N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 | CAS 2107273-68-5

N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 is a Cy5-based fluorescent bioconjugation reagent featuring a PEG spacer architecture that supports aqueous labeling and flexible presentation of the dye and the biotin handle. The biotin functionality enables strong avidin/streptavidin capture for downstream immobilization, enrichment, and patterned assembly, while the Cy5 fluorophore supports fluorescence imaging and detection in the red spectral window. This combination is widely used in workflows that require both fluorescent readout and affinity-based targeting or surface attachment.

1. Avidin Streptavidin Pull-Down

N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 is used to create fluorescent affinity probes for capturing biotin-binding proteins, immobilized streptavidin surfaces, or biotinylated targets in analytical workflows. Researchers commonly incorporate it into pull-down or enrichment steps where the biotin handle provides robust capture and the Cy5 label enables direct visualization or quantification of recovered material by fluorescence imaging or plate-based fluorescence readout. The PEG linkers help reduce steric hindrance, improving practical labeling density and signal uniformity across conjugate preparations.

2. Fluorescence Imaging Of Biotinylated Surfaces

N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 supports red fluorescence mapping of biomaterials and engineered surfaces that present biotin for streptavidin/avidin binding. In materials science and chemical biology, it is frequently applied to verify surface functionalization, assess spatial coverage, and monitor conjugate attachment on microarrays, hydrogels, and solid supports. The flexible PEG spacers help the Cy5 dye remain accessible to the optical field, which is particularly useful for microscopy-based localization studies and for comparing labeling conditions across surface batches.

3. Flow Cytometry Labeling With Avidin Capture

N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 is employed in flow cytometry workflows where a biotin-dependent avidin/streptavidin capture step is used to introduce a Cy5 fluorescence tag onto cells, particles, or biomolecular complexes. Typical use cases include labeling biotinylated cell-surface reagents, tagging streptavidin-coated beads, or generating fluorescent standards for gating and assay development. The PEGylated construct helps maintain conjugate dispersibility and reduces nonspecific aggregation that can complicate cytometric readouts, while Cy5 provides a convenient red-channel signal for multiplexing with other fluorophores when spectral separation is available.

4. Fluorescent Reporter In Binding Assays

N-(m-PEG4)-N'-(biotin-PEG3)-Cy5 is used as a fluorescent reporter in affinity-based binding and competition assays that rely on biotin-avidin interactions. In assay development settings, the reagent enables monitoring of binding events through Cy5 fluorescence while the biotin handle provides a defined, high-affinity capture mechanism to anchor the fluorescent signal to an avidin/streptavidin-coated format. This approach is frequently applied during reagent screening, surface chemistry optimization, and protocol qualification for systems where a consistent fluorescent readout is required alongside an affinity-defined capture step.