Background: Human leukocyte antigen B27 (HLA-B27) is a major histocompatibility complex class 1 molecule that is strongly associated with ankylosing spondylitis. The polymerase chain reaction-sequence specific primers (PCR-SSP) is currently used to detect HLA-B27 antigen. However, this technique still requires multiple steps, time consuming and cost-effective due to post PCR process. The real-time PCR and melting curve analysis provides a convenient and rapid way to detect this antigen.
Objective: To develop the real-time PCR and melting curve analysis for HLA-B27 allele in routine work.
Materials and Methods: All 50 patient samples had been previously detect HLA-B27 antigen with the conventional PCR-SSP method were enrolled in this study. The real-time PCR melting curve analysis for HLA-B27 was compared with previous HLA-B27 antigen results.
Results: 25 HLA-B27 negative samples had only one peak of β-globin gene at 84.90±0.09 °C while all 25 HLA-B27 positive samples showed single or double peaks at 87.28±1.36 ℃ and/or 84.90±0.09 °C. The results of real-time PCR melting curve analysis showed a perfect concordance with the conventional PCR-SSP method.
Conclusion: The real-time PCR methods appear to be reliable, very fast in the detection of HLA-B27 and therefore a suitable in routine laboratory practice.