Intended purpose
The SALSA MLPA Probemix P070 Subtelomeres Mix 2B is an in vitro diagnostic (IVD)¹ or research use only (RUO) semi-quantitative assay² for the detection of deletion(s) or duplication(s) in subtelomeric regions in genomic DNA isolated from human peripheral whole blood specimens, buccal swabs, (un)cultured amniotic fluid obtained in week 16 of the pregnancy or later and free from blood contamination, (un)cultured chorionic villi free from maternal contamination, foetal blood or products of conception free from maternal contamination. P070 Subtelomeres Mix 2B is intended to confirm a potential cause for and clinical diagnosis of intellectual disability, developmental delay, congenital abnormalities and/or pregnancy loss and for molecular genetic testing of at-risk family members.

It is recommended that results of P070 Subtelomeres Mix 2B are confirmed with P036 Subtelomeres Mix 1. Copy number variations (CNVs) detected with one or both probemixes should be confirmed with a different technique. In particular, CNVs detected by only a single probe always require confirmation by another method.

Assay results are intended to be used in conjunction with other clinical and diagnostic findings, consistent with professional standards of practice, including confirmation by alternative methods, parental evaluation, 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 testing, population screening, or for the detection of, or screening for, acquired or somatic genetic aberrations.

¹ Please note that this probemix is for in vitro diagnostic (IVD) use in the countries specified at the end of this product description.
² To be used in combination with a SALSA MLPA Reagent Kit and Coffalyser.Net analysis software.

Clinical background
In the general population, the prevalence of intellectual disability and developmental delay is estimated at 1-3% and the prevalence of congenital anomalies at 2-3% (Castells-Sarret et al. 2017). Clinical signs can include dysmorphic features, language impairment, seizures, learning disabilities, behavioural disturbances and autism spectrum disorders. It has been recognized that worldwide the genetic etiology of individuals with non-syndromic intellectual disability remains undetermined in the majority of cases. Determining the etiology of intellectual disability and developmental delay is important and useful for pediatric neurologists, geneticists, pediatricians, and patients’ families because it allows assessment of recurrence risk, appropriate genetic counselling, and focus on treatment options and prognosis.

Aberrant copy numbers of subtelomeric regions, e.g. due to an unbalanced translocation, are a frequent cause of intellectual disability, developmental delay and/or congenital abnormalities lacking distinct syndromic features. Subtelomeric regions are more likely to be involved in copy number changes as these are less likely to be lethal due to the low amount of critical genes located in these regions. The differences in size and breakpoint location of chromosomal CNVs make the phenotype highly variable among patients, ranging from a non-viable foetus to a phenotypically normal individual. Chromosomal anomalies of the foetus are found in almost half of the miscarriages and are the most common reason for pregnancy loss (Bernatowicz et al. 2019). More information can be found on Decipher (https://decipher.sanger.ac.uk/) and in the references listed at the end of this product description.

Probemix content
The SALSA MLPA Probemix P070-B3 Subtelomeres Mix 2B contains 46 MLPA probes with amplification products between 132 and 490 nucleotides (nt). This includes 41 probe(s) for the subtelomeric regions, no probes are present for the subtelomeric regions on the short p-arm of the 5 acrocentric chromosomes (13, 14, 15, 21 and 22). For these, an extra probe is included detecting the q-arm, close to the centromere. The subtelomeric probes for the X and Y chromosome are identical as they detect sequences in the pseudo-autosomal regions (PAR1 and PAR2) which are identical in chromosome X and Y. More information is present in Table 1 of this document. Complete probe sequences are available online (www.mrcholland.com).

This probemix contains ten quality control fragments generating amplification products between 64 and 121 nt: four DNA Quantity fragments (Q-fragments), two DNA Denaturation fragments (D-fragments), one Benchmark fragment, and one chromosome X and two chromosome Y-specific fragments (see table below). 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.