{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,3]],"date-time":"2026-09-03T12:36:08Z","timestamp":1788438968499,"version":"build-2803163510"},"reference-count":24,"publisher":"MDPI AG","issue":"2","license":[{"start":{"date-parts":[[2026,3,5]],"date-time":"2026-03-05T00:00:00Z","timestamp":1772668800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Antibodies"],"abstract":"<jats:p>Background: Anti-double-stranded DNA (anti-dsDNA) antibodies are a key serological marker for systemic lupus erythematosus (SLE) and are commonly assessed in conjunction with anti-nuclear antibody (ANA) testing by indirect immunofluorescence (IIF) on HEp-2 cells. However, their detection is influenced both by the heterogeneity of the autoimmune response and by the characteristics of the analytical method employed, thereby complicating diagnostic interpretation. Methods: In this retrospective single-center study, 3090 consecutive patients undergoing anti-dsDNA analysis were screened, and 138 positive individuals, with anti-dsDNA levels \u2265 15 IU\/mL by fluoroenzyme immunoassay (FEIA), were included in the study. A control group of 29 anti-dsDNA-negative patients was also analyzed. Anti-dsDNA-positive patients were stratified by antibody level (low, mild, high), and the results were correlated with HEp-2 IIF titers and fluorescence patterns. Furthermore, in a subset of 30 positive patients, anti-dsDNA antibodies were evaluated using immunoblotting (IB) and the Crithidia luciliae indirect immunofluorescence test (CLIFT). Statistical analyses assessed associations and concordance among methods. Results: Higher anti-dsDNA levels were generally associated with higher HEp-2 IIF titers. However, a considerable percentage (35%) of patients with positive anti-dsDNA were negative by HEp-2 IIF. Notably, high anti-dsDNA levels were detected in 19% of HEp-2 IIF-negative patients (titer &lt; 1:80), 18% of mildly HEp-2 IIF-positive patients (titer 1:80\u20131:160), and 25% of HEp-2 IIF-positive patients (titer &gt; 1:320). In the subset of 30 positive patients, FEIA analysis showed high concordance with the immunoblot in both IIF-positive (81%) and -negative (100%) patients, while CLIFT demonstrated lower agreement with both FEIA and IB independently of the IIF. Conclusions: Our findings indicate that anti-dsDNA antibody detection may occur independently of HEp-2 IIF positivity and that FEIA, especially when confirmed by immunoblot, represents a reliable approach for anti-dsDNA assessment. The observed results in this study likely reflect differences in epitope recognition and assay sensitivity among methods, suggesting the use of a multi-step diagnostic strategy in the serological evaluation of SLE.<\/jats:p>","DOI":"10.3390\/antib15020023","type":"journal-article","created":{"date-parts":[[2026,3,5]],"date-time":"2026-03-05T10:31:14Z","timestamp":1772706674000},"page":"23","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Evaluation of Anti-dsDNA Antibodies in Laboratory Practice: Management of Different Analytical Methods and Correlation with HEp-2 Immunofluorescence Patterns"],"prefix":"10.3390","volume":"15","author":[{"given":"Massimo","family":"Papale","sequence":"first","affiliation":[{"name":"Laboratory of Clinical Pathology, Department of Laboratory Diagnostics, Polyclinic University-Hospital Foggia, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2677-5747","authenticated-orcid":false,"given":"Carmela","family":"Paolillo","sequence":"additional","affiliation":[{"name":"Clinical Biochemistry, Department of Clinical and Experimental Medicine, University of Foggia, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Loredana","family":"Iafelice","sequence":"additional","affiliation":[{"name":"Laboratory of Clinical Pathology, Department of Laboratory Diagnostics, Polyclinic University-Hospital Foggia, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tiziana","family":"Trivisano","sequence":"additional","affiliation":[{"name":"Laboratory of Clinical Pathology, Department of Laboratory Diagnostics, Polyclinic University-Hospital Foggia, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3495-2707","authenticated-orcid":false,"given":"Giuseppe Stefano","family":"Netti","sequence":"additional","affiliation":[{"name":"Unit of Clinical Pathology, Center for Research and Innovation in Medicine (CREATE), Department of Medical and Surgical Sciences, University of Foggia, Viale Luigi