR10","doi-asserted-by":"publisher","first-page":"628","DOI":"10.1164\/rccm.201311-2069OE","volume":"189","author":"C Persson","year":"2014","unstructured":"Persson C, Uller L. Theirs but to die and do: primary lysis of eosinophils and free eosinophil granules in asthma. Am J Respir Crit Care Med. 2014;189:628\u201333.","journal-title":"Am J Respir Crit Care Med"},{"key":"567_CR11","doi-asserted-by":"publisher","first-page":"17","DOI":"10.1016\/j.jim.2013.11.013","volume":"403","author":"H Jin","year":"2014","unstructured":"Jin H, Hallstrand TS, Daly DS, Matzke MM, Nair P, Bigelow DJ, Pounds JG, Zangar RC. A halotyrosine antibody that detects increased protein modifications in asthma patients. J Immunol Methods. 2014;403:17\u201325.","journal-title":"J Immunol Methods"},{"key":"567_CR12","doi-asserted-by":"publisher","first-page":"1269","DOI":"10.1016\/j.jaci.2017.06.033","volume":"141","author":"M Mukherjee","year":"2018","unstructured":"Mukherjee M, Bulir DC, Radford K, Kjarsgaard M, Huang CM, Jacobsen EA, Ochkur SI, Catuneanu A, Lamothe-Kipnes H, Mahony J, Lee JJ, Lacy P, Nair PK. Sputum autoantibodies in patients with severe eosinophilic asthma. J Allergy Clin Immunol. 2018;141:1269\u201379.","journal-title":"J Allergy Clin Immunol"},{"key":"567_CR13","doi-asserted-by":"crossref","unstructured":"Mukherjee M, Forero DF, Tran S, Boulay ME, Bertrand M, Bhalla A, Cherukat J, Al-Hayyan H, Ayoub A, Revill SD, Javkar T, Radford K, Kjarsgaard M, Huang C, Dvorkin-Gheva A, Ask K, Olivenstein R, Dendukuri N, Lemiere C, Boulet LP, Martin JG, Nair P. Suboptimal treatment response to anti-IL-5 monoclonal antibodies in severe eosinophilic asthmatics with airway autoimmune phenomena. Eur Respir J 2020; 56.","DOI":"10.1183\/13993003.00117-2020"},{"key":"567_CR14","doi-asserted-by":"publisher","first-page":"2763","DOI":"10.3389\/fimmu.2018.02763","volume":"9","author":"M Mukherjee","year":"2018","unstructured":"Mukherjee M, Lacy P, Ueki S. Eosinophil Extracellular Traps and Inflammatory Pathologies-Untangling the Web! Front Immunol. 2018;9:2763.","journal-title":"Front Immunol"},{"key":"567_CR15","doi-asserted-by":"publisher","first-page":"35","DOI":"10.1007\/s11882-019-0868-0","volume":"19","author":"S Ueki","year":"2019","unstructured":"Ueki S, Miyabe Y, Yamamoto Y, Fukuchi M, Hirokawa M, Spencer LA, Weller PF. Charcot-Leyden Crystals in Eosinophilic Inflammation: Active Cytolysis Leads to Crystal Formation. Curr Allergy Asthma Rep. 2019;19:35.","journal-title":"Curr Allergy Asthma Rep"},{"key":"567_CR16","doi-asserted-by":"crossref","unstructured":"Persson EK, Verstraete K, Heyndrickx I, Gevaert E, Aegerter H, Percier JM, Deswarte K, Verschueren KHG, Dansercoer A, Gras D, Chanez P, Bachert C, Goncalves A, Van Gorp H, De Haard H, Blanchetot C, Saunders M, Hammad H, Savvides SN, Lambrecht BN. Protein crystallization promotes type 2 immunity and is reversible by antibody treatment. Science 2019; 364.","DOI":"10.1126\/science.aaw4295"},{"key":"567_CR17","doi-asserted-by":"crossref","unstructured":"Kulkarni NS, Hollins F, Sutcliffe A, Saunders R, Shah S, Siddiqui S, Gupta S, Haldar P, Green R, Pavord I, Wardlaw A, Brightling CE. Eosinophil protein in airway macrophages: a novel biomarker of eosinophilic inflammation in patients with asthma. J Allergy Clin Immunol 2010; 126: 61\u201369 e63.","DOI":"10.1016\/j.jaci.2010.03.026"},{"key":"567_CR18","doi-asserted-by":"crossref","unstructured":"Ghebre MA, Pang PH, Diver S, Desai D, Bafadhel M, Haldar K, Kebadze T, Cohen S, Newbold P, Rapley L, Woods J, Rugman P, Pavord ID, Johnston SL, Barer M, May RD, Brightling CE. Biological exacerbation clusters demonstrate asthma and chronic obstructive pulmonary disease