Rapid Detection of the Three Celiac Disease Risk Genotypes HLA-DQ2.2, HLA-DQ2.5, and HLA-DQ8 by Multiplex Ligation-Dependent Probe Amplification

Vijzelaar, Raymon; Zwan, Ellen van der; Gammeren, Adriaan van; Yilmaz, Rizkat; Verheul, Alice; Hoogstraten, Ingrid van; Baar, Ellen de; Schrauwen, Lianne; Kortlandt, W.

doi:10.1089/gtmb.2015.0233

Cited by 3 publications

(2 citation statements)

References 12 publications

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“…17 To address these limitations, more efficient and cost-effective alternatives have been proposed with the use of HLAtagging single nucleotide polymorphisms, 17 real-time PCR, 20 and multiplex ligation-dependent probe amplification. 21 A common feature of these approaches is a reliance on PCR-based technologies in combination with sensitive fluorescence-based detection of amplified products. This restricts their application to laboratory settings in which costly DNA purification, thermal-cycling, and fluorescence detection equipment are available.…”

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confidence: 99%

“…22,23 LAMP is a rapid, isothermal nucleic acid amplification technique that uses a strand displacement Bst DNA polymerase and four to six sequence-specific primers. 21,24 LAMP reactions typically require <60 minutes of incubation and function reliably across a broad temperature range (2 C to 4 C), eliminating the need for high accuracy thermal-cycling equipment. 25 Furthermore LAMP assays are more tolerant than PCRbased technologies to inhibitors in clinical samples and may be performed directly on minimally processed blood 22 or saliva.…”

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confidence: 99%

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Rapid, Loop-Mediated Isothermal Amplification Detection of Celiac Disease Risk Alleles

Erlichster

Tye–Din

Varney

et al. 2018

The Journal of Molecular Diagnostics

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Human leukocyte antigen (HLA) genotyping has become a useful investigation in the diagnostic work-up of celiac disease (CD), with utility in risk stratification and screening. However, broad application of this technology has been hindered by the cost and time burden of conventional laboratory-based assays. We have developed and validated CD-loop-mediated isothermal amplification (CD-LAMP), a LAMP assay, which enables rapid identification of the signature CD risk genotypes, HLA-DQ2.5, HLA-DQ8, HLA-DQ2.2, and HLA-DQA1*05. Sample-to-answer is achieved in approximately 65 minutes without DNA purification, thermal cycling, or specialized analytical equipment. CD-LAMP genotyping of samples was 100% concordant with accredited pathology genotyping on a panel of 40 blood and 20 saliva samples. In a panel of 100 purified DNA samples, genotyping of the high-risk DQ2.5 genotype was 100% concordant with accredited pathology genotyping, with slightly reduced sensitivity for the DQ8 genotype (97.1%) and reduced specificity for the DQ8 (93.9%) and DQ2.2 (95.1%) genotypes. CD-LAMP results are easily visualized and instrument free through the addition of a DNA intercalating dye after amplification. Combined with point-of-care antibody testing, CD-LAMP may enable immediate, confident CD diagnosis at a low cost in the clinical setting.

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confidence: 99%