Alexa Fluor 680 NHS ester | CAS 407628-15-3

Alexa Fluor 680 NHS ester is a reactive N-hydroxysuccinimide (NHS) ester dye designed for covalent fluorescent labeling of primary amines on proteins, peptides, and other amine-containing biomolecules. With excitation and emission well matched to far-red fluorescence instrumentation, it is commonly used to generate bright, stable conjugates for imaging, quantitation, and multiplex-compatible workflows. The NHS ester chemistry enables straightforward preparation of labeled reagents for downstream microscopy, gel-based analysis, and flow cytometry panels where far-red detection is advantageous.

1. Protein Conjugation Labeling

Alexa Fluor 680 NHS ester is frequently used by protein engineering and antibody labeling teams to prepare fluorescently tagged proteins for imaging and analytical workflows. Researchers typically target lysine residues and N-terminal amines to generate amine-reactive conjugates for Western blotting, fluorescence gel imaging, and quantitative binding experiments. Far-red readout helps reduce interference from shorter-wavelength fluorophores in multicolor studies, making the dye a practical choice when building labeled secondary reagents, assay standards, or fluorescence imaging probes from purified proteins.

2. Flow Cytometry Panel Staining

Alexa Fluor 680 NHS ester supports fluorescent labeling strategies used to build far-red flow cytometry reagents for cell-surface or intracellular targets when the labeling partner provides accessible primary amines. In practice, it is used to tag antibodies, antibody fragments, or other amine-containing binders prior to staining, enabling detection on instruments equipped for far-red channels. This labeling approach is especially useful for multiplex experiments where spectral separation from common green and red dyes is required to maintain clear population discrimination and robust gating across complex sample sets.

3. Fluorescence Microscopy Imaging

Alexa Fluor 680 NHS ester is widely applied in fluorescence microscopy workflows to generate labeled biomolecules for cellular imaging, including conjugates used for immunofluorescence and fluorescence localization studies. By covalently attaching the dye to amine-containing targeting reagents, researchers can visualize binding patterns with far-red excitation/emission compatible with modern microscope filter sets and laser lines. The NHS-ester labeling format is also used to create fluorescent standards or labeling reagents for co-localization experiments, benefiting studies that require reduced background from autofluorescence and improved spectral separation.

4. Bioconjugate Standards And Tracers

Alexa Fluor 680 NHS ester is commonly used in reagent development to create fluorescent tracers and calibration materials for quantitative fluorescence assays and imaging-based readouts. Teams in chemical biology and materials research use amine-reactive labeling to prepare dye-tagged proteins, peptides, or other amine-containing components that serve as internal standards, reference conjugates, or normalization controls. Far-red fluorescence compatibility makes these standards useful across gel imaging, plate-based fluorescence measurements, and instrument platforms that support Alexa Fluor 680 detection.