Applying label-free dynamic mass redistribution technology to frame signaling of G protein-coupled receptors noninvasively in living cells.
Label-free dynamic mass redistribution (DMR) is a cutting-edge assay technology that enables real-time detection of integrated cellular responses in living cells. It relies on detection of refractive index alterations on biosensor-coated microplates that originate from stimulus-induced changes in the total biomass proximal to the sensor surface. Here we describe a detailed protocol to apply DMR technology to frame functional behavior of G protein-coupled receptors that are traditionally examined with end point assays on the basis of detection of individual second messengers, such as cAMP, Ca(2+) or inositol phosphates. The method can be readily adapted across diverse cellular backgrounds (adherent or suspension), including primary human cells. Real-time recordings can be performed in 384-well microtiter plates and be completed in 2 h, or they can be extended to several hours depending on the biological question to be addressed. The entire procedure, including cell harvesting and DMR detection, takes 1-2 d.
Molecular-, Cellular- and Pharmacobiology Section, Institute of Pharmaceutical Biology, University of Bonn, Bonn, Germany.
Ralf
R
Schröder
Johannes
J
Schmidt
Stefanie
S
Blättermann
Lucas
L
Peters
Nicole
N
Janssen
Manuel
M
Grundmann
Wiebke
W
Seemann
Dorina
D
Kaufel
Nicole
N
Merten
Christel
C
Drewke
Jesus
J
Gomeza
Graeme
G
Milligan
Klaus
K
Mohr
Evi
E
Kostenis
D000818:Animals; D015374:Biosensing Techniques; D016466:CHO Cells; D006224:Cricetinae; D057809:HEK293 Cells; D006801:Humans; D016571:Neural Networks, Computer; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
D043562:Receptors, G-Protein-Coupled