Molecular Crowding Evolution for Enabling Discovery of Enthalpy-Driven Aptamers for Robust Biomedical Applications

Area of Science:

• Biotechnology
• Molecular Biology
• Chemical Biology

Background:

• Enthalpy-driven ligands offer superior specificity and affinity for practical applications compared to entropy-driven ligands.
• A significant gap exists in direct discovery strategies for identifying enthalpy-driven ligands.

Purpose of the Study:

• To develop a molecular crowding Systematic Evolution of Ligands by Exponential Enrichment (SELEX) strategy for discovering enthalpy-driven aptamers.
• To improve aptamer affinity and selectivity in complex biological samples.

Main Methods:

• A novel molecular crowding SELEX strategy was employed.
• Aptamers were evolved against a tumor biomarker protein.
• Thermodynamic analysis was used to confirm enthalpy-driven binding.
• Aptamers evolved via conventional SELEX and molecular crowding SELEX (SYL-H2C) were compared.

Main Results:

• Three enthalpy-driven aptamer sequences were successfully evolved against a tumor biomarker.
• Molecular crowding SELEX (SYL-H2C) yielded aptamers with significantly higher affinity than conventional SELEX.
• The SYL-H2C aptamer demonstrated excellent accuracy in detecting circulating tumor cells in patient blood samples (10/10).

Conclusions:

• Molecular crowding SELEX is a feasible and effective strategy for discovering enthalpy-driven aptamers.
• The developed enthalpy-driven aptamers show promise as robust binding probes for biomedical applications.
• This screening strategy offers a new direction for developing high-performance aptamers for complex diagnostic scenarios.