Napolitano, L. | bioRxiv Science

bioRxiv2025

Engineering a Dimeric Single-Domain Antibody for Improved Detection and neutralization of Amyloid-β oligomers

Soluble A{beta} oligomers are regarded as major neurotoxic agents in Alzheimers disease. Several monoclonal antibodies have been developed to target A{beta} oligomers, but most of them show limited specificity binding also to monomers and fibrils. To generate an antibody with high specificity for the oligomers, we aimed to increase the efficiency and sensitivity of a humanized A{beta}-oligomer-specific single domain antibody, called DesAb-O. We engineered a dimeric DesAb-O variant, DiDesAb-O, which showed a significantly improved binding properties for A{beta} oligomers as compared to the monomeric sdAb. Furthermore, DiDesAb-O detected A{beta}42 oligomers in cells, prevented their binding to cell membranes and the A{beta}42 oligomers-induced neurotoxicity from both synthetic A{beta}42 samples and cerebrospinal fluid of Alzheimers patients at lower concentrations compared to DesAb-O. Overall, our findings indicate that the rational engineering of dimeric sdAb variants is an effective strategy to improve their binding properties offering new opportunities for the development of clinical molecules in the early diagnosis and cure of Alzheimers disease. Significance StatementProtein aggregates, characterized by high structural heterogeneity, are central to numerous neurodegenerative diseases, yet the development of aggregate-specific molecular probes and therapeutic agents remains a significant challenge. We present a novel approach to enhance the binding of single-domain antibodies by engineering multivalent, specifically dimeric, molecules to exploit the avidity effect. As a proof of concept, we developed a dimeric version of a previously studied single-domain antibody named DesAb-O, which exhibits enhanced binding sensitivity and specificity to toxic amyloid-{beta} oligomers and greater efficacy in inhibiting their toxicity. These findings provide a robust foundation for creating next-generation antibody fragments with enhanced binding to heterogeneous protein aggregates, opening new avenues for innovative diagnostic tools and therapeutic strategies in neurodegenerative disease research and treatment.

biochemistry↗

bioRxiv2019

Rapid breath analysis for acute respiratory distress syndrome diagnostics using a portable 2-dimensional gas chromatography device

Acute respiratory distress syndrome (ARDS) is the most severe form of acute lung injury, responsible for high mortality and long-term morbidity. As a dynamic syndrome with multiple etiologies its timely diagnosis is difficult as is tracking the course of the syndrome. Therefore, there is a significant need for early, rapid detection and diagnosis as well as clinical trajectory monitoring of ARDS. Here we report our work on using human breath to differentiate ARDS and non-ARDS causes of respiratory failure. A fully automated portable 2-dimensional gas chromatography device with high peak capacity, high sensitivity, and rapid analysis capability was designed and made in-house for on-site analysis of patients breath. A total of 85 breath samples from 48 ARDS patients and controls were collected. Ninety-seven elution peaks were separated and detected in 13 minutes. An algorithm based on machine learning, principal component analysis (PCA), and linear discriminant analysis (LDA) was developed. As compared to the adjudications done by physicians based on the Berlin criteria, our device and algorithm achieved an overall accuracy of 87.1% with 94.1% positive predictive value and 82.4% negative predictive value. The high overall accuracy and high positive predicative value suggest that the breath analysis method can accurately diagnose ARDS. The ability to continuously and non-invasively monitor exhaled breath for early diagnosis, disease trajectory tracking, and outcome prediction monitoring of ARDS may have a significant impact on changing practice and improving patient outcomes.

bioengineering↗