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Simultaneous Detection of Three Foodborne Pathogens Based on Immunomagnetic Nanoparticles and Fluorescent Quantum Dots.

ABSTRACT: This paper presents a peptide-mediated immunomagnetic separation technique and an immunofluorescence quantum dot technique for simultaneous and rapid detection of Escherichia coli O157:H7, Staphylococcus aureus, and Vibrio parahaemolyticus. First, three peptides that can specifically recognize the three foodborne pathogens were combined with magnetic nanoparticles to form an immunomagnetic nanoparticle probe for capturing three kinds of target bacteria and then added three quantum dot probes (quantum dots-aptamer), which formed a sandwich composite structure. When the three quantum dot probes specifically combined with the three pathogenic bacteria, the remaining fluorescent signal in the supernatant will be reduced by magnetic separation. Therefore, the remaining fluorescent signal in the supernatant can be measured with a fluorescence spectrophotometer to indirectly determine the three pathogens in the sample. The linear range of the method was 10-107 cfu/mL, and in the buffer, the detection limits of E. coli O157:H7, S. aureus, and V. parahaemolyticus were 2.460, 5.407, and 3.770 cfu/mL, respectively. In the tap water simulation, the detection limits of E. coli O157:H7, S. aureus, and V. parahaemolyticus were 2.730, 1.990 × 101, and 4.480 cfu/mL, respectively. In the milk simulation sample, the detection limits of E. coli O157:H7, S. aureus, and V. parahaemolyticus were 6.660, 1.070 × 101, and 2.236 × 101 cfu/mL, respectively. The method we presented can detect three kinds of foodborne pathogens at the same time, and the entire experimental process did not exceed 4 h. It has high sensitivity and low detection limit and may be used in the sample detection of other pathogens.

SUBMITTER: Wang D

PROVIDER: S-EPMC7495797 | biostudies-literature | 2020 Sep

REPOSITORIES: biostudies-literature

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Simultaneous Detection of Three Foodborne Pathogens Based on Immunomagnetic Nanoparticles and Fluorescent Quantum Dots.

Wang Dan D Lian Fengnan F Yao Shuo S Liu Yi Y Wang Jinpeng J Song Xiuling X Ge Lirui L Wang Yue Y Zhao Yuyi Y Zhang Jiamei J Zhao Chao C Xu Kun K

ACS omega 20200903 36

This paper presents a peptide-mediated immunomagnetic separation technique and an immunofluorescence quantum dot technique for simultaneous and rapid detection of Escherichia coli O157:H7, Staphylococcus aureus, and Vibrio parahaemolyticus. First, three peptides that can specifically recognize the three foodborne pathogens were combined with magnetic nanoparticles to form an immunomagnetic nanoparticle probe for capturing three kinds of target bacteria and then added ...[more]

PMID: 32954157

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Project description:A fluorescent immunoassay based on europium nanoparticles (EuNPs-FIA) was developed for the simultaneous detection of antibiotic residues, solving the problems of single target detection and low sensitivity of traditional immunoassay methods. In the EuNPs-FIA, EuNPs were used as indictive probes by binding to anti-tetracyclines monoclonal antibodies (anti-TCs mAb), anti-sulphonamides monoclonal antibodies (anti-SAs mAb) and anti-fluoroquinolones monoclonal antibodies (anti-FQs mAb), respectively. Different artificial antigens were assigned to different regions of the nitrocellulose membrane as capture reagents. The EuNPs-FIA allowed for the simultaneous detection of three classes of antibiotics (tetracyclines, fluoroquinolones and sulphonamides) within 15 min. It enabled both the qualitative determination with the naked eye under UV light and the quantitative detection of target antibiotics by scanning the fluorescence intensity of the detection probes on the corresponding detection lines. For qualitative analysis, the cut-off values for tetracyclines (TCs), fluoroquinolones (FQs) and sulphonamides (SAs) were 3.2 ng/ml, 2.4 ng/ml and 4.0 ng/ml, respectively, which were much lower than the maximum residue limit in food. For quantitative analysis, these ranged from 0.06 to 6.85 ng/ml for TCs, 0.03-5.14 ng/ml for FQs, and 0.04-4.40 ng/ml for SAs. The linear correlation coefficients were higher than 0.97. The mean spiked recoveries ranged from 92.1 to 106.2% with relative standard deviations less than 8.75%. Among them, the three monoclonal antibodies could recognize four types of TCs, seven types of FQs and 13 types of SAs, respectively, and the detection range could cover 24 antibiotic residues with different structural formulations. The results of the detection of antibiotic residues in real samples using this method were highly correlated with those of high performance liquid chromatography (R 2 > 0.98). The accuracy and precision of the EuNPs-FIA also met the requirements for quantitative analysis. These results suggested that this multiplex immunoassay method was a promising method for rapid screening of three families of antibiotic residues.

Project description:Foodborne pathogens, especially bacteria, are explicitly threatening public health worldwide. Biosensors represent advances in rapid diagnosis with high sensitivity and selectivity. However, multiplexed analysis and minimal pretreatment are still challenging. We fabricate a gold nanoparticle (Au NP)-amplified microcantilever array biosensor that is capable of determining ultralow concentrations of foodborne bacteria including Escherichia coli O157:H7, Vibrio parahaemolyticus, Salmonella, Staphylococcus aureus, Listeria monocytogenes, Shigella, etc. The method is much faster than using conventional tools without germiculturing and PCR amplification. The six pairs of ssDNA probes (ssDNA1 + ssDNA2 partially complementary to the target gene) that originated from the sequence analysis of the specific gene of the bacteria were developed and validated. The ssDNA1 probes were modified with -S-(CH2)6 at the 5'-end and ready to immobilize on the self-assembled monolayers (SAMs) of the sensing cantilevers in the array and couple with Au NPs, while 6-mercapto-1-hexanol SAM modification was carried out on the reference cantilevers to eliminate the interferences by detecting the deflection from the environment induced by non-specific interactions. For multianalyte sensing, the target gene sequence was captured by the ssDNA2-Au NPs in the solution, and then the Au NPs-ssDNA2-target complex was hybridized with ssNDA1 fixed on the beam of the cantilever sensor, which results in a secondary cascade amplification effect. Integrated with the enrichment of the Au NP platform and the microcantilever array sensor detection, multiple bacteria could be rapidly and accurately determined as low as 1-9 cells/mL, and the working ranges were three to four orders of magnitude. There was virtually no cross-reaction among the various probes with different species. As described herein, it holds great potential for rapid, multiplexed, and ultrasensitive detection in food, environment, clinical, and communal samples.

Dataset Information

Simultaneous Detection of Three Foodborne Pathogens Based on Immunomagnetic Nanoparticles and Fluorescent Quantum Dots.

Publications

Simultaneous Detection of Three Foodborne Pathogens Based on Immunomagnetic Nanoparticles and Fluorescent Quantum Dots.

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