N-(DBCO-PEG4)-N-Biotin-PEG4-NHS | CAS 2353409-72-8

N-(DBCO-PEG4)-N-Biotin-PEG4-NHS is a biotinylated, NHS-activated DBCO (dibenzocyclooctyne) click chemistry reagent designed for strain-promoted azide–alkyne cycloaddition (SPAAC) workflows. The molecule combines an NHS ester for stable amide coupling with a PEG spacer architecture that improves solubility and reduces steric effects, while the DBCO handle enables rapid, catalyst-free conjugation to azide-bearing partners. This reagent is commonly used to generate azide-reactive biotinylated probes, immobilization reagents, and multivalent labeling constructs for biochemical assays, affinity capture, and molecular imaging tool development.

1. Biotin-Based Affinity Labeling

N-(DBCO-PEG4)-N-Biotin-PEG4-NHS is widely used to introduce both a biotin tag and a DBCO click handle onto proteins, peptides, and other NHS-reactive targets, enabling downstream affinity capture and orthogonal conjugation. Researchers often use the biotin functionality to streamline pull-down workflows with streptavidin or neutravidin surfaces, while the DBCO moiety provides a second handle for SPAAC coupling to azide-modified dyes, polymers, or biomolecule conjugates. This dual functionality supports modular probe assembly, where the biotin-containing intermediate can be clicked to azide reporters at a later stage to build multicomponent labeling reagents for research-grade assays and imaging experiments.

2. Surface Immobilization For Assays

N-(DBCO-PEG4)-N-Biotin-PEG4-NHS supports immobilization strategies that combine covalent attachment via the NHS ester with subsequent click-based loading of azide-functional components. In assay development and biosensor prototyping, the NHS ester is used to decorate amine-containing surfaces or biomolecular coatings, after which azide-bearing detection elements can be coupled through SPAAC to create defined, spatially controlled binding layers. The PEG spacers help maintain accessibility of both the biotin and DBCO-derived reactive sites, which is particularly useful when constructing complex assay surfaces such as streptavidin-coated platforms, microarray formats, or bead-based reagent systems requiring consistent conjugate presentation.

3. Multivalent Probe And Imaging Reagents

N-(DBCO-PEG4)-N-Biotin-PEG4-NHS is commonly incorporated into probe pipelines for fluorescence labeling, enrichment, and imaging reagent construction where multivalency improves signal generation and workflow efficiency. The reagent’s biotin group enables convenient enrichment or standardized capture on streptavidin scaffolds, while the DBCO handle allows rapid coupling to azide-functional imaging reporters, such as azide-modified fluorophores, polymeric tags, or imaging-compatible scaffolds. This design is especially valuable for building stepwise imaging conjugates, since the NHS coupling step can be performed first to functionalize a targeting or scaffold molecule, followed by click attachment of the final imaging component under mild, catalyst-free conditions.

4. Click-Ready Biomolecule Conjugates

N-(DBCO-PEG4)-N-Biotin-PEG4-NHS is used to generate click-ready biomolecule conjugates that can be further assembled into complex reagent libraries. Typical workflows involve NHS-mediated functionalization of proteins, antibodies, antibody fragments, or enzyme conjugates bearing accessible primary amines, producing intermediates that retain a DBCO handle for subsequent SPAAC coupling to azide-bearing partners. The PEG4 spacing and biotin tag support robust handling in aqueous buffers and compatibility with affinity-based workflows, making the reagent a practical choice for constructing modular research tools such as biotin-tagged enzyme conjugates, affinity-enriched labeling reagents, and multicomponent biochemical probes used in chemical biology and molecular imaging method development.