Aptamer-based fluorometric determination of Salmonella Typhimurium using Fe3O4 magnetic separation and CdTe quantum dots

Abstract

Based on the high sensitivity and stable fluorescence of CdTe quantum dots (QDs) in conjunction with a specific DNA aptamer, the authors describe an aptamer-based fluorescence assay for the determination of Salmonella Typhimurium. The fluorescence detection and quantification of S. Typhimurium is based on a magnetic separation system, a combination of aptamer-coated Fe3O4 magnetic particles (Apt-MNPs) and QD-labeled ssDNA2 (complementary strand of the aptamer). Apt-MNPs are employed for the specific capture of S. Typhimurium. CdTe QD-labeled ssDNA2 was used as a signaling probe. Simply, the as-prepared CdTe QD-labeled ssDNA2 was first incubated with the Apt-MNPs to form the aptamer-ssDNA2 duplex. After the addition of S. Typhimurium, they could specifically bind the DNA aptamer, leading to cleavage of the aptamer-ssDNA2 duplex, accompanied by the release of CdTe QD-labeled DNA. Thus, an increased fluorescence signal can be achieved after magnetic removal of the Apt-MNPs. The fluorescence of CdTe QDs (λexc/em = 327/612 nm) increases linearly in the concentration range of 10 to 1010 cfu•mL-1, and the limit of detection is determined to be 1 cfu•mL-1. The detection process can be performed within 2 h and is successfully applied to the analysis of spiked food samples with good recoveries from 90% to 105%.

Fig 1

The flow chart diagram of synthesis of QDs (a), QDs-ssDNA2 (b) and aptamer@MNPs (c).

Fig 2

Schematic diagram of (a) the synthesis of streptavidin magnetic nanoparticles and carboxyl CdTe QDs, (b) illustration of the detection of S. Typhimurium.

Fig 3

TEM (a) and HRTEM (b) images of CdTe QDs.

Fig 4

(a) Fluorescence spectra of QDs (curve a) and QDs@ssDNA2 (curve b); (b) UV-vis absorption spectrum of QDs and QDs@ssDNA2.

Fig 5

UV-vis absorption spectrum of MNPs@aptamer (curve a) and aptamer (curve b).

Fig 6

(a) UV-visible absorption spectrum of 10 μL of 1 mg•mL^-1 streptavidin-coated MNPs decorated with 50 μL of 10 nM aptamer. (b) Fluorescence spectra of different concentrations (from 70 μL to 10 μL) of ssDNA2@CdTe QDs of 30 μg·mL^-1 ssDNA2@CdTe QDs. (c) Fluorescence spectra of aptamer&QDs-ssDNA2@MNPs after different incubation times with S. Typhimurium. (d) Fluorescence spectra of aptamer&QDs-ssDNA2@MNPs incubated with S. Typhimurium at different incubation temperatures.

Fig 7

(a) Fluorescence spectra of aptasensors with different concentrations (from a to h: 10¹⁰, 10⁷, 10⁵, 10⁴, 10³, 10², 10, 0 cfu•mL^-1) of S. Typhimurium; (b) calibration curve of the fluorescence intensity of the QDs@ssDNA2 at 612 nm for S. Typhimurium detection.

Fig 8. Specificity result for the detection of S. enteritidis, S. aureus, E. coli O157:H7, L. monocytogenes, B. cereus, P. aeruginosa and S. Typhimurium.