Abstract
Microscopic viscosity is a critical parameter in cellular processes such as diffusion-controlled reactions and the correct folding of nascent proteins. However, a lack of a suitable technology has hampered the analysis of this fundamental parameter within the organelles of living cells. We have established a new technique called Rotor-based organelle viscosity imaging (ROVI). In our technique, environment-sensitive fluorescent dyes termed ‘molecular rotors’ are directed via active targeting to specific intracellular compartments of live cells for the real time dynamic measurement of microscopic viscosity. Molecular rotors are small synthetic fluorophores whose photophysical properties strongly depend upon the viscosity and molecular crowding of their immediate environment. By measuring the fluorescence lifetime of molecular rotors via time-correlated single photon counting fluorescence lifetime imaging (TCSPC FLIM) it is possible to quantitatively determine the microviscosity of the local environment. Using ROVI, we were able to measure and compare the microviscosity in multiple cellular compartments, namely the cytosol, nucleus, ER and mitochondria. Microviscosity was measured both basally and during manipulations that alter organelle function. We report significant variations of microviscosity between the studied organelles. In addition to providing measurements of microviscosity in multiple compartments of the cell with a single ‘molecular rotor’ probe, we also report striking variation of microviscosity between and within individual matrices of mitochondria, the cell's energy repository. Furthermore, we were able to detect changes of ER microviscosity upon drug administration. The ability to measure the viscoelastic response of the ER upon stress is extremely important and highly pertinent to health and disease, as well as being novel fundamental science. We believe that ROVI offers novel avenues to study cell biology and can aid to monitoring the progression and treatment of diseases.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.