N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 | CAS 2107273-30-1

N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 is a Cy5-based NHS-ester conjugation dye designed for water-soluble fluorescent labeling of amine-containing biomolecules and polymers. The PEG spacers improve handling and reduce steric constraints during coupling, enabling robust attachment to proteins, peptides, and other primary amines for fluorescence microscopy, flow cytometry, and fluorescence-based assay development. Cy5 excitation/emission in the far-red region supports low-background imaging workflows and convenient multiplexing with common visible fluorophores.

1. Protein And Peptide Labeling

N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 is widely used to fluorescently label antibodies, enzymes, receptors, and peptide reagents through NHS-ester coupling to lysine residues and N-termini. Researchers in chemical biology and biomolecular imaging workflows rely on the PEG4 spacers to maintain dye accessibility and reduce aggregation during conjugate preparation, supporting consistent staining of purified targets and labeled binding reagents. The resulting Cy5 conjugates are commonly incorporated into binding studies, fluorescence readouts for affinity reagents, and imaging experiments where far-red signal is advantageous for separating labeled targets from autofluorescence.

2. Flow Cytometry Staining

N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 is frequently selected for labeling cell-surface or extracellular targets using amine-reactive coupling to generate fluorescent antibodies or labeling reagents compatible with flow cytometry. The far-red Cy5 channel helps reduce spectral overlap with standard green/orange fluorophores, while the PEG functionality supports practical conjugate handling for staining panels. Typical workflows include preparing Cy5-labeled affinity reagents for staining fixed or permeabilized samples, followed by instrument-based quantification of fluorescence intensity to monitor binding, receptor abundance, or labeling consistency across experimental conditions.

3. Biomaterial Surface Functionalization

N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 is used to introduce far-red fluorescence onto biomaterial surfaces and PEGylated materials by coupling the NHS ester to amine-bearing coatings or functionalized polymers. Materials scientists and biomaterials researchers incorporate the dye into surface-modified hydrogels, polymer films, and functional layers to enable visualization of surface attachment, tracking of material localization, and monitoring of coating uniformity. The PEG spacers help maintain dye presentation away from the surface, supporting reliable fluorescence readout for microscopy-based characterization and fluorescence imaging of engineered interfaces.

4. Fluorescence Imaging Tracers

N-(m-PEG4)-N'-(PEG4-NHS ester)-Cy5 is applied as a fluorescent tracer for microscopy experiments where labeling of amine-functional biomolecules is required to follow distribution, uptake, or localization patterns. Investigators often prepare Cy5 conjugates with carrier proteins, enzymes, or labeled probe constructs to generate far-red imaging reagents that remain compatible with common fluorescence imaging setups. The NHS-ester reactivity supports straightforward conjugate generation from primary-amine-containing components, enabling researchers to tailor labeling density for imaging workflows while leveraging the Cy5 spectral window for reduced background in complex biological samples.