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Rapid detection of HLA-A*31:01 allele in DNA and blood samples using loop-mediated isothermal amplification
1Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong SAR, China.
Background:
The human leucocyte antigen (HLA) allele, HLA-A*31:01, is a biomarker for adverse cutaneous reactions to carbamazepine, a first-line antiepileptic drug.
Objectives:
To develop a platform that can rapidly detect the HLA-A*31:01 allele in blood samples to facilitate pretreatment screening.
Methods:
A novel protocol based on loop-mediated isothermal amplification (LAMP) was designed and optimized. It was applied to purified genomic DNA samples derived from B-cell lines with known HLA genotypes, and to DNA and whole blood samples collected from patients with epilepsy, in whom HLA-A genotypes were determined by sequence-based typing.
Results:
The turnaround time for the LAMP-based protocol was < 45 min. In the DNA samples derived from B-cell lines (n = 66), the sensitivity, specificity, positive predictive value and negative predictive value of the LAMP-based protocol for detecting HLA-A*31:01 were 1·00 [95% confidence interval (CI) 0·88-1·00], 0·95 (95% CI 0·82-0·99), 0·94 and 1·00, respectively. The LAMP-based protocol produced the same results in the DNA and whole blood samples collected from patients (n = 34). Its sensitivity, specificity, positive predictive value and negative predictive value in detecting HLA-A*31:01 in the patient samples were 1·00 (95% CI 0·57-1·00), 0·97 (95% CI 0·83-0·99), 0·83 and 1·00, respectively.
Conclusions:
The findings demonstrated the feasibility of accurately detecting HLA-A*31:01 in DNA and whole blood samples using a LAMP-based protocol. Given its rapid turnaround time, this novel platform has the potential to be adapted into a point-of-care screening test.
