Systematic functional screening of switchable aptamer beacon probes

Area of Science:

• Molecular Biology
• Biotechnology
• Analytical Chemistry

Background:

• Immunoassays rely on affinity probes like antibodies and aptamers, but complex steps hinder their use in challenging environments.
• Developing wash-free assays that signal upon target detection is a key goal for simplifying molecular analysis.

Purpose of the Study:

• To establish a systematic functional screening platform for developing switchable aptamer beacon probes.
• To enable target-responsive detection for simplified immunoassays.

Main Methods:

• Constructed a library of stem-loop, hairpin-shaped aptamer beacons on microbeads.
• Screened the library using target-responsive fluorescence-activated sorting.
• Utilized computational modeling to understand aptamer binding and structural switching mechanisms.

Main Results:

• Selected aptamer beacons demonstrated strong affinities and triggered fluorescence only upon target binding.
• Successfully enabled wash-free immunoassays for detecting intracellular and membrane proteins.
• Identified specific protein-aptamer interactions driving aptamer structural changes for signal activation.

Conclusions:

• The developed platform standardizes the generation of switchable aptameric tools.
• These aptamer beacons offer potential for advanced diagnostics and molecular biology research.
• Wash-free detection simplifies complex assay environments like intracellular settings and microfluidics.