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Determination of 21-hydroxylase autoantibodies: inter-laboratory concordance in the Euradrenal International Serum Exchange Program

  • Alberto Falorni
  • Vittorio Bini
  • Corrado Betterle
  • Annalisa Brozzetti
  • Luis Castano
  • Marta Fichna
  • Olle Kampe
  • Gunnar Mellgren
  • Part Peterson
  • Shu Chen
  • Johan Ronnelid
  • Jochen Seissler
  • Claudio Tiberti
  • Raivo Uibo
  • Liping Yu
  • Åke Lernmark
  • Eystein Husebye
Publishing year: 2015
Language: English
Pages: 1761-1770
Publication/Series: Clinical Chemistry and Laboratory Medicine
Volume: 53
Issue: 11
Document type: Journal article
Publisher: De Gruyter

Abstract english

Background: 21-Hydroxylase autoantibodies (21OHAb) are markers of an adrenal autoimmune process that identifies individuals with autoimmune Addison's disease (AAD). Quality and inter-laboratory agreement of various 21OHAb tests are incompletely known. The objective of the study was to determine inter-laboratory concordance for 21OHAb determinations. Methods: Sixty-nine sera from 51 patients with AAD and 51 sera from 51 healthy subjects were blindly coded by a randomization center and distributed to 14 laboratories that determined 21OHAb, either by an "in-house" assay (n=9) using in vitro-translated S-35-21OH or luciferase-labeled 21OH or a commercial kit with I-125-21OH (n=5). Main outcome measures were diagnostic accuracy of each participating laboratory and inter-laboratory agreement of 21OHAb assays. Results: Intra-assay coefficient of variation ranged from 2.6% to 5.3% for laboratories using the commercial kit and from 5.1% to 23% for laboratories using "in-house" assays. Diagnostic accuracy, expressed as area under ROC curve (AUC), varied from 0.625 to 0.947 with the commercial kit and from 0.562 to 0.978 with "in-house" methods. Cohen's. of inter-rater agreement was 0.603 among all 14 laboratories, 0.691 among "in-house" laboratories, and 0.502 among commercial kit users. Optimized cutoff levels, calculated on the basis of AUCs, increased the diagnostic accuracy of every laboratory (AUC >0.9 for 11/14 laboratories) and increased the Cohen's. of inter-rater agreement. Discrepancies in quantitation of 21OHAb levels among different laboratories increased with increasing autoantibody levels. Conclusions: The quality of 21OHAb analytical procedures is mainly influenced by selection of cutoff value and correct handling of assay materials. A standardization program is needed to identify common standard sera and common measuring units.


  • Rheumatology and Autoimmunity
  • Addison's disease
  • adrenal antibodies
  • autoimmunity
  • diagnosis
  • RIA
  • standardization


  • Diabetes and Celiac Unit
  • ISSN: 1434-6621
E-mail: ake [dot] lernmark [at] med [dot] lu [dot] se

Principal investigator

Diabetes and Celiac Unit

+46 40 39 19 01

+46 70 616 47 79


Jan Waldenströms gata 35, Malmö


Lund University Diabetes Centre, CRC, SUS Malmö, Jan Waldenströms gata 35, House 91:12. SE-214 28 Malmö. Telephone: +46 40 39 10 00