Intended purpose
The SALSA MLPA Probemix P062 LDLR is an in vitro diagnostic (IVD)
1 or research use only (RUO) semi-quantitative assay
2 for the detection of deletions or duplications in the
LDLR gene in genomic DNA isolated from human peripheral whole blood specimens. P062 LDLR is intended to confirm a potential cause for and clinical diagnosis of familial hypercholesterolaemia and for molecular genetic testing of at-risk family members.
Copy number variations (CNVs) detected with P062 LDLR should be confirmed with a different technique. In particular, CNVs detected by only a single probe always require confirmation by another method. Most defects in the
LDLR gene are point mutations, none of which will be detected by MLPA. It is therefore recommended to use this assay in combination with sequence analysis.
Assay results are intended to be used in combination with other clinical and diagnostic findings, consistent with professional standards of practice, including confirmation by alternative methods, clinical genetic evaluation, and counselling, as appropriate. The results of this test should be interpreted by a clinical molecular geneticist or equivalent.
This device is not intended to be used for standalone diagnostic purposes, pre-implantation or prenatal testing, population screening, or for the detection of, or screening for, acquired or somatic genetic aberrations.
1Please note that this probemix is for in vitro diagnostic (IVD) use in the countries specified at the end of this product description. In all other countries, the product is for research use only (RUO).
2To be used in combination with a SALSA MLPA Reagent Kit and Coffalyser.Net analysis software.
Clinical background
The low-density lipoprotein receptor (LDLR) is a cell surface receptor that plays an important role in cholesterol homeostasis. The receptor binds and internalises LDL cholesterol particles by endocytosis. This process takes place in all nucleated cells, but mainly in the liver, which is responsible for ~70% of the total clearance of LDL cholesterol from the circulation.
Familial hypercholesterolaemia (FH) is a disease characterised by significantly elevated LDL cholesterol levels that cause atherosclerotic plaque deposition in arteries, which may lead to coronary artery disease (CAD) or other cardiovascular disease manifestations at an early age. Heterozygous FH is an autosomal dominant disease with a worldwide prevalence of ~1:250 that is caused by pathogenic mutations in the
LDLR,
APOB or
PCSK9 genes. Mutations in
LDLR are the most common, with >50% of FH cases caused by pathogenic variants in the
LDLR gene. Mutations in
APOB and
PCSK9 are found in 5-10% and <1% of FH cases, respectively. In the remaining ~40% of FH cases, the underlying genetic defect is unknown. Homozygous FH is much rarer (1:160,000-400,000) and results from biallelic mutations (homozygous or compound heterozygous) in one of the aforementioned genes or one mutation in each of two different genes. Autosomal recessive FH has a prevalence lower than 1:1,000,000 and is caused by biallelic mutations in the LDL-receptor adaptor protein 1 (
LDLRAP1) gene. Less than 1% of FH cases are due to mutations in
LDLRAP1.
The severity of symptoms is dependent on the type of mutation; complete loss of function variants usually lead to more severe disease. Heterozygous FH patients are at an approximately 20-fold increased risk for CAD, whereas most homozygous FH patients experience severe CAD in their mid-20s and undergo coronary bypass surgery in their teenage years (
https://www.ncbi.nlm.nih.gov/books/NBK174884/; Nordestgaard et al. 2013; Raal et al. 2011; Kassner et al. 2014).
Probemix content
The SALSA MLPA Probemix P062-D2 LDLR contains 33 MLPA probes with amplification products between 136 and 490 nucleotides (nt). This includes 20 probes for the
LDLR gene and one flanking probe for the
SMARCA4 gene located upstream of the
LDLR gene. In addition, 12 reference probes are included that detect autosomal chromosomal locations. Complete probe sequences and the identity of the genes detected by the reference probes are available online (
www.mrcholland.com).
This probemix contains nine quality control fragments generating amplification products between 64 and 105 nt: four DNA Quantity fragments (Q-fragments), two DNA Denaturation fragments (D-fragments), one Benchmark fragment, and one chromosome X and one chromosome Y-specific fragment. More information on how to interpret observations on these control fragments can be found in the MLPA General Protocol and online at
www.mrcholland.com.