HFE C282Y PCR Assay
The Viennalab HFE C282Y RealFast Assay detects the C282Y variant in the HFE gene by real-time PCR and discriminates the normal, heterozygous and homozygous mutant genotypes in a human genomic DNA extract. C282Y is the most common genetic risk factor for hereditary haemochromatosis: about 80 per cent of patients are homozygous for this variant. Homozygous carriers run the highest risk of iron overload with liver cirrhosis, diabetes and cardiomyopathy, so identifying them early allows treatment before organ damage occurs.
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This kit in detail
- Detects HFE C282Y (g.10633G>A, rs1800562)
- Discriminates normal, heterozygous and homozygous mutant
- Real-time PCR with allele-specific hydrolysis probes, 144 bp amplicon
- WT-Control and MUT-Control supplied
- Whole peripheral blood, dried blood spots, buccal swabs or saliva
- 32 or 100 reactions per kit, REF 7-133 and REF 7-130
The Viennalab HFE C282Y RealFast Assay determines the genotype of the C282Y variant in the HFE gene, the most common genetic risk factor for hereditary haemochromatosis.
Clinical background
The HFE protein takes part in the regulation of iron absorption in the small intestine. The C282Y variant disturbs that regulation, so iron accumulates in the liver, heart and pancreas. About 80 per cent of patients with hereditary haemochromatosis are homozygous for C282Y. Compound heterozygotes, who carry both C282Y and H63D, are less common and have a milder iron absorption phenotype.
Interpretation
Homozygous carriers run the highest risk of iron overload with liver cirrhosis, diabetes and cardiomyopathy. Heterozygous carriers can develop iron-related complaints when other genetic or environmental factors are present. Because treatment by therapeutic phlebotomy is effective, identifying carriers before organ damage occurs is what determines the outcome.
Method
The assay is a real-time PCR based on the 5′ nuclease principle. A 144 bp fragment of the HFE gene is amplified and read with two allele-specific hydrolysis probes: HEX for the wild-type allele and FAM for the C282Y allele. Heterozygous samples give a signal in both channels. The kit supplies the RealFast 2x Genotyping Mix, the HFE C282Y Assay Mix, a WT-Control and a MUT-Control. The recommended DNA input is 10 to 100 ng per reaction, at a concentration of 2 to 20 ng/µl, and the limit of detection is 0.2 ng of genomic DNA per reaction. DNA isolated from whole peripheral blood, dried blood spots, buccal swabs or saliva can be used, and DNA extraction reagents are not supplied with the kit.
- RealFast™ 2x Genotyping Mix: Contains HotStart Taq DNA polymerase and dNTPs in an optimized buffer system for PCR amplification.
- HFE C282Y Assay Mix: Includes gene-specific primers targeting the HFE gene and two allele-specific, dual-labeled hydrolysis probes for genotype discrimination.
- HFE C282Y WT-Control: Used for validation, representing the wild-type (normal) genotype.
- HFE C282Y MUT-Control: Helps in accurate determination of results, representing the homozygous mutant genotype.
Datasheets and regulatory documentation
Everything your quality team needs for evaluation and procurement.
Instructions for Use
Frequently paired with this kit
Other assays and instruments lab teams add alongside the HFE C282Y PCR Assay.
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