研究目的
To develop and characterize a fluorescent biosensor that can detect the R248Q mutant of p53, which is prone to aggregation, both in vitro and in living cells.
研究成果
The developed fluorescent peptide biosensor can detect nanomolar concentrations and discriminate between R248Q and WT p53 in lung cancer cell extracts and in living cells, providing a useful tool for ex vivo diagnostics and potential drug discovery applications.
研究不足
The efficiency of internalization of the cell-penetrating peptide/biosensor complexes may differ between cell lines, requiring normalization of biosensor response to its intracellular levels if different cell lines are being studied.
1：Experimental Design and Method Selection:
The biosensor was engineered through conjugation of an environmentally-sensitive probe onto a peptide derived from the primary aggregation segment of p
2：Sample Selection and Data Sources:
The biosensor was tested in vitro and in living cells, specifically in PC9 lung cancer cell line expressing the R248Q mutant.
3：List of Experimental Equipment and Materials:
Fluorescent probes (FITC, TP2Rho), recombinant GST-tagged p53 proteins, cell lines (A549, PC9, H1299), and fluorescence microscopy equipment.
4：Experimental Procedures and Operational Workflow:
The biosensor was characterized by fluorescence titration assays, thermodenaturation experiments, and fluorescence imaging in living cells.
5：Data Analysis Methods:
Fluorescence emission was quantified, and dissociation constants were determined using curve fitting software.
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