High performance aptamer affinity chromatography for single-step selective extraction and screening of basic protein...

Area of Science:

• Biochemistry
• Analytical Chemistry
• Materials Science

Background:

• Affinity chromatography is crucial for protein purification.
• Developing selective methods for basic proteins like lysozyme remains a challenge.
• DNA aptamers offer high specificity for molecular recognition.

Purpose of the Study:

• To develop a high-performance affinity chromatography method for selective lysozyme extraction.
• To demonstrate the specificity of DNA aptamers for lysozyme capture.
• To create an efficient screening strategy for lysozyme in complex biological matrices.

Main Methods:

• Synthesis of a poly(glycidyl methacrylate-co-ethylene dimethacrylate) monolithic column.
• Covalent immobilization of an anti-lysozyme DNA aptamer onto the monolith surface via a spacer arm.
• High-performance affinity chromatography for protein separation and analysis.

Main Results:

• The aptamer-functionalized monolith selectively trapped lysozyme, distinguishing it from non-target proteins.
• Specific recognition, not electrostatic interaction, was confirmed as the primary trapping mechanism.
• High inter-day precision was achieved (0.8% for migration time, 4.2% for peak area).
• Successful selective extraction of lysozyme from chicken egg white was demonstrated.

Conclusions:

• DNA aptamer affinity chromatography provides a highly selective and efficient method for lysozyme purification and screening.
• The developed method offers advantages of low cost and ease of operation.
• This approach has significant potential for applications in diagnostics and biotechnology.