The SALSA MLPA Probemix P178 F8 is an in vitro diagnostic (IVD)1
or research use only (RUO) semi-quantitative assay2
for the detection of deletions or duplications in the F8
gene in genomic DNA isolated from human peripheral whole blood specimens. P178 F8 is intended to confirm a potential cause for and clinical diagnosis of hemophilia A, and for molecular genetic testing of at-risk family members.
Copy number variations (CNVs) detected with P178 F8 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 F8
gene are intron 22 and intron 1 inversions and point mutations, none of which will be detected by MLPA. It is therefore recommended to use this assay in combination with targeted intron 22 and intron 1 inversion analysis and sequence analysis.
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, 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.
Please 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).
To be used in combination with a SALSA MLPA Reagent Kit and Coffalyser.Net analysis software.
Hemophilia A, one of the most common coagulation disorders, is caused by complete or partial deficiency in factor VIII (FVIII) clotting activity. Depending on the remaining level of FVIII clotting activity, three types of hemophilia A can be discriminated: (1) severe hemophilia A with <1% FVIII activity, (2) moderate hemophilia A with 1-5% FVIII activity, and (3) mild hemophilia A with 6-40% FVIII activity. Patients with hemophilia A experience prolonged bleeding or oozing following an injury, tooth extraction or surgery, and delayed or recurrent bleeding prior to complete wound healing. The major cause of disability from bleeding is chronic joint disease, and the leading cause of death related to bleeding is intracranial hemorrhage.
Hemophilia A is an X-linked recessive disease with a prevalence of ~1:5,000 live male births that is caused by mutations in the F8
gene encoding FVIII. The most common genetic defects in the F8
gene are intron 22 and intron 1 inversions, which occur in 43-45% and 2-5% of severe hemophilia A cases, respectively. The remaining types of pathogenic variants span the entire mutation spectrum, i.e. missense mutations, small deletions or insertions, nonsense mutations, splice site mutations, deletions and duplications. Deletions and duplications are found in ~5% of hemophilia A patients (http://f8-db.eahad.org/
; Kim et al. 2012; Lannoy et al. 2012; Miller et al. 2012; Vencesla et al. 2012), and include single- and multi-exon deletions and duplications throughout the F8
gene, as well as whole gene deletions. More information about hemophilia A is available at https://www.ncbi.nlm.nih.gov/books/NBK1404/
The SALSA MLPA Probemix P178-B4 F8 contains 43 MLPA probes with amplification products between 136 and 477 nt. This includes 33 probes for the F8
gene. Each exon is covered by at least one probe. Two probes are present for exon 1, 3, 7, 12 and 26, and three probes are present for exon 14. In addition, ten reference probes are included that detect locations on the X chromosome. 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