Fully automated measurement of plasma Aß42/40 and p-tau181: Analytical robustness and concordance with cerebrospinal fluid profile along the Alzheimer's disease continuum in two independent cohorts.

INTRODUCTION: For routine clinical implementation of Alzheimer's disease (AD) plasma biomarkers, fully automated random-access platforms are crucial to ensure reproducible measurements. We aimed to perform an analytical validation and to establish cutoffs for AD plasma biomarkers measured with Lumipulse. METHODS: Two cohorts were included. UNIPG: n = 450 paired cerebrospinal fluid (CSF)/plasma samples from subjects along the AD-continuum, subjects affected by other neurodegenerative diseases, and controls with known CSF profile; AMS: n = 40 plasma samples from AD and n = 40 controls. Plasma amyloid ß (Aß)42, Aß40, and p-tau181 were measured with Lumipulse. We evaluated analytical and diagnostic performance. RESULTS: Lumipulse assays showed high analytical performance. Plasma p-tau181 levels accurately reflected CSF A+/T+ profile in AD-dementia and mild cognitive impairment (MCI)-AD, but not in asymptomatic-AD. Plasma and CSF Aß42/40 values were concordant across clinical AD stages. Cutoffs and probability-based models performed satisfactorily in both cohorts. DISCUSSION: The identified cutoffs and probability-based models represent a significant step toward plasma AD molecular diagnosis.

Neurofilament light protein as a biomarker for spinal muscular atrophy: a review and reference ranges.

Spinal muscular atrophy (SMA) is the leading genetic cause of infant mortality, characterized by progressive neuromuscular degeneration resulting from mutations in the survival motor neuron (SMN1) gene. The availability of disease-modifying therapies for SMA therapies highlights the pressing need for easily accessible and cost-effective blood biomarkers to monitor treatment response and for better disease management. Additionally, the wide implementation of newborn genetic screening programs in Western countries enables presymptomatic diagnosis of SMA and immediate treatment administration. However, the absence of monitoring and prognostic blood biomarkers for neurodegeneration in SMA hinders effective disease management. Neurofilament light protein (NfL) is a promising biomarker of neuroaxonal damage in SMA and reflects disease progression in children with SMA undergoing treatment. Recently, the European Medicines Agency issued a letter of support endorsing the potential utilization of NfL as a biomarker of pediatric neurological diseases, including SMA. Within this review, we comprehensively assess the potential applications of NfL as a monitoring biomarker for disease severity and treatment response in pediatric-onset SMA. We provide reference ranges for normal levels of serum based NfL in neurologically healthy children aged 0-18 years. These reference ranges enable accurate interpretation of NfL levels in children and can accelerate the implementation of NfL into clinical practice.

Methods to Discover and Validate Biofluid-Based Biomarkers in Neurodegenerative Dementias.

Neurodegenerative dementias are progressive diseases that cause neuronal network breakdown in different brain regions often because of accumulation of misfolded proteins in the brain extracellular matrix, such as amyloids or inside neurons or other cell types of the brain. Several diagnostic protein biomarkers in body fluids are being used and implemented, such as for Alzheimer's disease. However, there is still a lack of biomarkers for co-pathologies and other causes of dementia. Such biofluid-based biomarkers enable precision medicine approaches for diagnosis and treatment, allow to learn more about underlying disease processes, and facilitate the development of patient inclusion and evaluation tools in clinical trials. When designing studies to discover novel biofluid-based biomarkers, choice of technology is an important starting point. But there are so many technologies to choose among. To address this, we here review the technologies that are currently available in research settings and, in some cases, in clinical laboratory practice. This presents a form of lexicon on each technology addressing its use in research and clinics, its strengths and limitations, and a future perspective.

Multi-Omics Interdisciplinary Research Integration to Accelerate Dementia Biomarker Development (MIRIADE).

Proteomics studies have shown differential expression of numerous proteins in dementias but have rarely led to novel biomarker tests for clinical use. The Marie Curie MIRIADE project is designed to experimentally evaluate development strategies to accelerate the validation and ultimate implementation of novel biomarkers in clinical practice, using proteomics-based biomarker development for main dementias as experimental case studies. We address several knowledge gaps that have been identified in the field. First, there is the technology-translation gap of different technologies for the discovery (e.g., mass spectrometry) and the large-scale validation (e.g., immunoassays) of biomarkers. In addition, there is a limited understanding of conformational states of biomarker proteins in different matrices, which affect the selection of reagents for assay development. In this review, we aim to understand the decisions taken in the initial steps of biomarker development, which is done via an interim narrative update of the work of each ESR subproject. The results describe the decision process to shortlist biomarkers from a proteomics to develop immunoassays or mass spectrometry assays for Alzheimer's disease, Lewy body dementia, and frontotemporal dementia. In addition, we explain the approach to prepare the market implementation of novel biomarkers and assays. Moreover, we describe the development of computational protein state and interaction prediction models to support biomarker development, such as the prediction of epitopes. Lastly, we reflect upon activities involved in the biomarker development process to deduce a best-practice roadmap for biomarker development.

Clinical and analytical comparison of six Simoa assays for plasma P-tau isoforms P-tau181, P-tau217, and P-tau231.

