The rise of neurodegenerative disorders (NDD) in the ageing population is increasing demand on already stretched Health Services and is a major financial burden for society (1). The development of tools for early diagnosis is one of the responses to this problem (2). Cerebrospinal fluid (CSF) is the main repository of by-products of neuronal destruction (3); as serial lumbar punctures to procure CSF may be impractical in advanced patients (4), blood may represent the ideal source to track any meaningful disease signal of neurodegeneration.High-costs and low energy efficiency have pushed alternative means of samples collection and storage. Noviplex dried plasma spot (Np-DPS) and dried blood spot (DBS) on filter paper can be a remote, quick and inexpensive way of obtaining blood microsamples for the measurement of a large number of analytes in non-hospitalized, public health settings (5).We have used commercially-available and highly sensitive immunodetection assays (6) to test neurofilament light chain (Nf-L) expression in elute from DBS and from Np-DPS and compared these to standard Nf-L plasma measurement of healthy controls and patients with amyotrophic lateral sclerosis (ALS).We show that DBS and Np-DP cards can be used for remote blood collection and measurement of Nf-L. Nf-L measurement using elute from Noviplex DPS cards is equivalent to that obtained in plasma from the same blood samples, while levels of the same analyte in DBS elute are different, but provide discrimination between controls and ALS patients. Normalization using known loading protein concentration may also be needed for DBS analysis to adjust for the chromatographic effect of retained haemoglobin in the spot elute. Conversely, a simple elution step is required for Np-DPS, which reconstitutes a test fluid which is indistinguishable from plasma originated by conventional blood processing.
Objective. To evaluate serum and cerebrospinal fluid (CSF) levels of phosphorylated neurofilament heavy (pNfH), and to compare these to levels of neurofilament light (NfL), as biomarkers of pre-symptomatic ALS. Design. The study population includes 34 controls, 79 individuals at-risk for ALS, 22 ALS patients, and 14 phenoconverters. At-risk individuals are enrolled through Pre-Symptomatic Familial ALS (Pre-fALS), a longitudinal natural history and biomarker study of individuals who are carriers of any ALS-associated gene mutation, but who demonstrate no clinical evidence of disease at the time of enrollment. pNfH and NfL in serum and CSF were quantified using established enzyme-linked immunosorbent assays. Results. There is a longitudinal increase in serum pNfH in advance of the emergence of clinically manifest ALS. A similar pattern is observed for NfL, but with the absolute levels also frequently exceeding a normative threshold. Although CSF data are more sparse, similar patterns are observed for both neurofilaments, with absolute levels exceeding a normative threshold prior to phenoconversion. In serum, these changes are observed in the 6-12 months prior to disease among SOD1 A4V mutation carriers, and as far back as 2 and 3.5 years, respectively, in individuals with a FUS c.521del6 mutation and a C9ORF72 hexanucleotide repeat expansion. Conclusions. Serum and CSF pNfH increase prior to phenoconversion. In CSF, the temporal course of these changes is similar to NfL. In serum, however, pNfH is less sensitive to pre-symptomatic disease than NfL. The duration of pre-symptomatic disease, as defined by changes in neurofilaments, may vary depending on underlying genotype.
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