Novel Time-Resolved Fluorescence Immunochromatography Paper-Based Sensor with Signal Amplification Strategy for Detection of Deoxynivalenol

Abstract

Immunoassay has the advantages of high sensitivity, high specificity, and simple operation, and has been widely used in the detection of mycotoxins. For several years, time-resolved fluorescence immunochromatography (TRFIA) paper-based sensors have attracted much attention as a simple and low-cost field detection technology. However, a traditional TRFIA paper-based sensor is based on antibody labeling, which cannot easily meet the current detection requirements. A second antibody labeling method was used to amplify the fluorescence signal and improve the detection sensitivity. Polystyrene fluorescent microspheres were combined with sheep anti-mouse IgG to prepare fluorescent probes (Eu-IgGs). After the probe fully reacted with the antibody (Eu-IgGs-Abs) in the sample cell, it was deployed on the paper-based sensor using chromatography. Eu-IgGs-Abs that were not bound to the target were captured on the T-line, while those that were bound were captured on the C-line. The paper-based sensor reflected the corresponding fluorescence intensity change. Because a single molecule of the deoxynivalenol antibody could bind to multiple Eu-IgGs, this method could amplify the fluorescence signal intensity on the unit antibody and improve the detection sensitivity. The working standard curve of the sensor was established under the optimum working conditions. It showed the lower limit of detection and higher recovery rate when it was applied to actual samples and compared with other methods. This sensor has the advantages of high sensitivity, good accuracy, and good specificity, saving the amount of antibody consumed and being suitable for rapid field detection of deoxynivalenol.

Highly sensitive time‐resolved fluoroimmunoassay for the quantitative onsite detection of Alternaria longipes in tobacco

“…To label the mAb against A. longipes, Eu (III) chelate nanospheres were chosen due to its high fluorescence intensity, long fluorescence lifetime, and a sharp emission peak at 610 nm which avoids spectra overlap with autofluorescence in real samples (Dong et al, 2020;Tang et al, 2019). It has been reported that Eu (III) nanosphere-conjugated antibody outperforms the same antibody labelled by gold nanoparticles, quantum dots and fluorescein isothiocyanate when detecting E. coli O157:H7 in milk samples, with the highest sensitivity, a wider working range, and better tolerance in real samples (Luo et al, 2017).…”

Indirect signal amplification strategy with a universal probe-based lateral flow immunoassay for the rapid quantitative detection of fumonisin B1

“…The universal detection probes were prepared following a procedure by Dong et al 32 100 μL polystyrene fluorescent nanoparticles were added into 400 μL boric acid buffer solution (0.2 M, pH 8.18) and mixed with a vortex oscillator. The mixed solution was ultrasonically treated for 10 min to disperse the polystyrene fluorescent nanoparticles homogeneously in the boric acid buffer solution.…”

“…The coated NC film was dried at 37 °C for 2 h before use. The sample pad was treated with blocking solution (2.9 g NaH 2 PO 4 ·12H 2 O, 0.3 g NaH 2 PO 4 ·12H 2 O, 1.0 g PVP-30, 0.5 g BSA, 1.0 g Tween-20 and 0.25 g EDTA were weighed and dissolved in 100.0 mL ultrapure water 32 ) for 15 min, and dried at 37 °C for 2 h. The absorbent pad was used without any treatment. The sample pad, NC membrane and absorbent pad were laminated and pasted on the backing pad successively with an overlap of 1–2 mm.…”

“…5 g of corn or feed sample were weighed after grinding and crushing, 20 mL 70% methanol aqueous solution (v/v) was added to the matrix for 30 min of extraction, and the obtained extract was filtered through a 0.45 μm organic phase filter membrane. 1 mL of the filtrate was taken out and added to 4 mL of sustained release solution (1 g sucrose + 0.5 g OVA + 2.5 g Tween-20 + 0.5 g PVP-30 were weighed and dissolved in 100.0 mL ultrapure water 32 ) to achieve a 5-fold dilution. Then, the diluent solution was filtered through a 0.45 μm organic phase filter membrane again and the filtrate was prepared for further detection.…”

“…However, in this mode, the monoclonal antibody simultaneously combined with the probes and the goat anti-mouse IgG on the C line, which may hinder the specific recognition of the monoclonal antibody and antigen, making the competition mode complicated and reducing sensitivity. On the basis of that, some researchers have proposed novel test strips with goat anti-mouse IgG labeled with fluorescent molecules to detect other small molecule substances; for example, Urusov et al 29 detected zearalenone and aflatoxin B1 with LOD of 6 and 0.6 ng mL −1 , respectively; Majdinasab et al 30 detected ochratoxin A with a LOD as low as 0.4 pg mL; Li et al 31 simultaneously detected aflatoxin M1 and melamine in milk with LOD of 0.009 and 0.024 ng mL −1 respectively; and Dong et al 32 detected deoxynivalenol with a LOD of 0.121 ng mL −1 , which all prove that this indirect LFI method could overcome the limitations mentioned above and achieve excellent performance. However, there were few reports on FB1 detection this assay.…”