Anouk Donners

163 General Discussion mutation affecting the assays, neither OSA nor CSA perfectly predict patients’ bleeding phenotypes in people with non-severe haemophilia A [31]. The second future application is to use LC-MS/MS bioanalysis in cases of discrepancy between the FVIII activity and the clinical phenotype. As mentioned, the non-null mutations partially synthesise FVIII, and LC-MS/MS analysis can quantify these mutated peptides (non-functional or partially functional) in human plasma. As many mutated peptides as desired can be included (multiplexing), beginning with the most common. The entire mutated-peptide profile could eventually be screened. The FVIII concentration measurements using an LC-MS/MS method could add value to the existing clotting assays, identifying genetic factors in PwHA at risk of discrepancies, and thus misdiagnosis, when using common clotting assays. Measurements using LC-MS/MS could eventually be included in the first steps of diagnosing of haemophilia A. 3. Neonatal screening A third future application of LC-MS/MS could be the inclusion of haemophilia A in a neonatal screening programme. The Dutch neonatal screening programme aims to detect 26 treatable and severe disorders early in life [32]. The selection of these disorders was based on Wilson and Jungner’s principles of screening, published in 1963, which are still consulted regarding the inclusion of new disorders [33]. Haemophilia A is a potential candidate for the screening programme when applying these principles because half of new-borns have no family history and it being a serious and severe disorder that can be treated with access to accurate diagnostic tools and adequate therapies in the Netherlands. Why is this application of value? Diagnostic tests are currently performed for new-borns with a known family history of haemophilia A [34, 35]. Therefore, neonatal screening would help new-borns without a known family history of haemophilia A (±50% of cases), because this diagnosis rarely occurs until later in life, usually when a bleed occurs. Overall, approximately 80% of PwHA are at risk of a bleed during their first two years of life [36]. The most serious bleed, the intracranial haemorrhage (ICH), is also most likely to occur in the first two years of life [36, 37] and has been estimated to occur in 3–4% of newborns [38]. Including haemophilia A in neonatal screening with LC-MS/MS analysis could facilitate earlier diagnosis, allowing clinicians to begin prophylaxis with FVIII concentrates sooner and preventing bleeds in the first days, weeks or years of life. 4. Dried blood-spot sampling The second benefit of using LC-MS/MS analysis in neonatal screening is the sampling approach [39]. Sampling for neonatal screening involves collecting heel-prick blood drops from a neonate in the first week after delivery on a screening filter paper card. The dried blood spots (DBS) are then punched and analysed within a week [32, 40]. The LC-MS/MS approach has very high sensitivity with the ability to measure very low FVIII 9