Fluorescence immunity chromatography test strip for escherichia coli O157:H7 detection
A technique of fluorescence immunochromatography and O157, which is applied in the detection field to achieve the effects of fast detection speed, high sensitivity and strong specificity
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Embodiment 1
[0029] Example 1 Preparation and condition optimization of water-soluble core-shell quantum dots
[0030] CdTe / CdS core-shell quantum dots were synthesized in aqueous solution by using an inorganic aqueous phase synthesis method with mercaptopropionic acid as a stabilizer. The preparation conditions of CdTe / CdS quantum dots were optimized through single factor experiments, and the optimal level of each influencing factor was obtained: Cd 2+ with Te 2- The molar ratio is 5:1, the stabilizer MPA and Cd 2+ The molar ratio of CdTe is 2.5:1, the pH of the reaction system is 9, the reaction temperature is 95°C, the reflux time of CdTe core is 1h; the reflux time of CdS shell is 1h. Under these conditions, the fluorescence intensity of the synthesized CdTe / CdS is 726.41, which is 50.10% higher than the initial condition. The particle size of the quantum dots is about 3.0nm, and the dispersion is good. Compared with CdTe quantum dots, the core-shell structure The fluorescence...
Embodiment 2
[0039] Example 2 Labeling of E. coli O157:H7 rabbit polyclonal antibody and preparation of fluorescent immunochromatographic test strips
[0040] 1) Conditions for quantum dot-labeled Escherichia coli O157:H7 rabbit polyclonal antibody
[0041] Covalently coupled with EDC·HCL and NHS as coupling agent, take 100 μL of quantum dots, add 40 μL 0.1 mol / L EDC·HCl and 20 μL 0.1 mol / L NHS respectively, supplement pH 7.4, 0.01 mol / L PBS to The total volume was 800 μL, activated at room temperature for 15 minutes, then 80 μL of rabbit IgG with a concentration of 1 mg / mL was added, and the mixture was reacted in a shaker at 37 °C in the dark for 1.5 h, and finally the reaction was terminated with 100 μL of 1 mg / mL glycine, placed in overnight at 4°C.
[0042] The reaction mixture was repeatedly ultrafiltered with a 100k ultrafiltration membrane to remove the CdTe / CdS quantum dots that were not coupled to the Escherichia coli O157:H7 polyclonal antibody, and the CdTe / CdS-a...
Embodiment 3
[0048] The test of embodiment 3 samples and result judgment
[0049] Add the sample liquid to the sample pad dropwise, or insert one end of the sample pad into the sample liquid. After the immunochromatography is finished, irradiate the result under the ultraviolet light. If the detection line (C line) and the quality control line (T line) appear at the same time A yellow fluorescent band is a positive result; if there is a fluorescent band on the C line and no fluorescent band on the T line, it is a negative result; if neither the C line nor the T line has a fluorescent band, or the T line has a fluorescent band but the C line does not. If the fluorescent band appears, it is judged that the test strip is invalid ( Figure 8 ).
[0050] 1) Detection limit: heat inactivate the pure culture of Escherichia coli O157:H7 and dilute to different concentrations respectively, add dropwise to the sample pad, carry out chromatographic reaction, irradiate the results under ultraviolet l...
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