Up-converting and down-converting nanoparticle-based aptasensor model for multiplex detection of foodborne pathogens

Advances in the selection of synthetic affinity agents, called aptamers, have provided us with the opportunity to develop highly efficient and specific biosensors. Coupling of aptamers with fluorescent nanoparticles have led... [ view full abstract ]

Advances in the selection of synthetic affinity agents, called aptamers, have provided us with the opportunity to develop highly efficient and specific biosensors. Coupling of aptamers with fluorescent nanoparticles have led to the rapid detection of various molecules from ions to disease biomarkers (1). However, only one type of nanoparticle has been used in these multiplexed assays. Herein, we report a multiplex sensing method based on the unique excitation profiles of two down-converting quantum dots (QD) and two up-converting nanoparticles (UCNP) that are coupled with the target-specific DNA aptamers. Our group recently reported the dual-excitation strategy for detection of Salmonella typhimurium and Staphylococcus aureus (2). In the current work, the dual excitation method is extended to detection of four foodborne pathogens, S. typhimurium, S. aureus, Listeria monocytogenes, and Pseudomonas aeruginosa.
Pathogen-specific aptamers were coupled with CdTe QDs at 510 and 720 nm emissions, and NaYF4-based UCNPs at 545 and 800 nm emissions. The resulting nanoparticles were conjugated with the magnetic beads that were previously coupled with the corresponding partial complementary DNA sequences. These conjugates were simultaneously used as molecular recognition elements for the detection. Partially complementary ssDNA-decorated magnetic beads were employed in the sensing system for separation purposes. Aptamer coupled QD and UCNP conjugates, and cDNA decorated magnetic beads were characterized using UV-Vis spectroscopy, Circular Dichroism, and Dynamic Light Scattering techniques. The multiplex detection of the selected foodborne pathogens was implemented using consecutive 335 nm and 980 nm excitations that eliminated the signal overlap between the nanoparticle species. Dual-excitation multiplex aptasensor enabled detection of the targets at trace amounts. The absence of signal cross-talk between the nanoparticles rendered multiplex detection of foodborne pathogens, offering a promising alternative to the conventional methods for detection.

Acknowledgment This project is supported by The Scientific and Technological Research Council of Turkey, TUBITAK Grant ID: 114Z379 (Cost Action CM1403).

1. Yüce, M., Ullah, N. & Budak, H. Trends in aptamer selection methods and applications. Analyst 140, 5379–5399 (2015).
2. Kurt, H., Yüce, M., Hussain, B. & Budak, H. Dual-excitation upconverting nanoparticle and quantum dot aptasensor for multiplexed food pathogen detection. Biosens. Bioelectron. 81, 280–286 (2016).