Rapid chemoselective bioconjugation through oxidative coupling of anilines and aminophenols.

A highly efficient protein bioconjugation method is described involving addition of anilines to o-aminophenols in the presence of sodium periodate. The reaction takes place in aqueous buffer at pH 6.5 and can reach high conversion in 2-5 min. The major product was characterized using X-ray crystallography, which revealed that an unprecedented oxidative ring contraction occurs after the coupling step. The compatibility of the reaction with protein substrates has been demonstrated through attachment of small molecules, polymer chains, and peptides to p-aminophenylalanine residues introduced into viral capsids through amber stop codon suppression. Coupling of anilines to o-aminophenol groups derived from tyrosine residues is also described. The compatibility of this method with thiol modification chemistry is shown through attachment of a near-IR fluorescent chromophore to cysteine residues inside the viral capsid shells, followed by attachment of integrin-targeting RGD peptides to anilines on the exterior surface.

Controlled synthesis and single-particle imaging of bright, sub-10 nm lanthanide-doped upconverting nanocrystals.

Phosphorescent nanocrystals that upconvert near-infrared light to emit at higher energies in the visible have shown promise as photostable, nonblinking, and background-free probes for biological imaging. However, synthetic control over upconverting nanocrystal size has been difficult, particularly for the brightest system, Yb(3+)- and Er(3+)-doped ß-phase NaYF(4), for which there have been no reports of methods capable of producing sub-10 nm nanocrystals. Here we describe conditions for the controlled synthesis of protein-sized ß-phase NaYF(4): 20% Yb(3+), 2% Er(3+) nanocrystals, from 4.5 to 15 nm in diameter. The size of the nanocrystals was modulated by varying the concentration of basic surfactants, Y(3+):F(-) ratio, and reaction temperature, variables that also affected their crystalline phase. Increased reaction times favor formation of the desired ß-phase nanocrystals while having only a modest effect on nanocrystal size. Core/shell ß-phase NaYF(4): 20% Yb(3+), 2% Er(3+)/NaYF(4) nanoparticles less than 10 nm in total diameter exhibit higher luminescence quantum yields than comparable >25 nm diameter core nanoparticles. Single-particle imaging of 9 nm core/shell nanoparticles also demonstrates that they exhibit no measurable photobleaching or blinking. These results establish that small lanthanide-doped upconverting nanoparticles can be synthesized without sacrificing brightness or stability, and these sub-10 nm nanoparticles are ideally suited for single-particle imaging.