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Preparation method and application of kit for simultaneously detecting various nafil substances based on novel multi-quantum dot-protein polymer probe

A kit and technology of fluorescent quantum dots, which are used in measurement devices, material excitation analysis, fluorescence/phosphorescence, etc., to achieve the effect of simplifying the preparation method

Pending Publication Date: 2022-08-09
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Although the lateral flow immunochromatography method currently used for on-site screening of nafil-like western medicines that may be illegally added in food has high sensitivity, for example, patent CN114308153A, a solid-phase extraction microfluidic chip for detecting nafil-like substances and its application system for the detection of nafils, but there is still room for further improvement in terms of sensitivity and ease of probe preparation

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  • Preparation method and application of kit for simultaneously detecting various nafil substances based on novel multi-quantum dot-protein polymer probe
  • Preparation method and application of kit for simultaneously detecting various nafil substances based on novel multi-quantum dot-protein polymer probe
  • Preparation method and application of kit for simultaneously detecting various nafil substances based on novel multi-quantum dot-protein polymer probe

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0073] Example 1 Preparation method of self-assembled quantum dot-protein complex fluorescent probe

[0074] 1. Experimental method

[0075] Using the emulsification method, the red oil-soluble quantum dots were added to the PBS solution containing BSA and antibody under the action of ultrasound, and then emulsification was continued to obtain a uniform product. The specific preparation steps are as follows:

[0076] 1. Dissolve red oil-soluble quantum dots ZnCdSe / ZnS QDs (Q1625, Wuhan Jiayuan Quantum Dots Co., Ltd.) in n-hexane to make the quantum dot solution concentration 2 mg / mL.

[0077] 2. Take 12 μL of the 2 mg / mL quantum dot solution prepared in step 1 and add it to 100 μL of stirring PBS solution (containing 1 mg / mL BSA, 0.3 mg / mL Ab Sildenafil Antibody 1 μL and Ab at 0.34 mg / mL Tadalafil Antibody 1 μL), stirred at a speed of 500 rpm for 3 h at 25-30° C. to obtain the stirred quantum dot polymer MQDs-BSA-Ab NPs. the Ab Sildenafil The preparation method is derived...

Embodiment 2

[0084] Example 2 Kit for detecting Nafil substances

[0085] 1. Preparation method

[0086] 1. Preparation of immunochromatographic test strips

[0087] like Figure 4 As shown, the immunochromatographic strips for the detection of sildenafil and tadalafil consist of a sample pad, a nitrocellulose membrane (NC membrane), an absorbent paper and a PVC bottom plate. Take a 30cm-long NC film and place it on the film marking machine. With the long side of the film as the horizontal side and the short side as the vertical side, draw three 1mm vertical lines from left to right, the first two are detection lines, and the third is the detection line. The first test line is 7mm from the left short side, the two test lines are separated by 6mm, and the quality control line and the second test line are separated by 6mm.

[0088] 20μL of tadalafil antigen and sildenafil antigen with a concentration of 0.19mg / mL and 0.38mg / mL were sprayed to the first detection line and the second line a...

Embodiment 3

[0095] Example 3 Detection flow of lateral flow immunochromatography

[0096] For the detection principle, see Figure 5 ,Specific steps are as follows:

[0097] 1. Use a small syringe pump to inject 1 mL of methanol and 1 mL of ultrapure water into the solid-phase extraction microfluidic chip successively to activate and clean the MCX filler in the chip.

[0098] 2. Use a small syringe pump to inject the sample extraction solution to be tested into the chip at a flow rate of 80 μL / min.

[0099] 3. After the sample extraction solution is exhausted, use a small syringe pump to inject methanol solution containing 0.1% acetic acid into the chip at a flow rate of 20 μL / min to elute the analyte on the chip. At the same time, in the solvent evaporation area of ​​the chip A ceramic heating plate was placed under it, and the temperature of the ceramic heating plate was set to 60°C to evaporate methanol.

[0100] 4. After the eluent is exhausted, inject the chromatographic fluid int...

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Abstract

The invention discloses a preparation method and application of a kit for simultaneously detecting various nafil substances based on a novel multi-quantum dot-protein polymer probe. The multi-quantum dot-protein immune probe is prepared by a one-step method based on a novel solvent-induced effect strategy, and the fluorescent immune probe with a large number of fluorescent quantum dots is prepared by mixing the quantum dots, the BSA and the antibody and utilizing hydrophobic interaction and pi-pi accumulation, so that the loading capacity of the quantum dots in the probe is improved, and the preparation method of the probe is simplified. Sildenafil, tadalafil and derivatives thereof are taken as detection objects, sildenafil and tadalafil antibodies and antigens are combined with the method to prepare a probe and a lateral flow immunochromatography test strip for specifically detecting sildenafil, tadalafil and derivatives thereof, a solid-phase extraction micro-fluidic chip is combined to construct a system for detecting nafil substances, and the system is used for detecting the sildenafil, tadalafil and derivatives thereof. Qualitative and quantitative detection of the 12 nafil drugs is achieved, the specificity is high, the sensitivity is high, the accuracy is good, and the detection result is reliable.

Description

technical field [0001] The invention relates to the technical field of food and drug safety detection, in particular to a preparation method and application of a kit for simultaneous detection of a variety of nafil substances based on novel multi-quantum dot-protein polymer probes. Background technique [0002] Compared with traditional probes such as colloidal gold, latex microspheres, and fluorescent dyes, fluorescent quantum dots (QDs) have become a popular choice for constructing highly sensitive immune systems due to their high fluorescence intensity, long fluorescence lifetime, resistance to photobleaching, and diverse luminescence colors. Popular material for chromatography. However, high-quality fluorescent quantum dots are mostly fat-soluble, and need to undergo tedious surface chemical modification to modify the antibody; and due to the small size of quantum dots, the coupling efficiency of quantum dots and antibodies is low, and the loading of quantum dots on anti...

Claims

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Application Information

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IPC IPC(8): G01N33/533G01N33/543G01N21/64
CPCG01N33/533G01N33/54387G01N21/6428
Inventor 雷红涛关甜沈玉栋王锦沈兴李向梅韦晓群徐小艳徐振林杨金易
Owner SOUTH CHINA AGRI UNIV
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