A one-step real-time PCR assay for detection of DQA1*05, DQB1*02 and DQB1*0302 to aid diagnosis of celiac disease.
Celiac disease is an autoimmune disorder that develops after dietary exposure of the small intestine to gluten peptides in cereals. Celiac disease has a strong genetic component associated with HLA-DQ2 and HLA-DQ8, and testing for absence of these genetic markers is useful when serological tests and biopsies are indeterminate, as it renders celiac disease highly unlikely. We have developed a new real-time PCR assay, using sequence-specific primers (PCR-SSP) and TaqMan probes, for detection of DQB1*05, DQB1*02 (coding for DQ2) and DQB1*0302 (coding for DQ8). PCR amplification and detection of DQ2 and DQ8 was accurately and unambiguously performed from genomic DNA isolated from cell lines and human DNA. Amplification was scored digitally, without laboratory manipulation of amplified PCR products and with a higher accuracy than PCR-SSP. This assay should increase accuracy and throughput, and reduce risks of contamination in laboratories where testing for HLA DQ2 and DQ8 is performed as part of diagnosis of celiac disease.
Reinton
Nils
N
Fürst Medisinsk Laboratorium, Søren Bulls vei 25, N-1051 Oslo, Norway.
Helgheim
Asle
A
Shegarfi
Hamid
H
Moghaddam
Amir
A
D002446:Celiac Disease / Q000097:blood / Q000235:genetics / Q000276:immunology*; D004247:DNA / Q000737:chemistry / Q000235:genetics; D017931:DNA Primers / Q000737:chemistry / Q000235:genetics; D015343:DNA Probes, HLA / Q000737:chemistry / Q000235:genetics; D006683:HLA-DQ Antigens / Q000097:blood* / Q000235:genetics / Q000276:immunology; D059848:HLA-DQ alpha-Chains; D059866:HLA-DQ beta-Chains; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA
D017931:DNA Primers; D015343:DNA Probes, HLA; D006683:HLA-DQ Antigens; D059848:HLA-DQ alpha-Chains; D059866:HLA-DQ beta-Chains; C109756:HLA-DQ8 antigen; C069050:HLA-DQA1 antigen; C069051:HLA-DQB1 antigen; D004247:DNA