Modular fluorescent nanoparticle DNA probes for detection of peptides and proteins.

Autor: Stawicki CM; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Rinker TE; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA. torrir@nautilus.bio., Burns M; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Tonapi SS; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Galimidi RP; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Anumala D; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Robinson JK; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Klein JS; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA., Mallick P; Nautilus Biotechnology, 201 Industrial Rd #310, San Carlos, CA, 94070, USA.
Jazyk: angličtina
Zdroj: Scientific reports [Sci Rep] 2021 Oct 07; Vol. 11 (1), pp. 19921. Date of Electronic Publication: 2021 Oct 07.
DOI: 10.1038/s41598-021-99084-4
Abstrakt: Fluorescently labeled antibody and aptamer probes are used in biological studies to characterize binding interactions, measure concentrations of analytes, and sort cells. Fluorescent nanoparticle labels offer an excellent alternative to standard fluorescent labeling strategies due to their enhanced brightness, stability and multivalency; however, challenges in functionalization and characterization have impeded their use. This work introduces a straightforward approach for preparation of fluorescent nanoparticle probes using commercially available reagents and common laboratory equipment. Fluorescent polystyrene nanoparticles, Thermo Fisher Scientific FluoSpheres, were used in these proof-of-principle studies. Particle passivation was achieved by covalent attachment of amine-PEG-azide to carboxylated particles, neutralizing the surface charge from - 43 to - 15 mV. A conjugation-annealing handle and DNA aptamer probe were attached to the azide-PEG nanoparticle surface either through reaction of pre-annealed handle and probe or through a stepwise reaction of the nanoparticles with the handle followed by aptamer annealing. Nanoparticles functionalized with DNA aptamers targeting histidine tags and VEGF protein had high affinity (EC 50 s ranging from 3 to 12 nM) and specificity, and were more stable than conventional labels. This protocol for preparation of nanoparticle probes relies solely on commercially available reagents and common equipment, breaking down the barriers to use nanoparticles in biological experiments.
(© 2021. The Author(s).)
Databáze: MEDLINE
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