A general double library SELEX strategy for aptamer selection using unmodified nonimmobilized targets

Area of Science:

• Biotechnology
• Molecular Biology
• Biochemistry

Background:

• Aptamer discovery for unmodified, non-immobilized targets presents a significant challenge.
• Existing SELEX (Systematic Evolution of Ligands by Exponential Enrichment) methods often require target modification or immobilization.

Purpose of the Study:

• To develop a novel and versatile aptamer selection strategy for unmodified targets.
• To demonstrate the utility of this strategy for discovering both ssDNA and RNA aptamers with high binding affinities.
• To enable the concurrent selection of aptamer pairs for sandwich assay development.

Main Methods:

• Implementation of a novel double library (DL) SELEX strategy.
• Selection of aptamers against small (doxycycline) and large (VEGF165) unmodified targets.
• Concurrent selection of aptamer pairs recognizing distinct epitopes on the same protein.

Main Results:

• Achieved nanomolar binding affinities for both ssDNA and RNA aptamers against diverse targets.
• Successfully selected aptamer pairs from the same selection cycles for sandwich biosensor applications.
• Demonstrated the strategy's effectiveness for targets that cannot be chemically modified.

Conclusions:

• The developed DL-SELEX method is a valuable and accessible strategy for aptamer discovery.
• This method is particularly advantageous when target molecule modification is not feasible or desired.
• The approach facilitates the development of aptamer-based biosensors using selected aptamer pairs.