Is it possible to measure binding kinetics in solution? Now it is.
Is it possible to measure binding kinetics in solution? Now it is.
Scientific progress is driven by continuous evolution - binding kinetics are no exception.
For decades, immobilization-based methods were the standard for studying molecular interactions. While these techniques provided valuable insights, they also imposed fundamental limitations that researchers have long accepted as trade-offs: surface effects, complex assay setup, and restricted sample compatibility. But science moves forward, and we move with it. Now, in-solution kinetics is reshaping how we measure interactions—offering a more natural, efficient, and precise alternative.
Understanding molecular interactions is at the heart of structural biology, drug discovery, and biophysics. Binding kinetics—the rates at which molecules associate and dissociate—play a fundamental role in drug efficacy, protein function, and biomolecular stability.
How does surface (or lack of it) make a difference?
Traditional methods like Surface Plasmon Resonance (SPR) and Bio-Layer Interferometry (BLI) have long been used for kinetic studies. Importanlty, these methods (albeit highly useful) rely on surface immobilization, which can distort binding behavior. What if you could measure binding directly in solution, preserving native conditions and obtaining absolute kinetic data?
This is where Flow-Induced Dispersion Analysis (FIDA) offers a new perspective. Many biomolecular interactions occur in a complex, dynamic environment, where factors like protein conformation, oligomerization, and aggregation influence binding behavior. However, most kinetic assays require molecular immobilization, which introduces several challenges:
I. Steric hindrance: Surface attachment can block or alter binding sites.
II. Non-physiological conditions: Immobilized molecules may behave differently than in free solution
III. Mass transport effects: For fast kinetics, the mass transport to the surface may be the rate limiting step.Under such circumstances, assessment of binding kinetics becomes difficult or even impossible.
FIDA eliminates these barriers by measuring kinetics insolution, delivering absolute, label-free readouts.
Does FIDA measure kinetics differently?
Yes. Instead of relying on immobilization, FIDA tracks molecular size changes in free solution, using hydrodynamic radius (Rh)as a direct readout of binding interactions. Here’s how it works:
1. Baseline Measurement: The free molecule’s hydrodynamic radius (Rh) is determined in its native state.
2. Titration withBinding Partner: A series of increasing ligand or analyte concentrations is introduced.
3. Binding Detection: Any interaction that causes a size change (due to complex formation, conformational shifts, or oligomerization) is detected with absolute precision.
4. Binding (Kd) Analysis: By capturing size changes at different concentrations in equilibrated samples, the equilibrium interaction constant (Kd) can be determined without immobilization-induced artifacts.
5. Kinetic (kon and koff) Analysis: By controlling the reaction time, kinetic rate constants (kon and koff) can be obtained without immobilization-induced artifacts.
Watch an on-demand webinar, in which our CSO Henrik Jensen, Ph.D. explains in-solution kinetics in-depth. Or, if you prefer, take a look here.
Gaining scientific freedom and personal productivity
By removing immobilization-related artifacts and working directly in solution, you can work in more physiologically relevant and data-rich approach to kinetics.:
Preserve Native Molecular Behavior – No surface immobilization means no artificial constraints & a new level of scientific freedom
Work with Complex Samples – Measure unpurified samples, multi-subunit assemblies, and even membrane proteins.
Directly Measure Hydrodynamic Radius – Get insights into binding-induced conformational changes.
Minimize Sample Consumption – Use only µL-scale volumes, ideal for precious biomolecules.
Get Absolute Kinetic Readouts – Remove the need for assumption-based decisions, and use your data for ML and AI models
Read our user’s opinion on in-solution kinetics.
See how FIDA data matches SPR data.
Applications of In-solution Kinetics
What is it currenlty used for? FIDA’s absolute kinetic measurements are particularly valuable for:
🔬 Protein-Ligand Binding – Detect even subtle conformational changes upon binding.
🧬Oligomerization & Self-Assembly – Study how molecular size shifts during complex formation.
💊Drug Discovery & Screening – Evaluate binding kinetics for lead optimization.
🧪Structural Biology & Biophysics – Link kinetic profiles to protein stability and aggregation tendencies.
Feeling left with questions or doubts? We're happy to connect.
