An automated assay platform that robustly quantifies antibodies against up to 100 antigens in a single reaction, using Luminex suspension array technology, alongside generating ample quality control measures is described.
Extractable nuclear antigen (ENA) antibody testing is universal to the diagnosis of systemic autoimmune rheumatic diseases (SARDs). However, current commercial multiplex platforms require expensive instruments and are often cost-prohibitive for routine testing in low- and middle-income settings. Pictor PictHealth® Immunoscreen ENA assay was developed as an affordable, scalable alternative for the simultaneous detection of ENA antibodies. The assay performance was compared with the Euroimmun ENA Line immunoassay and BioPlex 2200 ENA assays using clinical sera from patients with autoimmune diseases. Concordance among assays was evaluated using three-way and pairwise analyses, expressed as positive percent agreement (PPA), negative percent agreement (NPA), and overall concordance. Analytical precision was assessed within and between manufacturing lots. The Pictor multiplex assay showed high concordance with established commercial platforms. Three-way agreement across all assays ranged from 67.3% for TRIM21/Ro52 to 100% for Jo-1, with Sm, CENP-B showing concordance of 95%. Pairwise comparisons demonstrated PPA and NPA values exceeding 85% for most autoantibodies. The Pictor PictHealth® Immunoscreen ENA assay provides diagnostic performance comparable to the Euroimmun and BioPlex platforms while maintaining the ease of implementation and scalability of a traditional ELISA format. These results support its potential as a cost-effective and accessible tool for ENA antibody testing in laboratories with varying resource levels.
Jung-Hyun Rho, Andrea Kinga, B. Shetty et al.· JIM - Journal of Immunologic...· 0 citations
Chimeric antigen receptor T-cell (CAR-T) therapy has revolutionized the treatment of refractory B-cell malignancies, yet severe cytokine release syndrome (CRS) and unpredictable long-term outcomes remain major clinical challenges. Here, we developed a rapid, scalable, 96-well plate-based strategy for the high-throughput preparation of lanthanide-encoded classifier beads. This approach substantially increases production throughput, from several types per day to 96 types within 8 hours, enabling sensitive multiplexed cytokine profiling by mass cytometry. By applying this platform to 136 longitudinal plasma samples from 32 patients with diffuse large B-cell lymphoma receiving CD19-directed CAR-T therapy, we achieved fg mL−1-level sensitivity and broad dynamic ranges across 14 key cytokines. Longitudinal profiling revealed distinct cytokine dynamics between patients with low- and high-grade CRS. Baseline TGF-β1 showed a significant univariate association with subsequent high-grade CRS (p = 0.001) and inferior event-free survival (p = 0.02). These findings demonstrate the utility of a scalable bead-based platform for high-resolution immune monitoring in CAR-T therapy and identify baseline TGF-β1 as a candidate exploratory biomarker associated with CRS severity and clinical outcome. This approach provides a practical tool for sensitive and high-throughput cytokine profiling and deeper understanding of immune responses in immunotherapy.
Keyuan Pu, Wanying He, Jiayi Feng et al.· Chemical Science· 0 citations
Monitoring prognostic biomarkers is essential for evaluating cancer treatment efficacy in real time. Here, we present a rapid, proof-of-concept microfluidic assay for cancer biomarker quantification and functional therapy evaluation. The assay requires only 5 µL of sample, eliminates tedious sample manipulation, and uses commercial antibodies to track both biomarker concentration and functional activity. We demonstrated the successful detection of VEGF and EML4-ALK proteins in both lung cancer cell culture supernatant and serum-spiked samples, achieving a detection sensitivity 15 times greater than standard ELISA using the same antibody pairs for the respective antigens. Crucially, we showcase the platform’s distinct applicability to functional assays by monitoring dynamic phosphorylation changes in EML4-ALK following treatment with the tyrosine kinase inhibitor alectinib, successfully capturing a significant, rapid decrease in phosphorylation levels. At this preclinical stage, the platform demonstrates analytical and functional feasibility for highly sensitive and rapid biomarker monitoring, providing a foundation for future validation in patient-derived ALK-positive samples for therapeutic-response assessment.
