A hydrogel-based versatile screening platform for specific biomolecular recognition in a well plate format.

Area of Science:

• Biomaterials Science
• Surface Chemistry
• Biotechnology

Background:

• High-throughput determination of biomolecular interactions requires specialized surface coatings for reliable ligand immobilization.
• Existing methods often face challenges with non-specific binding and limited versatility in immobilizing diverse biomolecules.

Purpose of the Study:

• To develop a one-step hydrogel coating system for 96-well microtiter plates that facilitates versatile biomolecule immobilization.
• To create a surface resistant to non-specific protein and cell adhesion while maintaining high specificity for biomolecular recognition.
• To demonstrate the immobilization and quantification of various biochemically relevant ligands.

Main Methods:

• Utilized isocyanate-functional six-arm poly(ethylene oxide)-based star polymers for hydrogel coating.
• Applied the coating to commercially available 96-well microtiter plates in a one-step process.
• Validated surface resistance to non-specific adsorption using fluorescently labeled bovine serum albumin and primary human dermal fibroblasts (HDF).
• Demonstrated immobilization of peptides (GRGDS, GRGDSK-biotin), biocytin, fibronectin, and carbohydrates (N-acetylglucosamine, N-acetyllactosamine).
• Established a simplified enzyme-linked immunosorbent assay (ELISA) protocol for ligand detection and quantification.
• Assessed cell adhesion on modified surfaces using HDF.

Main Results:

• The one-step hydrogel coating system effectively immobilizes a wide range of biomolecules, including peptides, proteins, and carbohydrates.
• The coating demonstrated significant resistance to non-specific adsorption of proteins and adhesion of human dermal fibroblasts.
• High specificity for biomolecular recognition processes was achieved upon ligand immobilization.
• A simplified ELISA protocol was successfully established for quantifying immobilized ligands.
• The system is compatible with standard high-throughput screening formats.

Conclusions:

• The presented hydrogel coating system offers a straightforward, robust, and versatile solution for biomolecule immobilization in microtiter plates.
• This technology enhances the precision of high-throughput screening assays by minimizing non-specific interactions and enabling specific biomolecular recognition.
• The system's adaptability to various biomolecules and its compatibility with routine testing make it suitable for diverse applications in life sciences research and diagnostics.