An iSCAT-Based Platform for Digital Determination of Affinity Constants (DiDAC)

Modern biological research relies heavily on antibodies to detect and measure proteins, from genetics research to diagnostics and drug development. Measuring binding kinetics for antibodies is important for inferring specificity, affinity, and its mechanism of action. Existing kinetics assays are powerful, but they often require large sample volumes and provide only averaged signals, making it difficult to precisely measure how strongly and how fast molecules bind.
This project aims to develop a new “digital” assay for measuring binding kinetics at the single-molecule level. Using an optical technique called interferometric scattering microscopy (iSCAT), individual binding and unbinding events will be directly observed on functionalized glass surfaces. Rather than inferring binding strength from bulk signals, this approach counts single binding events, enabling highly sensitive measurement of affinity constants with reduced reagent consumption.
The project will first establish the feasibility of digitally tracking single binding events using well-characterized molecular pairs, then benchmark the resulting measurements against surface plasmon resonance. Finally, the method will be extended to antibody “sandwich” assays, allowing direct visualization and troubleshooting of these binding interactions that are important for assay development.
By combining single-molecule sensitivity with surface-based measurements, this work aims to create a fast, low-cost, and highly informative tool for antibody characterization.

Faculty Supervisor:

David Juncker

Student:

Partner:

Max-Planck-Institut für die Physik des Lichts

Discipline:

Engineering

Sector:

Education

University:

McGill University

Program: