{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,21]],"date-time":"2026-04-21T08:56:31Z","timestamp":1776761791853,"version":"3.51.2"},"reference-count":36,"publisher":"Wiley","issue":"4","license":[{"start":{"date-parts":[[2026,3,30]],"date-time":"2026-03-30T00:00:00Z","timestamp":1774828800000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"},{"start":{"date-parts":[[2026,3,30]],"date-time":"2026-03-30T00:00:00Z","timestamp":1774828800000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/doi.wiley.com\/10.1002\/tdm_license_1.1"}],"content-domain":{"domain":["analyticalsciencejournals.onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["J Mass Spectrom"],"published-print":{"date-parts":[[2026,4]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                  <jats:p>\n                    Identification of hemoglobin (Hb) variants is of significant value in the clinical diagnosis of hemoglobinopathies. A common issue with the conventional methods for identifying Hb variants is their limited resolution to provide primary structures. Previous literature has shown the ability for high\u2010resolution mass spectrometry (HR\u2010MS) methods to accurately identify Hb variants, but ambiguity may still remain when distinguishing isomeric Hb variants due to inadequate AA sequence coverage. Using multiple fragmentation modes can be a solution to achieve higher sequence coverage in MS\n                    <jats:sup>2<\/jats:sup>\n                    fragment analysis. A liquid chromatography\u2010high\u2010resolution mass spectrometry (LC\u2010HR\u2010MS) method with multiple fragmentation modes was developed for identifying Hb variants, which could both effectively separate pairs of normal and variant Hb subunits and accurately distinguish isomeric Hb variant subunits. The separation was facilitated by a C4 reversed\u2010phase column, and the fragmentation modes in use were higher\u2010energy collision dissociation (HCD) and electron\u2010transfer\/higher\u2010energy collision dissociation (EThcD). The fragmentation modes HCD, ETD, and EThcD were tested with normal \u03b1 and \u03b2 subunits to evaluate the MS\n                    <jats:sup>2<\/jats:sup>\n                    performance of each mode. The combined use of EThcD and HCD was able to make high AA sequence coverage. Five clinical samples, including heterozygous Hb E and Hb Constant Spring (CS), Hb Hofu, Hb Raleigh, Hb P\u2010Nilotic, and Hb F\u2010Kuala Lumpur (KL), were analyzed to demonstrate the performance of the LC\u2010HR\u2010MS method for accurately identifying Hb variants samples. This article demonstrated the advantages of LC\u2010HR\u2010MS with multiple fragmentation modes in effectively solving clinical cases of Hb variants. In clinical practice, the combination of the LC\u2010HR\u2010MS and conventional Hb test methods is favorable, allowing for accurate diagnosis of Hb variants and significantly reducing the need for more costly genetic tests.\n                  <\/jats:p>","DOI":"10.1002\/jms.70054","type":"journal-article","created":{"date-parts":[[2026,3,31]],"date-time":"2026-03-31T05:26:13Z","timestamp":1774934773000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Establishment of a Top\u2010Down Liquid Chromatography\u2010High\u2010Resolution\u2010Mass Spectrometry Method With Multiple Fragmentation Modes for Clinical Diagnosis of Hemoglobin Variants"],"prefix":"10.1002","volume":"61","author":[{"given":"Danya","family":"Ortiz","sequence":"first","affiliation":[{"name":"Department of Pathology, School of Medicine Stanford University  Stanford California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Farida","family":"Elgebaly","sequence":"additional","affiliation":[{"name":"Department of Pathology, School of Medicine Stanford University  Stanford California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ryan\u00a0M.","family":"Aquino","sequence":"additional","affiliation":[{"name":"Clinical Laboratories Stanford Health Care  Palo Alto California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hector\u00a0A.","family":"Vidal","sequence":"additional","affiliation":[{"name":"Clinical Laboratories Stanford Health Care  Palo Alto California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Carolyn\u00a0V.","family":"Wong","sequence":"additional","affiliation":[{"name":"Clinical Laboratories Stanford Health Care  Palo Alto California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4483-8369","authenticated-orcid":false,"given":"Ruben\u00a0Y.","family":"Luo","sequence":"additional","affiliation":[{"name":"Department of Pathology, School of Medicine Stanford University  Stanford California USA"},{"name":"Clinical Laboratories Stanford Health Care  Palo Alto California USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2026,3,30]]},"reference":[{"key":"e_1_2_11_2_1","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkt911"},{"key":"e_1_2_11_3_1","doi-asserted-by":"publisher","DOI":"10.1093\/ajcp\/aqaa260"},{"key":"e_1_2_11_4_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00216-015-8525-5"},{"key":"e_1_2_11_5_1","doi-asserted-by":"publisher","DOI":"10.1373\/clinchem.2018.295766"},{"key":"e_1_2_11_6_1","doi-asserted-by":"publisher","DOI":"10.1255\/ejms.1340"},{"key":"e_1_2_11_7_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.cca.2016.07.006"},{"key":"e_1_2_11_8_1","doi-asserted-by":"publisher","DOI":"10.1515\/cclm-2019-0832"},{"key":"e_1_2_11_9_1","doi-asserted-by":"publisher","DOI":"10.1021\/jasms.1c00291"},{"key":"e_1_2_11_10_1","doi-asserted-by":"publisher","DOI":"10.1021\/acs.analchem.0c01350"},{"key":"e_1_2_11_11_1","doi-asserted-by":"publisher","DOI":"10.1093\/clinchem\/hvac171"},{"key":"e_1_2_11_12_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmsacl.2025.01.002"},{"key":"e_1_2_11_13_1","doi-asserted-by":"publisher","DOI":"10.1093\/labmed\/lmaf029"},{"key":"e_1_2_11_14_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.heliyon.2023.e19116"},{"key":"e_1_2_11_15_1","volume-title":"GeneReviews","author":"Tamary H.","year":"1993"},{"key":"e_1_2_11_16_1","doi-asserted-by":"publisher","DOI":"10.1590\/S1415-47572009005000071"},{"key":"e_1_2_11_17_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0031-3955(05)70427-2"},{"key":"e_1_2_11_18_1","first-page":"36","article-title":"Different Geographic Origins of Hb Constant Spring [alpha(2) Codon 142 TAA\u2010\u2010>CAA]","volume":"86","author":"Harteveld C. 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