研究目的
To develop a highly efficient genetically encoded mercury FRET sensor (MerFS) for measuring the cellular dynamics of Hg2+ at trace level in real time.
研究成果
The developed genetically encoded FRET-based nanosensor MerFS-51 is a highly efficient tool for the non-invasive monitoring of Hg2+ ions in living cells, offering high sensitivity and selectivity. It has potential applications in understanding Hg2+ homeostasis and in the diagnosis of metal-induced diseases.
研究不足
The study focuses on the detection of Hg2+ ions and does not address the detection of other heavy metals or the transformation of Hg2+ into other forms such as methylmercury in detail.
1：Experimental Design and Method Selection:
The study involved the construction of a genetically encoded FRET-based nanosensor using the periplasmic mercury-binding protein MerP sandwiched between enhanced cyan fluorescent protein (ECFP) and venus.
2：Sample Selection and Data Sources:
The sensor was tested in Escherichia coli (E. coli), yeast, and human embryonic kidney (HEK)-293T cells.
3：List of Experimental Equipment and Materials:
Equipment included a monochromator microplate reader (Biotek, USA), confocal microscope (Leica, Wetzlar, Germany), and various buffers and reagents for protein expression and purification.
4：Experimental Procedures and Operational Workflow:
The sensor protein was expressed, purified, and characterized for pH stability, metal specificity, and binding affinity. Live cell imaging was performed to monitor Hg2+ uptake.
5：Data Analysis Methods:
Binding constants (Kd) were calculated using GraphPad Prism software by fitting the ligand titration curve in a binding isotherm equation.
独家科研数据包，助您复现前沿成果，加速创新突破
获取完整内容