INTRODUCTION: Studies using different assays and technologies showed highly promising diagnostic value of plasma phosphorylated (P-)tau levels for Alzheimer's disease (AD). We aimed to compare six P-tau Simoa assays, including three P-tau181 (Eli Lilly, ADx, Quanterix), one P-tau217 (Eli Lilly), and two P-tau231 (ADx, Gothenburg). METHODS: We studied the analytical (sensitivity, precision, parallelism, dilution linearity, and recovery) and clinical (40 AD dementia patients, age 66±8years, 50%F; 40 age- and sex-matched controls) performance of the assays. RESULTS: All assays showed robust analytical performance, and particularly P-tau217 Eli Lilly; P-tau231 Gothenburg and all P-tau181 assays showed robust clinical performance to differentiate AD from controls, with AUCs 0.936-0.995 (P-tau231 ADx: AUC = 0.719). Results obtained with all P-tau181 assays, P-tau217 Eli Lilly assay, and P-tau231 Gothenburg assay strongly correlated (Spearman's rho > 0.86), while correlations with P-tau231 ADx results were moderate (rho < 0.65). DISCUSSION: P-tau isoforms can be measured robustly by several novel high-sensitive Simoa assays.

Sensitive and quantitative detection of cardiac troponin I with upconverting nanoparticle lateral flow test with minimized interference.

Measurement of cardiac troponin I (cTnI) should be feasible for point-of-care testing (POCT) to diagnose acute myocardial infarction (AMI). Lateral flow immunoassays (LFIAs) have been long implemented in POCT and clinical settings. However, sensitivity, matrix effect and quantitation in lateral flow immunoassays (LFIAs) have been major limiting factors. The performance of LFIAs can be improved with upconverting nanoparticle (UCNP) reporters. Here we report a new methodological approach to quantify cTnI using UCNP-LFIA technology with minimized plasma interference. The performance of the developed UCNP-LFIA was evaluated using clinical plasma samples (n = 262). The developed UCNP-LFIA was compared to two reference assays, the Siemens Advia Centaur assay and an in-house well-based cTnI assay. By introducing an anti-IgM scrub line and dried EDTA in the LFIA strip, the detection of cTnI in plasma samples was fully recovered. The UCNP-LFIA was able to quantify cTnI concentrations in patient samples within the range of 30-10,000 ng/L. The LoB and LoD of the UCNP-LFIA were 8.4 ng/L and 30 ng/L. The method comparisons showed good correlation (Spearman's correlation 0.956 and 0.949, p < 0.0001). The developed UCNP-LFIA had LoD suitable for ruling in AMI in patients with elevated cTnI levels and was able to quantify cTnI concentrations in patient samples. The technology has potential to provide simple and rapid assay for POCT in ED setting.

Double-Antigen Lateral Flow Immunoassay for the Detection of Anti-HIV-1 and -2 Antibodies Using Upconverting Nanoparticle Reporters.

Rapid diagnostic tests (RDTs) are often used for the detection of anti-human immunodeficiency virus (HIV) antibodies in remote locations in low- and middle-income countries (LMIC) with low or limited access to central laboratories. The typical format of an RDT is a lateral flow assay (LFA) with visual interpretation prone to subjectivity. This risk of misinterpretation can be overcome with luminescent upconverting nanoparticle reporters (UCNPs) measured with a miniaturized easy-to-use reader instrument. An LFA with UCNPs for anti-HIV-1/2 antibodies was developed and the assay performance was evaluated extensively with challenging patient sample panels. Sensitivity (n = 145) of the UCNP-LFA was 96.6% (95% CI: 92.1-98.8%) and specificity (n = 309) was 98.7% (95% CI: 96.7-99.7%). Another set of samples (n = 200) was used for a comparison between the UCNP-LFA and a conventional visual RDT. In this comparison, the sensitivities for HIV-1 were 96.4% (95% CI: 89.8-99.3%) and 97.6% (95% CI: 91.6-99.7%), for the UCNP-LFA and conventional RDT, respectively. The specificity was 100% (95% CI: 96.4-100%) for both assays. The developed UCNP-LFA demonstrates the applicability of UCNPs for the detection of anti-HIV antibodies. The signal measurement is done by a reader instrument, which may facilitate automated result interpretation, archiving and transfer of data from de-centralized locations.

Glycovariant-based lateral flow immunoassay to detect ovarian cancer-associated serum CA125.

Cancer antigen 125 (CA125) is a widely used biomarker in monitoring of epithelial ovarian cancer (EOC). Due to insufficient cancer specificity of CA125, its diagnostic use is severely compromised. Abnormal glycosylation of CA125 is a unique feature of ovarian cancer cells and could improve differential diagnosis of the disease. Here we describe the development of a quantitative lateral flow immunoassay (LFIA) of aberrantly glycosylated CA125 which is widely superior to the conventional CA125 immunoassay (CA125IA). With a 30 min read-out time, the LFIA showed 72% sensitivity, at 98% specificity using diagnostically challenging samples with marginally elevated CA125 (35-200 U/mL), in comparison to 16% sensitivity with the CA125IA. We envision the clinical use of the developed LFIA to be based on the substantially enhanced disease specificity against the many benign conditions confounding the diagnostic evaluation and against other cancers.