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Zusammenfassung:
We present a fluorescence-based assay for the characterization of functionalized gold nanoparticles (AuNPs) capped with a self-assembled monolayer of mixed thiols derived from poly(ethylene glycol) (PEG). These water-soluble AuNPs carry primary amino groups at the solvent-exposed interface, which can be used for further conjugation of biologically active molecules. The reported assay allows quantification of the average number of functionalizable amino groups per particle (NNH2) with a relative uncertainty below or equal to ±14% (95% confidence interval), thus providing essential information for the successive derivatization of the AuNPs. Here, a fluorescently labeled derivative of peptide-neurotoxin conantokin-G was coupled to the amino groups of the particle ligand shell via a flexible linker. We quantitatively determined the average number of peptides per particle (Npept) and the yield of the two-step conjugation strategy. AuNPs carrying 50−70 copies of the peptide were obtained. In addition, we have gained insights into the deterioration of the self-assembled monolayer due to thiol desorption with time. Under ordinary storage conditions in solution and at room temperature, a decrease in NNH2 between 48% and 75% could be observed at the end of the period of investigation (42−56 days). Slow desorption of the conjugated peptides upon storage was also observed and quantified (∼25% in 14 days).
