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Accessible Unlicensed Requires Authentication Published by De Gruyter September 6, 2011

Rapid HLA-B27 screening with real-time TaqMan PCR: a clinical validation in the Dutch population

Elianne A. Roelandse-Koop, Bert Buisman, Erik J. van Hannen, Anneke van der Zee, Wouter Kortlandt, Mirjam H.A. Hermans, Arend-Jan van Houte and Rianne van Rhee-Luderer

Abstract

Background: Human leukocyte antigen B27 (HLA-B27) is strongly associated with ankylosing spondylitis. The B27 allele is present in 90% of patients with this disease, whereas it is present in only 9% of Caucasians. Molecular detection of HLA-B27 is traditionally based on allele specific amplification of exon 2 (Olerup method) or exon 3 (Dominguez method) by PCR, followed by gel analysis.

Methods: We developed a real-time TaqMan PCR based on the Dominguez method with a β-Globin PCR as internal control.

Results: A total of 544 clinical samples were used to compare the real-time TaqMan PCR with the traditional Dominguez PCR, the traditional Olerup PCR and a commercial Olerup based HLA-B27 detection kit (Olerup SSPTM HLA-B27, GenoVision). While 542 samples gave concordant results, two samples showed discrepancies and were further analyzed. One sample that showed a discrepancy was negative with the traditional Olerup method and positive with the three other procedures. Sequencing analysis showed the presence of HLA-B*2712 in this sample. The other sample, positive with both Olerup based PCRs and negative with both Dominguez based methods, turned out to be positive for HLA-B*2707 by sequence analysis.

Conclusions: With a correct result for 543 out of 544 samples (99.8%), we consider our real-time HLA-B27 PCR is a reliable method to detect HLA-B27 in the Dutch population, with reduced hands-on time and contamination risk compared to traditional PCR methods.


Corresponding author: Dr. Arend-Jan van Houte, Bosboomstraat 1, 3582 KE Utrecht, The Netherlands Phone: +31-882506697, Fax: +31-882506695

Received: 2010-11-29
Accepted: 2011-8-1
Published Online: 2011-9-6
Published in Print: 2011-12-1

©2011 by Walter de Gruyter Berlin Boston