研究目的
To explore the application of optogenetic tools in the microscopy of cardiac excitation-contraction coupling, focusing on genetically encoded biosensors and their use in observing and manipulating cellular functions.
研究成果
The study demonstrates the potential of optogenetic tools in cardiac research, highlighting the advantages of genetically encoded biosensors for observing and manipulating cellular functions. Future research should focus on improving the spectral range, fluorescence yield, and kinetics of these tools.
研究不足
The study is limited by the spectral properties of the biosensors, their biocompatibility, and the need for genetic manipulation of the cells. The temporal kinetics of some biosensors may not be fast enough to capture rapid cellular events.
1:Experimental Design and Method Selection:
The study employs genetically encoded biosensors based on F?rster Resonance Energy Transfer (FRET) and circular permutated ?uorescent proteins (CPFP) for observing cellular functions. Channelrhodopsins are used for manipulating cellular functions.
2:Sample Selection and Data Sources:
Adult cardiac myocytes and stem cell-derived cardiomyocytes are used as samples. Data is acquired through fluorescence microscopy.
3:List of Experimental Equipment and Materials:
Includes fluorescence microscopes, genetically encoded biosensors, and channelrhodopsins.
4:Experimental Procedures and Operational Workflow:
Involves the expression of biosensors in cells, illumination with appropriate wavelengths for activation, and observation of fluorescence changes.
5:Data Analysis Methods:
Quantitative FRET measurements are performed to analyze the data, including corrections for donor bleed-through and excitation crosstalk.
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