Ultrasensitive detection of Staphylococcus aureus using a non-fluorescent cDNA-grafted dark BBQ®-650 chromophore integrated hydrophilic upconversion nanoparticles/aptamer system.

Abstract

A highly structured fluorometric bioassay has been proposed for screening Staphylococcus aureus (S. aureus). The study exploits (i) the spectral attributes of the hexagonal NaYF4:Yb,Er upconversion nanoparticle (UCNP)-coated 3-aminopropyl)triethoxysilane; (ii) the intrinsic non-fluorescent quenching features of the highly stable dark blackberry (BBQ®-650) receptor; (iii) the aptamer (Apt-) biorecognition and binding affinity, and (iv) the complementary DNA hybridizer-linkage efficacy. The principle relied on the excited state energy transfer between the donor Apt-labeled NH2-UCNPs at the 3' end, and cDNA-grafted BBQ®-650 at the 5' end, as the effective receptors. The donor moieties in proximity (< 10.0nm) trigger hybridization with the cDNA-grafted dark BBQ®-650, as the receptors of energy from the 2F5/2 level of Yb3+ ions to initiate the Förster resonance energy transfer pathway. This was confirmed by the decline in the excited-state lifetimes from 223.52μs (τ1) to 179.26μs (τ2). The existence of the target S. aureus in the bioassay attracts the Apt- resulting in the detachment of the acceptor, and disintegration of the complex configuration via conformation reversal. The re-activated fluorescence monitored at λex/em = 980/652nm, as a function of the logarithmic concentration of S. aureus (42 to 4.2 × 108CFUmL-1), yielded an ultra-low detection response of 2.0CFUmL-1. The bioassay screening of S. aureus in real samples revealed satisfactory recoveries (92.44-107.82%) and validation results (p > 0.05). Hence, the comprehensive Apt-labeled NH2-UCNPs-cDNA-grafted dark BBQ®-650 bioassay offered fastand precise S. aureus screening in food and environmental settings.