In the present study we describe an ELISA to quantify the light subunit of the neurofilament triplet protein (NFL) in CSF. The method was validated by measuring CSF NFL concentrations in healthy individuals and in two well-characterized groups of patients with amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD). The levels were increased in ALS (1,743~1,661 ng/L; mean ± SD) and AD (346 ± 176 ng/L) compared with controls (138 ± 31 ng/L; p < 0.0001 for both). Within the ALS group, patients with lower motor neuron signs only had lower NFL levels (360 ± 237 ng/L) than those with signs of upper motor neuron disease (2,435 ± 1,633 ng/L) (p <0.05). In a second study patients with miscellaneous neurodegenerative diseases were investigated (vascular dementia, olivopontocerebellar atrophy, normal pressure hydrocephalus, cerebral infarctions, and multiple sclerosis), and the CSF NFL level was found to be increased (665 ± 385 ng/L; p < 0.0001). NFL is a main structural protein of axons, and we suggest that CSF NFL can be used to monitor neurodegeneration in general, but particularly in ALS with involvement of the pyramidal tract.

URL: https://www.clinicaltrials.gov. Unique identifier: NCT01872884.

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BACKGROUND: Little solid information is available on the possible risks for neuronal injury in amateur boxing. OBJECTIVE: To determine whether amateur boxing and severity of hits are associated with elevated levels of biochemical markers for neuronal injury in cerebrospinal fluid. DESIGN: Longitudinal study. SETTING: Referral center specializing in evaluation of neurodegenerative disorders. PARTICIPANTS: Fourteen amateur boxers (11 men and three women) and 10 healthy male nonathletic control subjects. INTERVENTIONS: The boxers underwent lumbar puncture 7–10 days and 3 months after a bout. The control subjects underwent LP once. MAIN OUTCOME MEASURES: Neurofilament light protein, total tau, glial fibrillary acidic protein, phosphorylated tau, and beta‐amyloid protein 1–40 (Abeta([1–40])) and 1–42 (Abeta([1–42])) concentrations in cerebrospinal fluid were measured. RESULTS: Increased levels after a bout compared with after 3 months of rest from boxing were found for two markers for neuronal and axonal injury, neurofilament light protein (mean±SD, 845±1140 vs 208±108 ng/L; P=0.008) and total tau (mean±SD, 449±176 vs 306±78 ng/L; P=0.006), and for the astroglial injury marker glial fibrillary acidic protein (mean±SD, 541±199 vs 405±138 ng/L; P=0.003). The increase was significantly higher among boxers who had received many hits (>15) or high‐impact hits to the head compared with boxers who reported few hits. In the boxers, concentrations of neurofilament light protein and glial fibrillary acidic protein, but not total tau, were significantly elevated after a bout compared with the nonathletic control subjects. With the exception of neurofilament light protein, there were no significant differences between boxers after 3 months of rest from boxing and the nonathletic control subjects. CONCLUSIONS: Amateur boxing is associated with acute neuronal and astroglial injury. If verified in longitudinal studies with extensive follow‐up regarding the clinical outcome, analyses of cerebrospinal fluid may provide a scientific basis for medical counseling of athletes after boxing or head injury.

Miniaturization and silicon integration of micro enzyme reactors for applications in micro total analysis systems (µTASs) require new methods to achieve structures with a large surface area onto which the enzyme can be coupled. This paper describes a method to accomplish a highly efficient silicon microstructured enzyme reactor utilizing porous silicon as the carrier matrix. The enzyme activity of microreactors with a porous layer was recorded and compared with a microreactor without the porous layer. The microreactors were fabricated as flow-through cells comprising 32 channels, 50 µm wide, spaced 50 µm apart and 250 µm deep micromachined in 110 oriented silicon, p type (20-70 cm), by anisotropic wet etching. The overall dimension of the microreactors was 13.1 × 3.15 mm. To make the porous silicon layer, the reactor structures were anodized in a solution of hydrofluoric acid and ethanol. In order to evaluate the surface enlarging effect of different pore morphologies, the anodization was performed at three different current densities, 10, 50 and 100 mA cm −2. Glucose oxidase was immobilized onto the three porous microreactors and a non-porous reference reactor. The enzyme activity of the reactors was monitored following a colorimetric assay. To evaluate the glucose monitoring capabilities, the reactor anodized at 50 mA cm −2 was connected to an FIA system for glucose monitoring. The system displayed a linear response of glucose up to 15 mM using an injection volume of 0.5 µl. The result from the studies of glucose turnover rate clearly demonstrates the potential of porous silicon as a surface enlarging matrix for micro enzyme reactors. An increase in enzyme activity by a factor of 100, compared to the non-porous reference, was achieved for the reactor anodized at 50 mA cm −2 .

