A living cell-based biosensor utilizing G-protein coupled receptors: principles and detection methods.

Area of Science:

• Biomedical Engineering
• Cell Biology
• Biosensor Technology

Background:

• G protein-coupled receptors (GPCRs) are crucial cell surface receptors involved in signal transduction.
• Cell-based biosensors offer high sensitivity and specificity for detecting various analytes.
• Existing biosensor technologies can be limited by cell viability and genetic modification capabilities.

Purpose of the Study:

• To investigate the feasibility of using bullfrog fibroblast (FT) cells expressing GPCRs as a core component of a living cell-based biosensor.
• To develop a robust platform for culturing FT cells on microelectrode arrays for long-term biosensing applications.
• To demonstrate the detection of GPCR activation using both optical and electrochemical methods.

Main Methods:

• Fabrication of gold microelectrode arrays on silicon dioxide substrates for cell culture.
• Culturing of bullfrog FT cells under ambient conditions on the fabricated substrates.
• Monitoring GPCR activation via optical detection of intracellular calcium and electrochemical detection of norepinephrine secretion.
• Genetic modification of FT cells to express reporter genes, indicating potential for novel GPCR expression.

Main Results:

• Successful long-term, robust growth of FT cells on microelectrode arrays at room temperature.
• Demonstrated detection of endogenous GPCR activation by ATP using optical and electrochemical methods.
• Confirmed the potential for genetic modification of FT cells for expressing novel GPCRs.

Conclusions:

• Bullfrog FT cells expressing GPCRs are a viable foundation for developing living cell-based biosensors.
• The developed MEMS device supports cell growth and enables monitoring of cellular responses.
• This approach facilitates the creation of next-generation BioMEMS devices leveraging the sensitivity and selectivity of GPCRs.