Multiplexed Molecular Tension Sensor Measurements Using PIE-FLIM.
Methods Mol Biol
; 2600: 221-237, 2023.
Article
em En
| MEDLINE
| ID: mdl-36587101
ABSTRACT
Genetically encoded Förster Resonance Energy Transfer (FRET)-based tension sensors were developed to enable the quantification of piconewton (pN)-scale forces that act across distinct proteins in living cells and organisms. An important extension of this technology is the multiplexing of tension sensors to monitor several independent FRET probes in parallel. Here we describe how pulsed interleaved excitation (PIE)-fluorescence lifetime imaging microscopy (FLIM) can be implemented to enable the analysis of two co-expressed tension sensor constructs. Our protocol covers all essential steps from biosensor expression and live cell PIE image acquisition to lifetime calculations.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Técnicas Biossensoriais
/
Transferência Ressonante de Energia de Fluorescência
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article