A Compartmentalization-Free Digital Immunoassay Based on Plasmonic-Fluorescence Nanoparticles

Area of Science:

• Biotechnology
• Nanotechnology
• Analytical Chemistry

Background:

• Ultrasensitive protein detection is vital for early disease diagnosis and precision medicine.
• Digital immunoassays offer operational simplicity and independence from large instrumentation.
• Effective single-molecule labels are critical for compartmentalization-free digital immunoassay performance.

Purpose of the Study:

• To develop a novel compartmentalization-free digital immunoassay using plasmonic-fluorescent nanoparticles (SA-Cy3@BSA-Bio@Au NPs) as single-molecule labels.
• To evaluate the performance of this new assay for ultrasensitive protein detection.
• To demonstrate a new label design strategy for sensitive and rapid digital immunoassays.

Main Methods:

• Synthesized plasmonic-fluorescent nanoparticles (SA-Cy3@BSA-Bio@Au NPs) by modifying gold nanoparticles (Au NPs) with biotinylated bovine serum albumin (BSA-Bio) and streptavidin-conjugated Cy3 (SA-Cy3).
• Developed a compartmentalization-free digital immunoassay platform utilizing these nanoparticles as labels.
• Evaluated the assay's performance using interleukin-6 (IL-6) as a model analyte, including detection limit and assay time, and validated with human serum samples.

Main Results:

• The synthesized SA-Cy3@BSA-Bio@Au NPs exhibited small size, high single-particle brightness, and good dispersibility.
• The developed compartmentalization-free digital immunoassay achieved an ultrasensitive detection limit of 95 fg/mL for IL-6.
• The assay time was significantly reduced to 75 minutes, and practical utility was confirmed in human serum samples.

Conclusions:

• This study presents a novel design strategy for single-molecule labels in digital immunoassays.
• The developed plasmonic-fluorescent nanoparticles enable sensitive, rapid, and compartmentalization-free protein detection.
• This approach holds promise for applications in early disease diagnosis, precision medicine, and life science research.