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Rapid detection of HLA ‐A*31:01 allele in DNA and blood samples using loop‐mediated isothermal amplification
Author(s) -
Cheung Y.K.,
Kwok M.,
Chan E.,
Kwan P.
Publication year - 2014
Publication title -
british journal of dermatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.304
H-Index - 179
eISSN - 1365-2133
pISSN - 0007-0963
DOI - 10.1111/bjd.12897
Subject(s) - loop mediated isothermal amplification , human leukocyte antigen , genotype , allele , typing , whole blood , genomic dna , confidence interval , medicine , dna , buccal swab , immunology , microbiology and biotechnology , antigen , biology , genetics , gene
Summary 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.