Fitc-ASTD-FMK integrates a FITC fluorophore with a tetrapeptide sequence and a fluoromethyl ketone reactive group. The construct enables visualization of protease engagement and covalent active-site labeling. Researchers use it to examine enzyme specificity and catalytic mechanism. Its fluorescent signature facilitates real-time monitoring in biochemical assays.
CAT No: HB00025
Fitc-ASTD-FMK is a synthetic peptide inhibitor distinguished by the incorporation of a fluorescein isothiocyanate (FITC) fluorescent label and a fluoromethyl ketone (FMK) reactive group. As a biochemically engineered peptide, it is designed to irreversibly bind and inhibit specific cysteine proteases, particularly within the caspase family. The FITC label enables direct visualization and tracking in cellular and biochemical assays, while the FMK moiety confers potent and selective inhibition by covalently modifying the active site of target enzymes. Fitc-ASTD-FMK is widely utilized in apoptosis research, protease activity profiling, and mechanistic studies requiring both functional inhibition and real-time detection, making it a valuable tool for researchers investigating proteolytic pathways and cell death mechanisms.
Protease Activity Profiling: As a mechanism-based inhibitor, Fitc-ASTD-FMK is highly effective for profiling the activity of caspases and related cysteine proteases in various biological samples. Upon entry into cells or lysates, it covalently binds to the active site cysteine of target enzymes, resulting in irreversible inhibition. The attached FITC fluorophore allows for sensitive detection and quantification of protease-bound inhibitor by fluorescence measurement, facilitating the assessment of protease activation status under different experimental conditions. This property is particularly advantageous for elucidating protease dynamics in apoptosis, inflammation, and other regulated cell death processes.
Apoptosis Detection: The compound's dual functionality as both an inhibitor and a fluorescent probe makes it especially useful in apoptosis research. By selectively binding to active caspases, Fitc-ASTD-FMK enables the identification of cells undergoing programmed cell death. The resulting fluorescent signal can be detected by flow cytometry, fluorescence microscopy, or plate-based assays, providing a direct readout of caspase activation at the single-cell or population level. This application supports the investigation of apoptotic pathways, drug screening for apoptosis modulators, and the validation of cell death mechanisms in diverse cellular models.
Live-Cell Imaging: The FITC label incorporated into the peptide backbone permits real-time visualization of protease activity in living cells. Researchers can apply Fitc-ASTD-FMK to cultured cells and monitor the spatial and temporal dynamics of caspase activation using fluorescence microscopy. This capability enables detailed studies of cell death progression, subcellular localization of active proteases, and the impact of experimental treatments on apoptotic signaling. The use of live-cell imaging with this probe enhances the temporal resolution of apoptosis assays and supports high-content screening applications.
Enzyme Kinetics and Inhibitor Evaluation: Fitc-ASTD-FMK serves as a valuable reagent for characterizing enzyme kinetics and evaluating the potency and selectivity of novel protease inhibitors. By acting as a well-defined, irreversible inhibitor with a fluorescent tag, it allows for the direct measurement of time-dependent inactivation rates and competition with other inhibitors. This information is crucial for the rational design and optimization of new inhibitory compounds targeting caspases or related proteases, supporting both basic research and early-stage drug discovery efforts.
Cell-Based Assay Development: The compound is frequently employed in the development and validation of cell-based assays for caspase activity and apoptosis. Its cell-permeable structure, combined with the specific and irreversible nature of FMK-mediated inhibition, ensures robust signal generation and minimal background. The FITC fluorescence readout enables multiplexing with other fluorescent probes and compatibility with automated platforms, streamlining assay workflows in high-throughput screening and functional genomics studies. By providing a reliable and sensitive tool for monitoring protease activity in cellular contexts, Fitc-ASTD-FMK supports the advancement of research in cell biology, toxicology, and molecular pharmacology.
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