研究目的
Investigating the use of fluorescently labeled small molecule affinity ligands for imaging Tie2 in endothelial signaling and angiogenesis.
研究成果
The Reb-TMR derivative effectively labels Tie2 in vitro and in vivo, offering a promising tool for studying Tie2's role in disease. Further optimization and development of companion imaging agents could enhance their utility in clinical settings.
研究不足
The Reb-SiR analog showed weak staining and high background fluorescence, indicating potential issues with cellular environment sensitivity or binding kinetics. The study focused primarily on Reb-TMR, leaving the optimization of Reb-650 for future work.
1：Experimental Design and Method Selection:
Synthesis of three fluorescent derivatives of rebastinib based on crystallography structures. Evaluation in in vitro and in vivo assays.
2：Sample Selection and Data Sources:
HUVEC and HEK293T cells for in vitro assays; Tie2-GFP mice for in vivo imaging.
3：List of Experimental Equipment and Materials:
Operetta High Content Screening System, IXM MetaExpress, Olympus BX63 microscope, Fugene 6 transfection kit, Corning 3764 384-well plates.
4：Experimental Procedures and Operational Workflow:
Treatment of cells with fluorescent derivatives, competition assays, transfection of Tie2 plasmid, intravital imaging in mice.
5：Data Analysis Methods:
Image analysis using Columbus server and ImageJ FIJI, fluorescent polarization binding assay.
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