Monitoring intracellular calcium in response to GPCR activation using thin-film silicon photodiodes with integrated fluorescence filters.
G-protein coupled receptor (GPCRs) drug discovery is a thriving strategy in the pharmaceutical industry. The standard approach uses living cells to test millions of compounds in a high-throughput format. Typically, changes in the intracellular levels of key elements in the signaling cascade are monitored using fluorescence or luminescence read-out systems, which require external equipment for signal acquisition. In this work, thin-film amorphous silicon photodiodes with an integrated fluorescence filter were developed to capture the intracellular calcium dynamics in response to the activation of the endogenous muscarinic M1 GPCR of HEK 293T cells. Using the new device it was possible to characterize the potency of carbachol (EC50=10.5 µM) and pirenzepine (IC50=4.2 μM), with the same accuracy as standard microscopy optical systems. The smaller foot-print provided by the detection system makes it an ideal candidate for the future integration in microfluidic devices for drug discovery.
INESC Microsistemas e Nanotecnologias (INESC MN), Rua Alves Redol 9, 1000-029 Lisbon, Portugal; IN-Institute of Nanoscience and Nanotechnology, Rua Alves Redol 9, 1000-029 Lisbon, Portugal; IBB-Institute for Biotechnology and Bioengineering, Instituto Superior Técnico, Avenida Rovisco Pais, 1049-001 Lisbon, Portugal.
S A M
SA
Martins
G
G
Moulas
J R C
JR
Trabuco
G A
GA
Monteiro
V
V
Chu
J P
JP
Conde
D M F
DM
Prazeres
D015374:Biosensing Techniques / Q000379:methods*; D002118:Calcium / Q000302:isolation & purification*; D002217:Carbachol / Q000737:chemistry; D005453:Fluorescence; D057809:HEK293 Cells; D006801:Humans; D046210:Microfluidic Analytical Techniques; D010890:Pirenzepine / Q000737:chemistry; D043643:Receptor, Muscarinic M1 / Q000737:chemistry; D012825:Silicon / Q000737:chemistry*
D043643:Receptor, Muscarinic M1; D010890:Pirenzepine; D002217:Carbachol; D002118:Calcium; D012825:Silicon
Amorphous silicon; Calcium; Cell; Fluorescence; GPCR; Integrated detection