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Updated: Jul 15, 2026
Determination of High-affinity Antibody-antigen Binding Kinetics Using Four Biosensor Platforms
Published on: April 17, 2017
Concentration dependence of IgG-protein A affinity studied by wireless-electrodeless QCM
Hirotsugu Ogi1, Kazuma Motohisa, Kenichi Hatanaka
1Graduate School of Engineering Science, Osaka University, Machikaneyama 1-3, Toyonaka, Osaka 560-8531, Japan. ogi@me.es.osaka-u.ac.jp
Insights
This study utilized a novel wireless-electrodeless quartz crystal microbalance (QCM) to precisely measure the binding affinity between human immunoglobulin G (IgG) and protein A. Results revealed varying equilibrium constants, highlighting the complexity of IgG-protein A complex formation.
Area of Science:
- Biophysics
- Analytical Chemistry
- Materials Science
Background:
- Protein A is crucial for purifying human immunoglobulin G (IgG).
- Accurate measurement of binding affinity is essential for optimizing purification processes.
- Existing methods may have limitations in sensitivity or non-contact measurement capabilities.
Purpose of the Study:
- To develop and validate a wireless-electrodeless quartz crystal microbalance (QCM) system for studying IgG-Protein A interactions.
- To quantify the binding affinity between human IgG and immobilized Protein A.
- To investigate the influence of IgG concentration on binding kinetics and complex formation.
Main Methods:
- Immobilization of Protein A onto an electrodeless AT-cut quartz crystal.
- Non-contact measurement of the quartz crystal's fundamental resonance frequency using a line antenna.
- Utilizing a flow-cell system for continuous frequency monitoring during IgG injection.
- Applying Langmuir kinetics to analyze frequency changes and determine equilibrium constants.
Main Results:
- The wireless-electrodeless QCM demonstrated high sensitivity for detecting binding events.
- Equilibrium constants (K(A)) ranged from 6 x 10^6 to 6 x 10^10 M^-1.
- Observed variations in K(A) correlated with different IgG concentrations, suggesting concentration-dependent complex formation.
Conclusions:
- The developed wireless-electrodeless QCM is a viable tool for sensitive, non-contact analysis of biomolecular interactions.
- The study provides valuable kinetic data on human IgG and Protein A binding.
- Understanding the concentration-dependent complex formation is key for refining immunopurification strategies.
Abstract:
The binding affinity between human immunoglobulin G (IgG) and protein A was studied by the homebuilt wireless-electrodeless quartz crystal microbalance (QCM). Protein A was immobilized on the electrodeless AT-cut quartz plate of 0.05 mm thick and its fundamental resonance frequency near 34 MHz was measured by a noncontacting manner using a line antenna. The vibrational analysis was performed to ensure higher sensitivity of the electrodeless QCM. A flow-cell system was fabricated to continuously measure the resonance frequency during the injection sequence of the IgG solutions with concentrations of 1-20,000 ng/mL. The exponential frequency changes were recorded to determine the affinity based on the Langmuir kinetics. The equilibrium constant K(A) significantly varied between 6 x 10(6) and 6 x 10(10) M(-1), depending on the IgG concentration, which is attributed to various formations of IgG-protein A complexes.
