Intracellular Pathogen Detection Based on Dual-Recognition Units Constructed Fluorescence Resonance Energy Transfer Nanoprobe

Abstract

The intracellular invasion and survival of a pathogen like Staphylococcus aureus (S. aureus) within host cells enable them to resist antibiotic treatment and colonize long-term in the host, which leads to a series of clinical issues. Rapid and specific detection of intracellular bacteria is important in diagnosis of infection and guiding antibiotic administration. Herein, this work reports a simple one-step fluorescence resonance energy transfer (FRET) platform-based strategy to achieve specific and rapid detection of S. aureus in specimens of phagocytic cells. The aptamer modified quantum dots (Aptamer-QDs) and antibiotic molecule of Teicoplanin functionalized-gold nanoparticles (Teico-AuNPs) dual-recognition units to S. aureus are employed as energy donor and acceptor, respectively. Based on the "off" to "on" signal readout mode, when in the presence of target S. aureus, the donor and acceptor are close to each other and bring high FRET efficiency, which is suitable for analysis of intracellular S. aureus. After it was incubated with the sample for 2 h, the as-prepared FRET sensor showed selectivity to the target S. aureus, and the changed fluorescence signal shows an obvious variation with increasing concentration of S. aureus in pure buffer. When the FRET strategy was further applied to assay intracellular S. aureus, there was an obvious fluorescence signal change obtained both by spectrum analysis and visual fluorescence microscope observation when the average number of S. aureus in one host cell (N S. aureus/cell ) was as low as 1, which can be attributed to the high fluorescence quenching efficiency of about 41.3%. It could be envisioned that this FRET nanoprobe with high fluorescence quenching efficiency may provide a simple approach for the facile, selective, and rapid diagnosis of an intracellular bacterial infection.

Principles and Recent Advances in Biosensors for Pathogens Detection

“…Over the last few decades, organic dyes have played a significant role in fluorescence-based biosensors and, therefore, the downsides of employing traditional dyes have been resolved by the utilization of quantum dots as fluorophores. [184] In addition to quantum dots, nanomaterials such as GaAs, InAs, PbTe, PbSe, InP, HgTe, PbS, CdTe, and CdSe have also been used as popular fluorophores for the detection of deadly pathogens. On the other hand, the indirect biosensing approach uses a sandwich-type detection strategy to recognize foodborne and waterborne pathogens in contaminated food and polluted water samples, respectively.…”

Ex vivo and in vivo fluorescence detection and imaging of adenosine triphosphate

“…As such, searching for FRET acceptors with prominent quenching efficiency against fluorescent dyes is pivotal for the construction of ultrasensitive fluorescent ATP probes. To date, the quenching efficiency of most reported FRET acceptors were less than 95% [ 22 , 23 ]. Besides, fluorescent probes suitable for simultaneously sensing ATP ex vivo and in vivo are still few.…”

Ex Vivo and In Vivo Fluorescence Detection and Imaging of Adenosine Triphosphate

“…As such, searching for FRET acceptors with prominent quenching e ciency against uorescent dyes is pivotal for the construction of ultrasensitive uorescent ATP probes. To date, the quenching e ciency of most reported FRET acceptors were less than 95% 22,23 . Besides, uorescent probes suitable for simultaneously sensing ATP ex vivo and in vivo are still few.…”

“…Most uorescent probes for ATP detection were based on the uorescence resonance energy transfer (FRET) sensing strategy. FRET is a nonradiative energy transfer process between donor chromophore and acceptor chromophore (quencher) [21][22][23] . When using the same donor in the FRET system, the acceptor featuring superior quenching e ciency would in principle lead to higher e ciency of the energy transfer.…”