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Cho, Lee, Park, and Lee: Utility of In-House PCR for HLA-B27 Typing: Comparison of Concordance Rate between PCR Kit and In-House PCR
Original Article

Korean J Leg Med 2008;28(3):239-243.

Published online: 14 February 2008

DOI: https://doi.org/10.3343/kjlm.2008.28.3.239

Utility of In-House PCR for HLA-B27 Typing: Comparison of Concordance Rate between PCR Kit and In-House PCR

Sun Young Cho, M.D., Kwang Gil Lee, M.T., Su Yon Park, M.D., Hee Joo Lee, M.D.

Department of Laboratory Medicine, Kyung-Hee Medical Center, School of Medicine, Kyoung-Hee University, Seoul, Korea

Received 30 January 200816 April 2008       Accepted 17 April 2008

Copyright © 2008 by the Korean Society for Legal Medicine

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background

Commercial kits of PCR method are widely used in HLA-B27 typing; however, their cost is relatively high. In this study, we evaluated the utility of an in-house PCR method by comparing it with that of a commercial kit.

Methods

HLA-B27 typing was done in 188 patients by using two PCR methods, Absolute™ HLA-B27 PCR kit (Biosewoom, Korea) and an in-house PCR method. The primers used in the in-house method were prepared by Bioneer (Korea). Both PCR tests were done by Gene Amp PCR System 9600 (Perkin-Elmer Centus Corp., USA).

Results

The commercial kit and in-house PCR showed 100% concordance rate with each other in HLA-B27 typing. Of 188 patients tested 72 (38.3%) were positive and 116 (61.7%) were negative by the both tests. Of 62 patients with ankylosing spondylitis, 50 were positive (80.7%).

Conclusions

The in-house PCR is a reliable and cost-effective method and can replace or supplement commercial kits for HLA-B27 typing.

Keywords: HLA-B27, In-house PCR, DNA typing

REFERENCES

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Fig. 1.
Results of HLA-B27 typing. (A) Absolute™ HLA-B27 PCR kit (Biosewoom), (B) In-house PCR with B27 280 primer.
kjlm-28-239f1.tif

Lanes 1-4, patient 1-4; P, positive control; N, negative control; a, internal control (beta-globin); c, internal control (P53 exon 8); b and d, HLA B27 band.

Table 1.
Patient characteristics and HLA-B27 typing results
Diseases N. of patients HLA- B27 positive cases N (%) Mean age yr Sex ratio M/F
Ankylosing spondylitis 62 50 (80.7) 35.9 48/14
Iridocyclitis 28 6 (21.4) 38.9 17/11
Rheumatism 16 5 (31.3) 32.1 11/5
Arthrosis 16 1 (6.3) 32.9 12/4
Spinal stenosis 11 2 (18.2) 36.2 6/5
Joint pain/myalgia 11 2 (18.2) 27.6 11/0
Optic neuritis 7 0 (0) 55.9 0/7
Other articular diseases∗ 8 4 (50.0) 34.8 5/3
Other diseases† 29 2 (6.7) 42.3 17/12
Total 188 72 (38.3) 37.0 127/61

∗ , HLA-B27 positive in synovitis, osteochondrosis and pyogenic arthritis;

† , HLA-B27 positive in diabetes and cerebral infarct.

Abbreviations: F, female; M, male; N, number.

Table 2.
Characteristics of primers used in the in-house PCR
Primer name   Primer sequence Locus Primer length (bp) Product length (bp)
B27 280 Sense 5′-GCTACGTGGACGACACGCT-3′ B∗2701-2709 19 149/150
  Antisense 5′-CTCGGTCAGTCTGTGCCTT-3′   19  
    5′-TCTCGGTAAGTCTGTGCCTT-3′   20  
Internal control Sense 5′-TAGGACCTGATTTCCTTACTGCCTC-3′ P53 exon 8 25 236
  Antisense 5′-AACTGCACCCTTGGTCTCCTCCACC-3′   25  

Abbreviation: bp, base pair.

Table 3.
Comparison of the test results in 188 patients by PCR kit and in-house PCR
    In-house PCR Total
Positive Negative
Commercial Kit Positive 72 0 72
 (Biosewoom) _negative 0 116 116
Total   72 116 188

Concordance rate=100%.

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