Reagentless, reusable, ultrasensitive electrochemical molecular beacon aptasensor

Area of Science:

• Biomedical Engineering
• Electrochemistry
• Nucleic Acid Aptamer Technology

Background:

• Thrombin is a key biomarker in hemostasis and thrombosis.
• Development of sensitive and specific biosensors for thrombin detection is crucial for clinical diagnostics.
• Aptamers offer a promising alternative to antibodies as biorecognition elements due to their stability and specificity.

Purpose of the Study:

• To synthesize a bifunctional thrombin-binding aptamer derivative.
• To develop an electrochemical aptasensor for sensitive thrombin detection.
• To characterize the electrochemical properties and performance of the developed aptasensor.

Main Methods:

• Synthesis of a ferrocene-labeled aptamer with a thiol group.
• Self-assembly of the aptamer on a gold electrode surface to form a mixed monolayer.
• Electrochemical characterization using cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS).
• Investigation of thrombin-binding using a fluorescent molecular beacon and electrochemical techniques.

Main Results:

• The aptamer-modified electrode exhibited a redox signal characteristic of the surface-confined ferrocenyl moiety.
• Blocking with 2-mercaptoethanol enhanced the DPV signal, indicating improved aptamer immobilization and specific thrombin interaction.
• Thrombin binding induced a conformational change in the aptamer, leading to a "signal-on" electrochemical response.
• A linear correlation between DPV current and thrombin concentration (5.0–35.0 nM) with a detection limit of 0.5 nM was achieved.
• The aptasensor demonstrated excellent regenerability over 25 cycles without loss of electrochemical signal.

Conclusions:

• The developed electrochemical aptasensor provides a sensitive and reliable method for thrombin detection.
• The aptasensor's performance is attributed to the specific aptamer-thrombin interaction and the conformational change upon binding.
• The high regenerability suggests potential for real-time monitoring applications.