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

Date Published: June 19, 2019

Publisher: Public Library of Science

Author(s): Junan Ren, Gang Liang, Yan Man, An Li, Xinxin Jin, Qingju Liu, Ligang Pan, Yogendra Kumar Mishra.

http://doi.org/10.1371/journal.pone.0218325

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%.

Partial Text

Bacterial infection is a major problem for human health because of their toxin system and probable antibiotic resistance [1,2]. Salmonella Typhimurium (S. Typhimurium) is one of the most important prevalent pathogens in humans, causing diarrhea, fever, and abdominal cramps [3,4]. This salmonellosis is mostly related to contaminated foods that mainly originate from animal sources, including poultry eggs, milk, beef, and raw food, and non-animal foods, such as fruits, vegetables, and spices [5]. Currently, S. Typhimurium is considered a key concern in many countries, such as the US, China, Europe, and Japan. Therefore, it is important to develop a method that can detect S. Typhimurium rapidly, accurately and sensitively for food quality control.

In this paper, we report a magnetic separation system-based fluorescence sensing strategy to detect and quantify S. Typhimurium with high sensitivity and selectivity. QD-ssDNA2 was incubated with Apt-MNPs to form an aptamer-complementary DNA duplex as a detection probe. Upon addition of S. Typhimurium, QD-ssDNA2 is replaced by the bacteria and released from the Fe3O4 MNPs, accompanied by the release of CdTe QD-labeled ssDNA2, resulting in a significantly increased fluorescence intensity. The difference in fluorescence intensity can be used to sensitively detect S. Typhimurium, with a low detection limit of 1 cfu•mL-1. Furthermore, the fluorescence sensor was also successfully applied to detect S. Typhimurium in water and milk. In addition, the aptamer and QD-modified complementary sequences are more cost-effective and more stable than antibodies, and the assay can capture targets with magnetic beads and use fluorescence to quantify S. Typhimurium simultaneously within 2 h. In conclusion, the sensing assay has the potential to be further extended to on-site screening of pathogenic bacteria S. Typhimurium-related food contamination and other pathogenic bacteria targets by changing the aptamer and cDNA.

 

Source:

http://doi.org/10.1371/journal.pone.0218325

 

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