Monitoring individual cell-signaling activity using combined metal-clad waveguide and surface-enhanced fluorescence imaging

摘要: Evanescent field based biosensing systems such as surface plasmon resonance (SPR), diffraction gratings, or metal-clad waveguides (MCWGs) are powerful tools for label-free real-time monitoring of signaling activity living cells exposed to hormones, pharmacological agents, and toxins. In particular, MCWG-based imaging is well suited studying relatively thick objects due its greater depth penetration into the sensing medium, compared SPR. Label-free methods, however, provide only indirect measurements in that measured signal arises from local changes material properties rather than specific biomolecular targets. case cells, situation especially complex may result a combination very diverse sources: morphological changes, intra-cellular reorganization, cascaded molecular events, protein expression etc. Consequently, deconvolving contributions sources particular cell response profile can be challenging. following, we present platform combines two distinct modalities, namely MCWG label-based enhanced fluorescence (SEF), designed facilitate identification underlying structural images. We demonstrate bimodal capabilities this experiments visualize actin cytoskeleton organization vascular smooth muscle cells. then monitored HEK293 expressing Angiotensin 1 receptor (AT1R), when stimulated by agonist II (AngII). The analysis simultaneous obtained intracellular calcium resulting form AT1R activation, SEF, allows relating features markers activation. Our results show levels normally observed following activation not required initial burst cellular but indicates involving kinase ROCK.