Surfactant-Assisted Label-Free Fluorescent Aptamer Biosensors and Binding Assays

Area of Science:

• Biochemistry
• Analytical Chemistry
• Materials Science

Background:

• Label-free detection of aptamer binding commonly employs DNA staining dyes such as SYBR Green I (SGI) and thioflavin T (ThT).
• Cationic nature of these dyes leads to adsorption onto reaction vessel surfaces, causing signal instability and potential misinterpretation in binding assays and biosensor development.

Purpose of the Study:

• To evaluate and enhance the signal stability of DNA staining dyes in label-free aptamer detection.
• To identify methods for mitigating fluorescence signal decrease caused by dye adsorption in biosensing applications.

Main Methods:

• Signal stability assessment using thioflavin T (ThT) and an adenosine aptamer in different reaction vessels (polystyrene microplate vs. quartz cuvette).
• Screening of polymers and surfactants to counteract dye adsorption, with a focus on Triton X-100.
• Testing the sensitivity and signal stability of three aptamers (Hg²⁺, adenosine, cortisol) with and without Triton X-100.

Main Results:

• Signal instability was observed in polystyrene microplates, while quartz cuvettes provided more stable signals.
• Equilibration time was found to improve signal stability.
• 0.01% Triton X-100 demonstrated the most effective protection against fluorescence signal decrease due to dye adsorption.
• Aptamer sensitivity remained similar with Triton X-100, but signal stability was significantly enhanced.

Conclusions:

• Careful experimental design and control experiments are crucial for reliable results in dye-based aptamer detection.
• Incorporating Triton X-100 and employing longer equilibration times can substantially improve the reliability and stability of biosensor signals.