AF647 NHS ester | CAS 407627-60-5

AF647 NHS ester is a reactive Alexa Fluor 647 dye derivative designed for efficient covalent labeling of primary amine-containing biomolecules. As an NHS ester, it readily forms stable amide bonds with lysine residues on proteins and peptides, enabling bright far-red fluorescence for microscopy, flow cytometry, and fluorescence-based assay development. The far-red emission of Alexa Fluor 647 helps reduce background from autofluorescence and supports multiplex-friendly imaging workflows when paired with spectrally distinct fluorophores.

1. Protein And Antibody Labeling

AF647 NHS ester is widely used by antibody and protein engineering groups to generate fluorescently labeled antibodies, immunoreagents, and protein conjugates for binding studies and imaging workflows. Researchers typically use it to label lysine-rich targets such as IgG, affinity reagents, enzymes, and receptor-binding proteins, producing far-red conjugates for fluorescence microscopy and quantitative fluorescence assays. The NHS ester reactivity supports straightforward preparation of dye-protein conjugates used in immunostaining panels, binding kinetics studies, and assay reagent development where stable amide linkage is required.

2. Flow Cytometry Conjugates

AF647 NHS ester is commonly employed to prepare far-red flow cytometry dyes for cell-surface and intracellular labeling workflows that rely on fluorescent antibody conjugates. Cytometry users conjugate the dye to antibodies or other amine-bearing targeting ligands to create reagents for multiparameter panels, benefiting from the Alexa Fluor 647 spectral region to help separate signals from common cellular autofluorescence. These conjugates are frequently used for receptor expression analysis, phenotyping workflows, and fluorescence-based gating strategies in research and industrial cell analytics settings.

3. Fluorescence Microscopy Staining

AF647 NHS ester supports fluorescent staining and labeling for fluorescence microscopy when robust covalent attachment to amine-containing biomolecules is required. Imaging teams use dye-protein conjugates to visualize targets such as extracellular markers, labeled affinity probes, and protein components in fixed-cell or fixed-tissue workflows, where stable labeling helps maintain signal during washing and imaging. The far-red fluorescence of Alexa Fluor 647 is particularly useful for multicolor microscopy experiments that require spectral separation and reduced background from endogenous fluorescence.

4. Fluorescent Assay Reagent Development

AF647 NHS ester is used in fluorescence assay development to create labeled detection reagents, including dye-tagged proteins and affinity reagents used as readout components in binding assays and fluorescence-based screening formats. Assay developers conjugate the dye to amine-bearing components to produce stable fluorescent probes for endpoint measurements and imaging-compatible assays, often leveraging the far-red emission to improve robustness against background. In molecular biology and chemical biology workflows, these labeled reagents are used to track binding events, quantify interactions, and support fluorescence readouts in research-grade assay systems.