Background and purposeNeurofilament light chain (NfL) is a marker of neuroaxonal damage. We aimed to study associations between serum NfL (sNfL) concentrations at different time points after ischemic stroke and outcomes.MethodsWe prospectively included ischemic stroke cases (n = 595, mean age 59 years, 64% males) and assessed outcomes by both the modified Rankin Scale (mRS) and the NIH stroke scale (NIHSS) at 3 months and by mRS at 2 years. In a subsample, long-term (7-year) outcomes were also assessed by both mRS and NIHSS. We used the ultrasensitive single-molecule array assay to measure sNfL in the acute phase (range 1–14, median 4 days), after 3 months and 7 years in cases and once in controls (n = 595).ResultsAcute-phase sNfL increased by the time to blood-draw and highest concentrations were observed at 3 months post-stroke. High sNfL associated to stroke severity and poor outcomes, and both associations were strongest for 3-month sNfL. After adjusting for age, previous stroke, stroke severity, and day of blood draw, 3-month sNfL was significantly associated to both outcomes at all time points (p < 0.01 throughout). For all main etiological subtypes, both acute phase and 3-month sNfL were significantly higher than in controls, but the dynamics of sNfL differed by stroke subtype.ConclusionsThe results from this study inform on sNfL in ischemic stroke and subtypes over time, and show that sNfL predicts short- and long-term neurological and functional outcomes. Our findings suggest a potential utility of sNfL in ischemic stroke outcome prediction.Electronic supplementary materialThe online version of this article (10.1007/s00415-019-09477-9) contains supplementary material, which is available to authorized users.

Background Neurofilament light (NFL) is a well-validated biomarker for neuronal injury and neurodegeneration. Increased cerebrospinal fluid (CSF) levels have been shown after stroke, as well as in patients with a broad range of neurodegenerative and neuroinflammatory diseases. Neurofilament heavy (NFH) belongs to the same family of structural proteins but it is less extensively studied. The potential of phosphorylated NFH (pNFH) as a stroke biomarker and for the prediction of clinical outcome is unknown. In this study, we aimed to examine the temporal pattern of NFL and pNFH concentrations in serum and CSF after acute ischemic stroke. Materials and MethodsA quantitative Enzyme-Linked ImmunoSorbent Assay (ELISA) for pNFH was developed and tested on CSF and serum samples. NFL and pNFH were analysed in serum and CSF of acute ischemic stroke patients, who were followed over time (Day 0-1, Day 2-3, Day 7-9, three weeks, and 3-5 months after stroke).Results NFL and pNFH concentrations in serum and CSF increased after stroke, peaked during the 3rd week, and then decreased back to almost baseline levels at 3-5 months. CSF-NFL and serum-NFL correlated to the outcome measured by Barthel Index after 3-5 months, whilst no such association was seen for pNFH.Discussion These findings suggest that NFL and pNFH in both CSF and serum reflect the temporal pattern of the post ischemic axonal injury and that this process does not seem to progress after 3-5 months.Conclusion NFL and pNFH in CSF and serum are promising biomarkers for axonal injury following stroke. Further studies in larger populations are needed to fully understand the progression of the neuronal damage after acute ischemic stroke and to evaluate if these biomarkers can provide additive information and how they relate to outcome.

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