Fluorescent microsphere immunochromatographic test strip for triple quantitative detection of fusarium toxin, preparation method and application of Fluorescent microsphere immunochromatographic test strip
A technology of immunochromatographic test strips and fluorescent microspheres, which is applied in measurement devices, analytical materials, instruments, etc., can solve the problems of single detection, limited sensitivity, instability of quantum dots, etc., and achieves high speed, improved sensitivity, coupled antibodies Simple and fast effects
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Embodiment 1
[0043] Example 1 Preparation of Triple Quantitative Detection Fusarium Toxin Fluorescent Immunochromatographic Test Strips
[0044] 1. Preparation of FB1, DON and ZEN monoclonal antibody-fluorescent microsphere markers. Take 10ug fluorescent microspheres, 0.5-1ug monoclonal antibody and a certain proportion of EDC (the mass ratio of fluorescent microspheres to EDC is 40:1) and add them to 400uL phosphate buffered saline solution (pH=6), then mix well Shake and incubate at 37°C for 10-20min, add 10% BSA to a final concentration of 0.1%, mix well and shake and incubate at 37°C for 10-20min; centrifuge at 12000rpm for 10min, discard the supernatant, and mix the FB1, DON and ZEN monoclonal antibodies - The fluorescent microsphere pellet was resuspended in a storage solution containing 0.9% sodium chloride, 1% BSA and 0.09% sodium azide, and stored at 4°C after ultrasonication for 20s.
[0045] 2. Spray the gold-labeled antibody onto the gold-labeled pad. Before coating the gol...
Embodiment 2
[0049] Embodiment 2 establishes standard curve
[0050] Prepare a range of concentrations (0 / 0 / 0, 0.47 / 0.195 / 0.094, 0.94 / 0.39 / 0.188, 1.875 / 0.78 / 0.375, 3.75 / 1.56 / 0.75, 7.5 / 3.12 / 1.25, 15 / 6.25 / 2.5, 30 / 12.5 / 5, 60 / 25 / 10, 120 / 50 / 20, 240 / 100 / 40ng / mL) FB1, DON and ZEN standards, the prepared solution is containing 5% methanol, 0.5% Tween20, 0.5% PvP k30 10 mM PBS solution. Get 100 μ L of standard solution and drop on the test strip sample pad prepared in embodiment 1, measure the fluorescence value on each detection line and control line on the test strip after 25min, obtain the detection line and the fluorescence intensity ratio on the control line (F=T / C), then take the natural logarithm (LogC) of the concentration value of the standard series as the abscissa and the fluorescence intensity value percentage (F / F o ) to establish a corresponding standard curve for the ordinate, such as figure 2 shown, where F o It is the T / C value when the standard substance concentration is ...
Embodiment 3
[0051] Example 3 sample detection and result analysis
[0052] Add 5 g of sample to 20 mL of extract (methanol: water = 60: 40, v / v), shake vigorously in a shaker for 30 min. After standing still for 30 min, the supernatant was centrifuged at 6000 rpm for 10 min, then the supernatant was filtered with a 0.22 μm polytetrafluoroethylene filter, and diluted 15 times with 10 mM PBS containing 0.5% PvPk30 and 0.5% Tween20, and finally the diluted The sample was centrifuged again at 12000rpm for 10min, and the supernatant was collected. Take 100 μL of the treated sample solution and drop it on the sample pad of the test strip, measure the fluorescence value on the test line and the control line on the test strip after 25 minutes, calculate the content of each toxin in the sample according to the corresponding fluorescence value and the standard curve, like figure 2 shown.
[0053] The results showed that the IC50 values of the above test strips for simultaneous detection of ...
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