Two-Dimensional Protein Nanoarray as a Carrier of Sensing Elements for Gold-Based Immunosensing Systems

Area of Science:

• Biotechnology
• Biosensing
• Surface Chemistry

Background:

• High-density protein immobilization on gold surfaces is crucial for sensitive immunosensing but challenging to achieve without activity loss.
• Existing methods often involve harsh chemical cross-linking, potentially damaging protein structure and function.
• A mild, biologically compatible method is needed for advanced biosensor development.

Purpose of the Study:

• To develop an improved, biologically mild method for homogeneous and high-density protein immobilization on gold surfaces.
• To enhance the sensitivity and performance of immunosensing platforms.
• To preserve the activity of immobilized proteins for reliable diagnostics.

Main Methods:

• Utilized a genetically encoded SpyTag-SpyCatcher interaction system for protein conjugation.
• Engineered a bacterial S-layer for ordered, two-dimensional nanoarray formation on gold.
• Combined the S-layer with the SpyTag-SpyCatcher system for covalent protein linkage.

Main Results:

• Achieved homogeneous and high-density protein immobilization on a gold surface.
• Significantly preserved the biological activity of immobilized African swine fever virus proteins.
• Demonstrated an eightfold increase in detection sensitivity compared to conventional chemical cross-linking methods.

Conclusions:

• The S-layer-based immobilization method offers an innovative approach for creating high-quality gold biosensing interfaces.
• This technique significantly enhances immunosensing sensitivity, aiding in pathogen detection and understanding host immunity.
• The biologically mild and efficient immobilization strategy holds promise for advanced biosensor applications.