Flow Cytometric Human Leukocyte Antigen-B27 Typing with Stored Samples for Batch Testing.
10.3343/alm.2013.33.3.174
- Author:
Bo Young SEO
1
;
Dong Il WON
Author Information
1. Department of Clinical Pathology, Kyungpook National University School of Medicine, Daegu, Korea. wondi@knu.ac.kr
- Publication Type:Original Article ; Research Support, Non-U.S. Gov't
- Keywords:
HLA-B27 typing;
Flow cytometry;
Sample storage;
Frozen platelets
- MeSH:
Blood Platelets/metabolism;
Erythrocytes/metabolism;
*Flow Cytometry;
Freezing;
HLA-B27 Antigen/*blood;
HLA-B7 Antigen/blood;
Histocompatibility Testing;
Humans;
Leukocytes, Mononuclear/metabolism;
Real-Time Polymerase Chain Reaction;
Spondylarthropathies/diagnosis;
Temperature
- From:Annals of Laboratory Medicine
2013;33(3):174-183
- CountryRepublic of Korea
- Language:English
-
Abstract:
BACKGROUND: Flow cytometry (FC) HLA-B27 typing is still used extensively for the diagnosis of spondyloarthropathies. If patient blood samples are stored for a prolonged duration, this testing can be performed in a batch manner, and in-house cellular controls could easily be procured. In this study, we investigated various methods of storing patient blood samples. METHODS: We compared four storage methods: three methods of analyzing lymphocytes (whole blood stored at room temperature, frozen mononuclear cells, and frozen white blood cells [WBCs] after lysing red blood cells [RBCs]), and one method using frozen platelets (FPLT). We used three ratios associated with mean fluorescence intensities (MFI) for HLAB27 assignment: the B27 MFI ratio (sample/control) for HLA-B27 fluorescein-5-isothiocyanate (FITC); the B7 MFI ratio for HLA-B7 phycoerythrin (PE); and the ratio of these two ratios, B7/B27 ratio. RESULTS: Comparing the B27 MFI ratios of each storage method for the HLA-B27+ samples and the B7/B27 ratios for the HLA-B7+ samples revealed that FPLT was the best of the four methods. FPLT had a sensitivity of 100% and a specificity of 99.3% for HLA-B27 assignment in DNA-typed samples (N=164) when the two criteria, namely, B27 MFI ratio >4.0 and B7/B27 ratio <1.5, were used. CONCLUSIONS: The FPLT method was found to offer a simple, economical, and accurate method of FC HLA-B27 typing by using stored patient samples. If stored samples are used, this method has the potential to replace the standard FC typing method when used in combination with a complementary DNA-based method.
