A microfluidic linear node array for the study of protein-ligand interactions.

Area of Science:

• Biochemistry
• Chemical Engineering
• Drug Discovery

Background:

• Protein-ligand binding is crucial for drug discovery.
• Existing methods for analyzing binding can be complex and time-consuming.
• Continuous separation techniques are needed for efficient analysis.

Purpose of the Study:

• To develop and validate a microfluidic device for continuous separation of small molecules from protein mixtures.
• To demonstrate the device's utility in studying protein-ligand interactions.
• To assess the device's performance using various fluorophores and binding assays.

Main Methods:

• A microfluidic device utilizing R and L nodes was engineered for continuous separation.
• High flow rates (up to 1300 μL h⁻¹) were employed to create a solution barrier for filtering protein complexes.
• The device's tunability was tested by adjusting flow rate and distribution ratios.
• Binding studies involved bovine serum albumin (BSA) and Eosin Y (EY) as a model system.

Main Results:

• The microfluidic device successfully separated unbound small molecules from protein-ligand complexes.
• Dose-dependent binding between BSA and EY was observed, reaching a plateau at a BSA:EY ratio above 1.
• The percentage decrease in small molecules correlated with increasing protein concentration, indicating binding.
• The microfluidic filter's performance was tunable and effective for various fluorophores.

Conclusions:

• The developed microfluidic device offers a novel and efficient method for continuous separation of small molecules.
• This technology is a promising tool for preliminary drug screening and studying protein-ligand binding.
• The device's high flow rate operation facilitates easy off-chip detection and analysis.