overlap with distinct mediator and microbiome profiles. J Allergy Clin Immunol 2018; 141: 2027\u20132036 e2012.","DOI":"10.1016\/j.jaci.2018.04.013"},{"key":"567_CR19","doi-asserted-by":"crossref","unstructured":"Wang Z, Locantore N, Haldar K, Ramsheh MY, Beech AS, Ma W, Brown JR, Tal-Singer R, Barer MR, Bafadhel M, Donaldson GC, Wedzicha JA, Singh D, Wilkinson TMA, Miller BE, Brightling CE. Inflammatory Endotype Associated Airway Microbiome in COPD Clinical Stability and Exacerbations - A Multi-Cohort Longitudinal Analysis. Am J Respir Crit Care Med 2020.","DOI":"10.1164\/rccm.202009-3448OC"},{"key":"567_CR20","doi-asserted-by":"publisher","first-page":"1555","DOI":"10.1002\/JLB.5MA0720-770RR","volume":"108","author":"E Stewart","year":"2020","unstructured":"Stewart E, Wang X, Chupp GL, Montgomery RR. Profiling cellular heterogeneity in asthma with single cell multiparameter CyTOF. J Leukoc Biol. 2020;108:1555\u201364.","journal-title":"J Leukoc Biol"},{"key":"567_CR21","doi-asserted-by":"publisher","first-page":"696","DOI":"10.1111\/cea.13609","volume":"50","author":"M Fricker","year":"2020","unstructured":"Fricker M, Qin L, Niessen N, Baines KJ, McDonald VM, Scott HA, Simpson JL, Gibson PG. Relationship of sputum mast cells with clinical and inflammatory characteristics of asthma. Clin Exp Allergy. 2020;50:696\u2013707.","journal-title":"Clin Exp Allergy"}],"container-title":["Allergy, Asthma &amp; Clinical Immunology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s13223-021-00567-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1186\/s13223-021-00567-w\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1186\/s13223-021-00567-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,11,5]],"date-time":"2023-11-05T12:16:02Z","timestamp":1699186562000},"score":1,"resource":{"primary":{"URL":"https:\/\/aacijournal.biomedcentral.com\/articles\/10.1186\/s13223-021-00567-w"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,7,5]]},"references-count":21,"journal-issue":{"issue":"1","published-print":{"date-parts":[[2021,12]]}},"alternative-id":["567"],"URL":"https:\/\/doi.org\/10.1186\/s13223-021-00567-w","relation":{"has-preprint":[{"id-type":"doi","id":"10.21203\/rs.3.rs-297911\/v1","asserted-by":"object"}]},"ISSN":["1710-1492"],"issn-type":[{"value":"1710-1492","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,7,5]]},"assertion":[{"value":"3 March 2021","order":1,"name":"received","label":"Received","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"24 June 2021","order":2,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"5 July 2021","order":3,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"This study was approved by the Research Ethics Board of St. Joseph\u2019s Healthcare.Hamilton (RP#11020), for the collection and maintenance of a sputum cell count database for research.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Ethics approval and consent to participate"}},{"value":"Not applicable.","order":3,"name":"Ethics","group":{"name":"EthicsHeading","label":"Consent for publication"}},{"value":"The other authors declare no competing interests or conflicts of interest. PN reports grants and personal fees from AstraZeneca, grants from Novartis, grants and personal fees from Teva, grants from Sanofi, grants and personal fees from Roche, personal fees from Novartis, personal fees from Merck and personal fees from Equillium, outside the submitted work. MM reports a grant from Methapharm Specialty Pharmaceuticals and personal fees from AstraZeneca and GlaxoSmithKline, outside the submitted work.","order":4,"name":"Ethics","group":{"name":"EthicsHeading","label":"Competing interests"}}],"article-number":"63"}}