Context-Aware Diagnostic Specificity (CADS).

Rapid detection of proteins is critical in a vast array of diagnostic or monitoring applications [...].

Prognostic Value of Histidine-Rich Glycoprotein for Community-Acquired Pneumonia.

PURPOSE: Histidine-rich glycoprotein (HRG) is abundant in serum and has been implicated in several processes including blood coagulation and immune response. This prospective study is aimed at exploring HRG as a biomarker in patients hospitalized for community-acquired pneumonia (CAP). METHODS: A total of 160 patients (73 severe CAP, 57 nonsevere CAP), and 30 healthy controls were enrolled in 2019. Demographic and clinical data were recorded for all patients. Serum HRG concentration was measured upon admission using ELISA. RESULTS: HRG levels were significantly lower in severe CAP patients compared with other groups, regardless of etiology, and were negatively correlated with serum interleukin-6 and disease severity index scores. Combination of CURB-65, PSI, and APACHE II scores with HRG values significantly improved the accuracy of predicting 30-day mortality in these patients. Cox regression analysis showed that HRG could serve as an independent risk factor for 30-day mortality. Notably, patients with HRG ≤ 16.92 µg/mL had significantly lower cumulative survival than those with HRG > 16.92 µg/mL. CONCLUSION: Serum HRG levels are lower in patients with severe CAP and are negatively correlated with disease severity scores. Measurement of HRG upon admission can provide valuable prognostic information for patients with CAP.

Recommendations and discussion points on immunogenicity, biomarkers, automation/technology and protein-MS from the 2021 European Bioanalysis Forum Focus Workshops.

During the first half of 2021, and due to the SARS-CoV-2 pandemic preventing in-person meetings, the European Bioanalysis Forum organized four workshops as live interactive online meetings. The themes discussed at the workshops were carefully selected to match the cyberspace dynamics of the meeting format. The first workshop was a training day on challenges related to immunogenicity. The second one focused on biomarkers and continued the important discussion on integrating the principles of Context of Use (CoU) in biomarker research. The third workshop was dedicated to technology, that is, cutting-edge development in cell-based and ligand-binding assays and automation strategies. The fourth was on progress and the continued scientific and regulatory challenges related to peptide and protein analysis with MS. In all four workshops, the European Bioanalysis Forum included a mixture of scientific and regulatory themes, while reminding the audience of important strategic aspects and our responsibility toward the patient.

Intranasal versus intratracheal exposure to lipopolysaccharides in a murine model of acute respiratory distress syndrome.

Due to frequent and often severe lung affections caused by COVID-19, murine models of acute respiratory distress syndrome (ARDS) are increasingly used in experimental lung research. The one induced by a single lipopolysaccharide (LPS) exposure is practical. However, whether it is preferable to administer LPS intranasally or intratracheally remains an open question. Herein, female C57Bl/6 J mice were exposed intranasally or intratracheally to one dose of either saline or 3 mg/kg of LPS. They were studied 24 h later. The groups treated with LPS, either intranasally or intratracheally, exhibited a pronounced neutrophilic inflammation, signs of lung tissue damage and protein extravasation into the alveoli, and mild lung dysfunction. The magnitude of the response was generally not different between groups exposed intranasally versus intratracheally. However, the variability of some the responses was smaller in the LPS-treated groups exposed intranasally versus intratracheally. Notably, the saline-treated mice exposed intratracheally demonstrated a mild neutrophilic inflammation and alterations of the airway epithelium. We conclude that an intranasal exposure is as effective as an intratracheal exposure in a murine model of ARDS induced by LPS. Additionally, the groups exposed intranasally demonstrated less variability in the responses to LPS and less complications associated with the sham procedure.

Sequential Protein Capture in Multiplex Single Molecule Arrays: A Strategy for Eliminating Assay Cross-Reactivity.

Measurements of multiple biomolecules within the same biological sample are important for many clinical applications to enable accurate disease diagnosis or classification. These disease-related biomarkers often exist at very low levels in biological fluids, necessitating ultrasensitive measurement methods. Single-molecule arrays (Simoa), a bead-based digital enzyme-linked immunosorbent assay, is the current state of the art for ultrasensitive protein detection and can detect sub-femtomolar protein concentrations, but its ability to achieve high-order multiplexing without cross-reactivity remains a challenge. Here, a sequential protein capture approach for multiplex Simoa assays is implemented to eliminate cross-reactivity between binding reagents by sequentially capturing each protein analyte and then incubating each capture bead with only its corresponding detection antibody. This strategy not only reduces cross-reactivity to background levels and significantly improves measurement accuracies, but also enables higher-order multiplexing. As a proof of concept, the sequential multiplex Simoa assay is used to measure five different cytokines in plasma samples from Coronavirus Disease 2019 (COVID-19) patients. The ultrasensitive sequential multiplex Simoa assays will enable the simultaneous measurements of multiple low-abundance analytes in a time- and cost-effective manner and will prove especially critical in many cases where sample volumes are limited.