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Colloidal gold test paper strip for detecting Listeria or Listeria monocytogenes in food based on monoclonal antibody, and applications thereof

A technology of Listeria monocytogenes and monoclonal antibody, which is applied in the field of immunoassay, can solve the problems such as cross-reaction is difficult to meet the requirements, and achieve the effect of simple and fast analysis, high sensitivity and high accuracy

Inactive Publication Date: 2016-11-09
JIANGNAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there have been studies on Listeria and Listeria monocytogenes colloidal gold test paper methods, but the main problem is that the cross-reactivity of the monoclonal antibody or polyclonal antibody and the established detection method is difficult to meet the requirements, that is, for Listeria monocytogenes. Bacteria of different species within the genus Tetrabacterium or different serotypes of Listeria monocytogenes can be identified, and there is no cross-reaction to other miscellaneous bacteria, which is very important for practical application in sample detection

Method used

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  • Colloidal gold test paper strip for detecting Listeria or Listeria monocytogenes in food based on monoclonal antibody, and applications thereof
  • Colloidal gold test paper strip for detecting Listeria or Listeria monocytogenes in food based on monoclonal antibody, and applications thereof
  • Colloidal gold test paper strip for detecting Listeria or Listeria monocytogenes in food based on monoclonal antibody, and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Embodiment 1 Colloidal gold test strip

[0046] Colloidal gold test strips for the detection of Listeria or Listeria monocytogenes in food based on monoclonal antibodies, including absorbent pad 1, nitrocellulose membrane 2, quality control line 3, detection line 4, sample pad 5, PVC Bottom plate 7, equipped with gold-labeled antibody 6;

[0047] The sample pad 5, the nitrocellulose membrane 2 and the water-absorbing pad 1 are sequentially arranged on the PVC bottom plate 7; the nitrocellulose membrane 2 is directly arranged on the PVC bottom plate 7; directly set on the PVC bottom plate 7; one end of the water-absorbing pad 1 is set on the PVC bottom plate 7, and the other end is lapped on the nitrocellulose membrane 2;

[0048] The nitrocellulose membrane 2 is sequentially provided with a quality control line 3 and a detection line 4; the quality control line 3 is coated with a goat anti-mouse IgG secondary antibody, and the detection line 4 is coated with antibody 2...

Embodiment 2

[0051] The preparation method of the colloidal gold test strip of Listeria genus or Listeria monocytogenes in the detection food based on monoclonal antibody, the steps are:

[0052] (1) Preparation of gold-labeled antibody 6:

[0053] a, the synthesis of colloidal gold solution: adopt the citric acid reduction method to synthesize 30nm gold nanoparticles, add 300mL of chloroauric acid solution with a mass concentration of 0.01% in the cleaned flask, heat to a complete boil under magnetic stirring, and pour into the solution Quickly add 4.8mL of 1% trisodium citrate solution, boil the solution for 10min until the color turns bright wine red, cool the flask at room temperature, and store at 4°C;

[0054] b. Coupling of gold-labeled antibody: Use antibody 2H8 or antibody No. A to label colloidal gold particles, the specific process is: use 0.1M potassium carbonate solution to adjust the pH of 1mL colloidal gold solution prepared in step a to 7.5; add monoclonal antibody 10 μg o...

Embodiment 3

[0057] Example 3 Wet detection of P60 protein

[0058] Sample dilution: P60 protein was diluted with 10mM PBS to 250ng / mL, 100ng / mL, 50ng / mL, 25ng / mL, 10ng / mL, 5ng / mL, 2.5ng / mL, 1ng / mL and blank PBS was used as control. Listeria colloidal gold test strips and Listeria monocytogenes colloidal gold test strips were used for detection. Take 7 μL of the gold-labeled antibody and 47 μL of the resuspension solution, that is, 0.01 M 10 mM phosphate buffer solution containing 0.2% Tween and 0.2% sucrose, and react 150 μL of the sample to be tested in a microwell plate for 5 minutes, and then the colloidal gold The test strips were inserted into the microwell plate, and read after 10 minutes of reaction. Interpretation basis: Positive samples: Quality control line and detection line are colored at the same time. Negative sample: the quality control line is colored, and the test line is not colored. The specific test results are shown in Figure 2. The test results show that the List...

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Abstract

The invention relates to a colloidal gold test paper strip for detecting Listeria or Listeria monocytogenes in food based on monoclonal antibody, and applications thereof, and belongs to the technical field of immunological analysis. According to the colloidal gold test paper strip, a sample pad, a nitrocellulose membrane and a water absorbing pad are sequentially arranged on a PVC bottom plate; the gold-labeled antibody is the colloidal gold solution of antibody 2H8 when the Listeria in the food is detected based on the monoclonal antibody; and the gold-labeled antibody is the colloidal gold solution of antibody A when the Listeria monocytogenes in the food is detected based on monoclonal antibody. According to the present invention, the colloidal gold test paper strip for detecting the Listeria or the Listeria monocytogenes in the food based on the monoclonal antibody is developed, and is suitable for the high-specificity, high-accuracy, high-sensitivity, rapid and convenient analysis of the Listeria or the Listeria monocytogenes in the food.

Description

technical field [0001] The invention relates to a colloidal gold test strip for detecting Listeria or Listeria monocytogenes in food based on a monoclonal antibody and an application thereof, belonging to the technical field of immune analysis. Background technique [0002] Listeria monocytogenes is a facultative anaerobic Gram-positive bacterium belonging to the genus Listeria. It is primarily food-borne and is one of the deadliest foodborne pathogens worldwide. Listeria is widely distributed in the environment, and meat, eggs, poultry, dairy products, vegetables, etc. have all been proven to be the source of Listeria infection. Infection with Listeria can cause symptoms such as muscle pain, nausea, and diarrhea. In severe cases, it can cause blood and brain tissue infections. Moreover, since the bacteria can still grow and reproduce at a refrigeration temperature of 4°C, contaminated refrigerated food is harmful to human health. Big threat. Therefore, many countries hav...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/569G01N33/558
CPCG01N33/56911G01N33/558G01N33/577
Inventor 胥传来王文彬匡华徐丽广马伟刘丽强吴晓玲宋珊珊胡拥明
Owner JIANGNAN UNIV
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