title: One-step molecular detection of the MYD88 L265P mutation by unlabeled probe genotyping analysis.
abstract: The aim of our study was to establish an unlabeled probe genotyping approach for rapid detection of the MYD88 L265P mutation in the differential diagnosis of Waldenstrӧm macroglobulinemia patients. Analytical and clinical validation of the assay was performed using serially diluted amplicon-cloned standards, 14 clinical bone marrow aspirate samples, and 30 peripheral blood samples from healthy donors, respectively. The analytical validation results showed that the assay is able to reproducibly identify as low as 10% of the L265P mutant. Clinical validation results showed improved detection sensitivity for the L265P mutation compared to Sanger sequencing. With the simplicity, cost-effectiveness, specificity and rapidity, we foresee that the unlabeled probe HRM assay is a good alternative to substitute current established methods for routine diagnostic testing of the MYD88 L265P mutation.
authors.affiliation: Molecular Diagnosis Centre, Department of Laboratory Medicine, National University Health System, Singapore.
authors.forename: Kok-Siong
authors.initials: KS
authors.lastname: Poon
authors.affiliation: Division of Haemato-Oncology, Department of Medicine, National University Health System, Singapore; Division of Haematology, Department of Laboratory Medicine, National University Health System, Singapore.
authors.forename: Te-Chih
authors.initials: TC
authors.lastname: Liu
authors.affiliation: Department of Laboratory Medicine, National University Health System, Singapore.
authors.forename: Karen Mei-Ling
authors.initials: KM
authors.lastname: Tan
authors.affiliation: Molecular Diagnosis Centre, Department of Laboratory Medicine, National University Health System, Singapore; Department of Pathology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore. Electronic address: evelyn_koay@nuhs.edu.sg.
authors.forename: Evelyn Siew-Chuan
authors.initials: ES
authors.lastname: Koay
mesh_terms: D000943:Antigens, Differentiation / Q000235:genetics*; D005838:Genotype; D006801:Humans; D007930:Leucine / Q000235:genetics; D025202:Molecular Diagnostic Techniques / Q000191:economics / Q000379:methods*; D009154:Mutation; D015345:Oligonucleotide Probes / Q000235:genetics*; D011392:Proline / Q000235:genetics; D015203:Reproducibility of Results; D017422:Sequence Analysis, DNA; D008258:Waldenstrom Macroglobulinemia / Q000134:cerebrospinal fluid / Q000175:diagnosis* / Q000235:genetics; D000097642:GADD45 Proteins
chemical_list: D000943:Antigens, Differentiation; D007930:Leucine; D015345:Oligonucleotide Probes; D011392:Proline; C086470:GADD45B protein, human; D000097642:GADD45 Proteins
keywords: High-resolution melt; L265P; MYD88; Unlabeled probe; Waldenstrӧm macroglobulinemia