Bifunctional Calix[4]arene-Coated Gold Nanoparticles for Orthogonal Conjugation

Abstract

Gold nanoparticles (AuNPs) are currently intensively exploited in the biomedical field as they possess interesting chemical and optical properties. Although their synthesis is well-known, their controlled surface modification with defined densities of ligands such as peptides, DNA, or antibodies remains challenging and has generally to be optimized case by case. This is particularly true for applications like in vivo drug delivery that require AuNPs with multiple ligands, for example a targeting ligand and a drug in well-defined proportions. In this context, we aimed to develop a calixarene-modification strategy that would allow the controlled orthogonal conjugation of AuNPs, respectively, via amide bond formation and copper­(I)-catalyzed azide–alkyne cycloaddition (CuAAC). To do this, we synthesized a calix[4]­arene-tetradiazonium salt bearing four PEG chains ended by an alkyne group (C1) and, after optimization of its grafting on 20 nm AuNPs, we demonstrated that CuAAC can be used to conjugate an azide containing dye (N3-cya7.5). It was observed that AuNPs coated with C1 (AuNPs-C1) can be conjugated to approximately 600 N3-cya7.5 that is much higher than the value obtained for AuNPs decorated with traditional thiolated PEG ligands terminated by an alkyne group. The control over the number of molecules conjugated via CuAAC was even possible by incorporating a non-functional calixarene (C2) into the coating layer. We then combined C1 with a calix[4]­arene-tetradiazonium salt bearing four carboxyl groups (C3) that allows conjugation of an amine (NH2-cya7.5) containing dye. The conjugation potential of these bifunctional AuNPs-C1/C3 was quantified by UV–vis spectroscopy: AuNPs decorated with equal amount of C1 and C3 could be conjugated to approximately 350 NH2-dyes and 300 N3-dyes using successively amide bond formation and CuAAC, demonstrating the control over the orthogonal conjugation. Such nanoconstructs could benefit to anyone in the need of a controlled modification of AuNPs with two different molecules via two different chemistries.

Ligation of Gold Nanoparticles with Self-Assembling, Coiled-Coil Peptides

“…Higher LA-PEG-Mal surface fractions lead to higher local concentrations of Cy5 since they are tethered to the AuNP surface, resulting in the formation of dimers and H-aggregates as shown by the increase in absorbance at 603 nm. , The number of CE21Cy5 per AuNP was calculated from the absorbance data using extinction coefficients of 2.71 × 10 5 and 9.2 × 10 8 M –1 cm –1 . When the AuNPs had a 40% surface fraction of LA-PEG-Mal, there were 293 ± 45 CE21Cy5 molecules per AuNP (Figure d). , Assuming a ligand foot-print-area of 0.8 nm 2 per LA-PEG-Mal, we calculated there to be 354 possible binding sites for CE21Cy5. , This indicates an 83 ± 13% coupling efficiency between CE21Cy5 and the LA-PEG-Mal on the AuNP surface. When there was a 0% surface fraction of LA-PEG-Mal, there were 24 ± 2 CE21Cy5 per AuNP demonstrating nondiscriminate adsorption of CE21Cy5 onto the AuNP.…”

Ultra-stable silver nanoplates: efficient and versatile colorimetric reporters for dipstick assays

“…Calixarene-coated Ag or AuNPs have demonstrated a superior stability compared to particles with traditional coatings, allowing their drying and resuspension or their use in biofluids. 21,[25][26][27] Very recently, we have shown that calixarene-coated AgNPs could advantageously replace gold nanoparticles of the same size (i.e. around 20 nm) in a serological dipstick assay for the detection of Anti-SARS-CoV-2 Immunoglobulin G. 22 An improvement by one order of magnitude of the limit of detection was indeed obtained compared to AuNPs due to the better optical properties of silver.…”

Di-Arginine Additives for Dissociation of Gold Nanoparticle Aggregates: A Matrix-Insensitive Approach with Applications in Protease Detection

“…22 While many in the community have shown the reversible assembly of a small number of nanoparticles, this usually requires sophisticated coating of the AuNP-citrate surface with responsive polymers, 23,24 DNA strands, 25,26 or other organic ligands. 27,28 However, we are unaware of work describing the assembly of a large amount of AuNPs−citrate into macroscopic aggregates that can easily be dissociated without the need for prior surface modifications.…”

“…Reversible aggregation of nanoparticles is challenging because, according to the Derjaguin–Landau–Verwey–Overbeek (DLVO) theory, the particles can be trapped in deep energetic minima during the aggregation, thus transforming the aggregates into larger insoluble materials that can be only slightly dissociated by aggressive sonication; their optical properties cannot be recovered . While many in the community have shown the reversible assembly of a small number of nanoparticles, this usually requires sophisticated coating of the AuNP-citrate surface with responsive polymers, , DNA strands, , or other organic ligands. , However, we are unaware of work describing the assembly of a large amount of AuNPs–citrate into macroscopic aggregates that can easily be dissociated without the need for prior surface modifications.…”