Related Experiment Video
Updated: May 21, 2025
Multimodal Analytical Platform on a Multiplexed Surface Plasmon Resonance Imaging Chip for the Analysis of Extracellular Vesicle Subsets
Published on: March 17, 2023
Partition-Less Digital Immunoassay Using Configurable Topographic Nanoarrays for Extracellular Vesicle Diagnosis of
Yunjie Wen1, Yutao Li1, Shibo Cheng1
1Department of Chemistry, University of Florida, Gainesville, Florida 32611, United States.
Abstract:
Emerging digital bioassays provide opportunities for bioanalysis and clinical diagnostics due to their improved analytical performance. Prevailing digitization strategies rely on partitioning single molecules into discrete compartments for which bead-based capture may be used. Herein we report a partition-free digital enzyme-linked immunosorbent assay (dELISA) named microfluidic Topographically Intensified, Partition-less dELISA (μTIP-dELISA). Our method builds on a single-molecule signal amplification technique that employs a simple micropost device to generate a topographic nanogap array to significantly enhance surface-bound enzymatic reactions. Compared to existing dELISA methods, our approach features appreciable simplicity and enhanced adaptability as it obviates the needs for sophisticated device fabrication, complicated workflow for off-line immunomagnetic capture, and ultralow-volume compartmentalization. Moreover, μTIP-dELISA integrates the inherent advantages of microfluidics in improving the assay performance and throughput and enhancing the scalability and automation. As a proof-of-concept for potential biomedical applications, we adapted μTIP-dELISA to extracellular vesicle (EV)-based liquid biopsy diagnosis of a pediatric cancer, Ewing sarcoma (EWS). Our technology confers >300-fold improvement over the conventional ELISA in detecting four EWS protein biomarkers. We demonstrated highly sensitive and specific detection of EWS cases with a machine-learning-defined four-marker EV signature. The four EV markers were further tested to assess age as a prognostic factor, resulting in an overall accuracy of 97% for classifying the control, pediatric and adult subjects. Overall, we envision that our μTIP single-molecule signal amplification strategy could promote the development and adaptation of digital bioassays and that the μTIP-dELISA could provide a useful tool for clinical diagnostics for many malignancies.