Pinto, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4996-3938","authenticated-orcid":false,"given":"Elena","family":"Ranieri","sequence":"additional","affiliation":[{"name":"Unit of Clinical Pathology, Center for Research and Innovation in Medicine (CREATE), Department of Medical and Surgical Sciences, University of Foggia, Viale Luigi Pinto, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4720-1320","authenticated-orcid":false,"given":"Gaetano","family":"Corso","sequence":"additional","affiliation":[{"name":"Laboratory of Clinical Pathology, Department of Laboratory Diagnostics, Polyclinic University-Hospital Foggia, 71122 Foggia, Italy"},{"name":"Clinical Biochemistry, Department of Clinical and Experimental Medicine, University of Foggia, 71122 Foggia, Italy"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,3,5]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1151","DOI":"10.1136\/annrheumdis-2018-214819","article-title":"2019 European League Against Rheumatism\/American College of Rheumatology classification criteria for systemic lupus erythematosus","volume":"78","author":"Aringer","year":"2019","journal-title":"Ann. Rheum. Dis."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"2677","DOI":"10.1002\/art.34473","article-title":"Derivation and validation of the Systemic Lupus International Collaborating Clinics classification criteria for systemic lupus erythematosus","volume":"64","author":"Petri","year":"2012","journal-title":"Arthritis Rheum."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1526","DOI":"10.1056\/NEJMoa021933","article-title":"Development of autoantibodies before the clinical onset of systemic lupus erythematosus","volume":"349","author":"Arbuckle","year":"2003","journal-title":"N. Engl. J. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1619","DOI":"10.1007\/s10067-013-2330-3","article-title":"Immunoserological parameters in SLE: High-avidity anti-dsDNA detected by ELISA are the most closely associated with the disease activity","volume":"32","author":"Andrejevic","year":"2013","journal-title":"Clin. Rheumatol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"56","DOI":"10.1007\/BF02200721","article-title":"Clinical significance of antibodies to double stranded DNA (dsDNA) for systemic lupus erythematosus (SLE)","volume":"6","author":"Swaak","year":"1987","journal-title":"Clin. Rheumatol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"98","DOI":"10.1038\/s41584-020-00544-4","article-title":"Toll-like receptor signalling in B cells during systemic lupus erythematosus","volume":"17","author":"Fillatreau","year":"2021","journal-title":"Nat. Rev. Rheumatol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"311","DOI":"10.3109\/03009742.2013.765032","article-title":"Low diagnostic and predictive value of anti-dsDNA antibodies in unselected patients with recent onset of rheumatic symptoms: Results from a long-term follow-up Scandinavian multicentre study","volume":"42","author":"Compagno","year":"2013","journal-title":"Scand. J. Rheumatol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1136\/annrheumdis-2013-203863","article-title":"International recommendations for the assessment of autoantibodies to cellular antigens referred to as anti-nuclear antibodies","volume":"73","author":"Damoiseaux","year":"2014","journal-title":"Ann. Rheum. Dis."},{"key":"ref_9","first-page":"50","article-title":"Forum Interdisciplinare per la Ricerca sulle Malattie Autoimmuni (FIRMA) studygroup. Diagnostic accuracy of currently available anti-double-stranded DNA antibody assays. An Italian multicentre study","volume":"29","author":"Ghirardello","year":"2011","journal-title":"Clin. Exp. Rheumatol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"e13123","DOI":"10.1111\/sji.13123","article-title":"Comparative analysis of three methods in anti-dsDNA antibodies detection: Implications for Systemic Lupus Erythematosus diagnosis","volume":"95","author":"Cuomo","year":"2022","journal-title":"Scand. J. Immunol."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Lu, R., Yu, R., Huang, R., Xue, C., Song, N., Zhao, J., Zeng, X., and Hu, C. (2025). Comparative evaluation of three anti-dsDNA antibody detection methods in systemic lupus erythematosus: Insights from a large monocentric cohort. Front. Immunol., 16.","DOI":"10.3389\/fimmu.2025.1529484"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Chang, H.C., Wu, Y.C., Chen, J.P., Wu, Y.D., Huang, W.N., Chen, Y.H., and Chen, Y.M. (2021). Comparisons of Anti-dsDNA Antibody Detection Methods by Chemiluminescent Immunoassay and Enzyme-Linked Immunosorbent Assay in Systemic Lupus Erythematosus. Diagnostics, 11.","DOI":"10.3390\/diagnostics11111940"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"160","DOI":"10.1007\/s00296-025-05910-7","article-title":"Sex- and gender-related differences in systemic lupus erythematosus: A scoping review","volume":"45","author":"Albrecht","year":"2025","journal-title":"Rheumatol. Int."