Enzyme-linked immunosorbent assay kit for detecting acid orange and its application
The technology of enzyme-linked immunosorbent reagent and acid orange is applied in the detection field of acid orange content in marinated pork, bean paste and hot sauce, which can solve the problem of low sensitivity of thin-layer chromatography scanning method, expensive equipment and unsuitable promotion and application, etc. problem, to achieve the effect of efficient detection method, simple structure and simple pre-processing process
- Summary
- Abstract
- Description
- Claims
- Application Information
AI Technical Summary
Problems solved by technology
Method used
Image
Examples
Embodiment 1
[0028] Embodiment 1 Preparation of kit components
[0029] 1. Preparation of acid orange hapten
[0030] Dissolve 280mg of p-aminobenzoic acid in 20mL of 0.2mol / L HCl solution, add 140mg of NaNO dropwise at 0-4°C 2 5mL aqueous solution, and react in the dark for 0.5~1h, then slowly dropwise add the mixture of 240mg acid orange II in 5mL0.5mol / L sodium acetate solution, and react in the dark for 1~3h; Wash with water, extract with ethyl acetate, and remove the solvent to obtain the Acid Orange II azo derivative recrystallized in ethanol-water, which is the Acid Orange hapten.
[0031] Get above-mentioned product and measure through proton nuclear magnetic resonance spectrum, such as figure 2 As shown, the increased carboxyl signal peak around 12.7ppm and the increased aromatic ring signal peak around 8.1-8.4ppm indicate that the hapten was successfully synthesized.
[0032] 2. Antigen preparation
[0033] Immunogen preparation—the acid orange hapten was coupled with bovine...
Embodiment 2
[0050] Example 2 The formation of the enzyme-linked immunosorbent assay kit for detecting acid orange
[0051] An enzyme-linked immunosorbent assay kit for acid orange detection was constructed to include the following components:
[0052] (1) A microtiter plate coated with acid orange conjugated antigen;
[0053] (2) 6 bottles of acid orange standard solution, the concentrations are 0μg / L, 0.5μg / L, 1.5μg / L, 4.5μg / L, 13.5μg / L, 40.5μg / L;
[0054] (3) Acid orange monoclonal antibody working solution;
[0055] (4) goat anti-mouse anti-antibody labeled with horseradish peroxidase;
[0056] (5) Substrate chromogenic solution is made up of A liquid and B liquid, and A liquid is carbamide peroxide, and B liquid is tetramethylbenzidine;
[0057] (6) The stop solution is 2mol / L sulfuric acid;
[0058] (7) The washing solution has a pH value of 7.4, contains 0.5% to 1.0% Tween-20, 0.01‰ to 0.03‰ sodium azide preservative, and 0.1 to 0.3mol / L phosphate buffer, and the percentage is ...
Embodiment 3
[0060] Example 3 Detection of sour orange in marinated pork, bean paste and hot sauce samples
[0061] 1. Sample pretreatment
[0062] Marinated Pork and Chili Sauce Samples:
[0063] Weigh 2.0g±0.05g sample into a 50mL polystyrene centrifuge tube, add 4mL acetonitrile, then add 6mL deionized water, vortex with a vortexer until uniform, above 3000g, and centrifuge at room temperature (20-25°C) for 5min; Take 100 μL supernatant in a 2 mL polystyrene centrifuge tube, add 900 μL complex solution, then add 800 μL n-hexane, vortex for 30 s with a vortexer, centrifuge at room temperature (20-25 ° C) for 5 min; remove the upper organic phase , remove 50 μL of the layer for analysis.
[0064] Bean paste sample:
[0065] Weigh 2.0g±0.05g of sample into a 50mL polystyrene centrifuge tube, add 6mL of acetonitrile, then add 4mL of 0.1mol / L carbonate buffer (pH9.1), vortex with a vortexer until uniform, more than 3000g , centrifuge at room temperature (20-25°C) for 5 min; take 100 μL o...
PUM
Login to View More Abstract
Description
Claims
Application Information
Login to View More