期刊
SENSORS
卷 16, 期 8, 页码 -出版社
MDPI
DOI: 10.3390/s16081312
关键词
FRET; FLIM; AMPK; spheroid; 2-photon; biosensor; TCSPC; 3D culture
资金
- UK Biotechnology and Biological Sciences Research Council [BBSRC BB/M006786/1]
- MRC Clinical Sciences Centre
- Imperial College London
- Institute of Chemical Biology EPSRC
- BBSRC [BB/M006786/1] Funding Source: UKRI
- MRC [MC_U120027537] Funding Source: UKRI
- Biotechnology and Biological Sciences Research Council [BB/M006786/1] Funding Source: researchfish
- Medical Research Council [MC_U120027537] Funding Source: researchfish
- Medical Research Foundation [MRF-023-0002-S-CHENN] Funding Source: researchfish
We describe an approach to non-invasively map spatiotemporal biochemical and physiological changes in 3D cell culture using Forster Resonance Energy Transfer (FRET) biosensors expressed in tumour spheroids. In particular, we present an improved Adenosine Monophosphate (AMP) Activated Protein Kinase (AMPK) FRET biosensor, mTurquoise2 AMPK Activity Reporter (T2AMPKAR), for fluorescence lifetime imaging (FLIM) readouts that we have evaluated in 2D and 3D cultures. Our results in 2D cell culture indicate that replacing the FRET donor, enhanced Cyan Fluorescent Protein (ECFP), in the original FRET biosensor, AMPK activity reporter (AMPKAR), with mTurquoise2 (mTq2FP), increases the dynamic range of the response to activation of AMPK, as demonstrated using the direct AMPK activator, 991. We demonstrated 3D FLIM of this T2AMPKAR FRET biosensor expressed in tumour spheroids using two-photon excitation.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据