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"369","DOI":"10.1002\/art.38238","article-title":"Population-based incidence and prevalence of systemic lupus erythematosus: The Michigan Lupus Epidemiology and Surveillance program","volume":"66","author":"Somers","year":"2014","journal-title":"Arthritis. Rheumatol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"293","DOI":"10.1177\/0961203313515763","article-title":"A surge in anti-dsDNA titer predicts a severe lupus flare within six months","volume":"23","author":"Pan","year":"2014","journal-title":"Lupus"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"102","DOI":"10.1016\/S0377-1237(10)80118-2","article-title":"Correlation between Systemic Lupus Erythematosus Disease Activity Index, C3, C4 and Anti-dsDNA Antibodies","volume":"66","author":"Narayanan","year":"2010","journal-title":"Med. J. Armed. Forces India"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1702","DOI":"10.2215\/CJN.01910213","article-title":"Elevated subclinical double-stranded DNA antibodies and future proliferative lupus nephritis","volume":"8","author":"Olson","year":"2013","journal-title":"Clin. J. Am. Soc. Nephrol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1819","DOI":"10.1007\/s10067-023-06577-w","article-title":"Negative ANA-IIF in SLE patients: What is beyond?","volume":"42","author":"Abozaid","year":"2023","journal-title":"Clin. Rheumatol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"863","DOI":"10.1007\/s10067-024-07288-6","article-title":"Identification of patients with ANA-negative and double-stranded DNA positive: What is the significance in daily rheumatology practice?","volume":"44","author":"Uzun","year":"2025","journal-title":"Clin. Rheumatol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"541","DOI":"10.1515\/cclm-2020-1408","article-title":"The clinical relevance of anti-dsDNA antibodies determined by the Elia\u2122 dsDNA assay in patients with negative indirect immunofluorescence on the HEp-2 cell","volume":"59","author":"Robier","year":"2020","journal-title":"Clin. Chem. Lab. Med."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1093\/jalm\/jfab146","article-title":"Anti-dsDNA testing specificity for systemic lupus erythematosus: A systematic review","volume":"7","author":"Orme","year":"2022","journal-title":"J. Appl. Lab. Med."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"e000984","DOI":"10.1136\/lupus-2023-000984","article-title":"Longitudinal study of patients with discrepant results in CLIFT and a solid-phase dsDNA antibody assay: Does a gold standard dsDNA assay exist?","volume":"10","author":"Navas","year":"2023","journal-title":"Lupus Sci. Med."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e13220","DOI":"10.1111\/sji.13220","article-title":"Comparison of current methods for anti-dsDNA antibody detection and reshaping diagnostic strategies","volume":"96","author":"Infantino","year":"2022","journal-title":"Scand. J. Immunol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"97","DOI":"10.1016\/j.autrev.2012.07.002","article-title":"Are anti-nucleosome antibodies a better diagnostic marker than anti-dsDNA antibodies for systemic lupus erythematosus? A systematic review and a study of metanalysis","volume":"12","author":"Bizzaro","year":"2012","journal-title":"Autoimmun. Rev."}],"container-title":["Antibodies"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4468\/15\/2\/23\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,3,5]],"date-time":"2026-03-05T10:48:45Z","timestamp":1772707725000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4468\/15\/2\/23"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,3,5]]},"references-count":24,"journal-issue":{"issue":"2","published-online":{"date-parts":[[2026,4]]}},"alternative-id":["antib15020023"],"URL":"https:\/\/doi.org\/10.3390\/antib15020023","relation":{},"ISSN":["2073-4468"],"issn-type":[{"value":"2073-4468","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,3,5]]}}}