The multiplex‐bead immunoassay is characterized by its ability to detect multiple analytes simultaneously within a short timeframe and with high precision. Consequently, it is an appealing tool for identifying humoral immune responses to various targets, particularly respiratory viral infections that pose a significant risk to public health. In this study, we optimize the assay by using minimal reagent volumes to reduce both costs and turnaround time without compromising assay accuracy. We have developed a multiplex‐bead immunoassay to detect humoral responses against Severe Acute Respiratory Syndrome COVID‐19 2 (SARS‐CoV‐2) and Middle East Respiratory Syndrome COVID‐19 (MERS‐CoV) on a platform that can be extended to include additional targets. To validate the performance of the developed assay, serum samples from MERS‐CoV‐ and SARS‐CoV‐2‐seropositive and seronegative individuals were evaluated and compared with the standard enzyme‐linked immunosorbent assay (ELISA) method. The results demonstrate that the developed multiplex‐bead immunoassay exhibits high performance, achieving 100% sensitivity and 95.4%–96.6% specificity, a detection limit comparable to ELISA, and a strong correlation with ELISA (R 2 = 0.91–0.98) in detecting humoral responses in the tested samples using minimal reagents. Overall, the developed immunoassay offers a rapid method for simultaneously detecting humoral responses for multiple analytes, with high sensitivity and specificity and lower costs than ELISA.
Tarfa A. Altorki, M. Basabrain, Abdulaziz Kulthum et al.· Journal of Immunological Res...· 0 citations
Meningitis and encephalitis necessitate rapid pathogen identification to guide therapy, as conventional methods are time-consuming. This study evaluated both the wet-lab analytical performance and the clinical performance evaluation of the Bioeksen Meningitis/Encephalitis Panel (BS-MEP) integrated onto the fully automated, high-throughput Sigmoida Lab platform. Analytical limits of detection (LoD) were defined via Probit analysis (95% threshold) by spiking negative cerebrospinal fluid (CSF) matrices. Target detection was verified using characterized reference materials for all 14 analytes, and in silico primer/probe coverage was assessed against taxon-specific sequence databases. Cross-reactivity was evaluated using high-prevalence non-target organisms. Clinical performance was evaluated retrospectively using 500 archived, anonymized CSF specimens from a single-center repository, with analyte-specific classifications compared with prespecified routine comparator methods. The automated platform provided an 80-min sample-to-result turnaround for up to 23 samples. Verified LoDs ranged from 472 to 2118 genome copies/mL. All inclusivity strains were successfully detected in 5/5 replicates. In silico coverage exceeded 98% combined, and zero wet-lab cross-reactivity was observed. Precision coefficients of variation (CVs) were consistently ≤1.38%. In the clinical evaluation, the pooled clinical agreement was high, with PPA ranging from 90.9% to 100% and NPA of 100% across the evaluated analytes. Analyte-level clinical sensitivity ranged from 90.9% to 100%, with zero false-positive results across all targets. The fully automated molecular system demonstrates excellent analytical robustness and strong clinical agreement for syndromic detection of major CNS pathogens. The automated workflow combines broad pathogen coverage with an approximately 80-min sample-to-result turnaround and warrants further prospective evaluation in routine clinical settings.
Lateral flow assays (LFAs) have become the foundation of point‐of‐care (POC) diagnostics, valued for their user‐friendly format, cost‐effectiveness, and rapid turnaround time. However, despite their widespread success, most notably in pregnancy testing and infectious disease monitoring, conventional colourimetric LFAs often suffer from insufficient sensitivity, typically failing to reach sub‐picomolar detection limits, limiting their utility for the early‐stage detection of low‐abundance biomarkers. In recent years, aptamer‐based LFAs have emerged as a promising alternative to conventional antibody‐based formats owing to their superior stability, reproducibility and programmability. This review provides a critical analysis of recent strategies that have been developed to improve the sensitivity of aptamer‐based LFAs. Enhancement strategies are systematically classified into six distinct domains: (i) flow modulation techniques that optimise reaction kinetics; (ii) sample preconcentration methods; (iii) advanced signal transduction reporters beyond traditional gold nanoparticles; (iv) chemical signal amplification, including nanozyme and enzymatic catalysis; (v) structural strategies for maximising label accumulation; and (vi) the engineering of aptamers as programmable recognition elements. Particular emphasis is placed on amplification strategies that are compatible with or uniquely enabled by aptamer‐based detection systems. Finally, the emerging transition from passive biological selection to active molecular engineering is discussed, outlining the future trajectory of ultra‐sensitive next‐generation LFAs.
Aylar Eslami Saed, J. Giaretta, S. Farajikhah et al.· Visual Information Expert Wo...· 0 citations
We use cookies to run the site and, with your consent, for analytics and to show ads.
See our Cookie Policy.
