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Test paper box for detecting listeria monocytogenes

A technology of Listeria and test paper box, applied in the field of bacteriology and immunology, can solve the problems of obvious material matrix effect, human health and safety impact, poor stability, etc., and achieve the effects of simple process, convenient preparation and cost saving

Inactive Publication Date: 2015-01-21
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] 1. Colloidal gold is a kind of nanoparticle. It is difficult to prepare colloidal gold nanoparticles, and the particle diameter is difficult to control uniformly;
[0005] 2. A strong reducing agent is used in the colloidal gold labeling process, and at the same time, it is exposed to heavy metal ions, which has a certain impact on human health and safety;
[0006]3. The colloidal gold labeling process is a physical adsorption process, so the stability in the liquid phase is poor;
[0007]4. Only when the gold particles gather to a certain amount, the naked eye can observe the purple-red band, and the color band has little contrast with the background, thus limiting Improve the detection sensitivity;
[0008] 5. The matrix effect of different materials is obvious, and the background interference is very large;
Antigen pairing process takes a long time, increasing R&D costs

Method used

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  • Test paper box for detecting listeria monocytogenes
  • Test paper box for detecting listeria monocytogenes
  • Test paper box for detecting listeria monocytogenes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 : Single antibody test strip structure description and preparation method of the present invention

[0031] Production of single antibody test strips, spraying Listeria monocytogenes antibody on the nitrocellulose membrane as the detection line (T line) and fluorescent dye antibody spraying quality control line (C line);

[0032] The cultured bacteria were taken out from the shaker, washed 3 times with 1*PBS containing 5% Tween 20, washed once with 75% ethanol during this period, and centrifuged at 1W rpm for 5min. Stain the washed cells with propidium iodide (PI), and add PI directly. After staining for 5 minutes with PI concentration of 25ug / ml, wash 3 times with 1*PBS of 5% Tween 20 and resuspend the pellet.

[0033] Take 20uL of the mixed solution of various bacteria after staining, mix it with 90uL of 5% Tween 20 1*PBS, add it into the sample hole of the single antibody test strip and colloidal gold test strip prepared by us, and react for 15 minutes. ...

Embodiment 2

[0035] Example 2 : Test strips after bacteria stained with other dyes

[0036] Common dyes saffron, sand yellow, crystal violet and common fluorescent dye DiOC 18 (3) After staining Listeria monocytogenes with fluorescein isothiocyanate (FITC) for 10 minutes, load the sample onto the test strip of the present invention.

[0037] Spray the corresponding dye antibody on the C line;

[0038] The result is as image 3 As shown, other dyes were found to be unable to develop color on the test strip. The reason may be that these dyes are not suitable for use in test strips. When they combine with bacteria, they also combine with the nitrocellulose membrane or sample pad of the test strip, so that the stained bacteria cannot migrate on the test strip. , and finally cannot develop color. And four kinds of fluorescent dyes of propidium iodide, fluorescein diacetate (FDA), acridine orange or SYBR Green screened by the present invention can be preferably suitable for the use of test...

Embodiment 3

[0039] Example 3 : test paper box of the present invention detects Listeria monocytogenes

[0040] Get the test paper box that embodiment 1 prepares;

[0041] The number of cells taken was 10 3 、10 4 、10 5 、10 6 、10 7 、10 8 、10 10CFU / mL and blank group Listeria monocytogenes suspension were stained with 0.01% fluorescent dye solution for 5 min. Add 4 drops of the sample (about 100 μL) to the sample hole of the test strip after the stained bacterial liquid. After reacting for 15 minutes, judge the result under the ultraviolet light. There are two bands at the detection line, and the result is positive. There is no bright band in the detection line, and the result is negative;

[0042] Compared with the detection results of Listeria monocytogenes colloidal gold test strips with the same bacterial count, Figure 4 The test results of colloidal gold test strips for Listeria monocytogenes showed that the minimum detection limit was 10 bacteria 6 to 10 5 between. The ...

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Abstract

The invention relates to an immunochromatography test paper box based on a thallus fluorescent staining technology, which is used for detecting stained listeria monocytogenes by using a single antibody. The immunochromatography test paper box comprises a single antibody test paper strip and a fluorescent dye box. The immunochromatography test paper box has the advantages that thalli of the listeria monocytogenes are pre-stained by a fluorescent dye and a specific antibody of the listeria monocytogenes is sprayed on a T line of the test paper strip to capture the thalli of the listeria monocytogenes in a mixed sample, so that the rapid and specific detection of the thalli in the sample is realized.

Description

technical field [0001] The invention relates to the technical fields of bacteriology and immunology. Specifically, the invention relates to an immunochromatographic test strip for detecting bacteria with a single antibody and a preparation method thereof. Background technique [0002] Immunochromatography (immunochromatography) is a rapid detection technology established on the basis of immunofiltration in the late 1980s and early 1990s. The antibody (or antigen) of the antigen-antibody reaction is pre-coated on the specific position of the immunochromatographic test paper in the form of a strip, and the antigen (or antibody) in the sample to be tested is pre-labeled by the pigment during the process of developing with the developing solution. When the antigen (or antibody) in the sample to be tested marked by the dye is expanded to a specific position (that is, the position where the antibody (or antigen) is coated in a strip), the antigen (or antibody) in the sample to be...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
CPCG01N33/56916
Inventor 徐锋魏华陈星星武晓丽叶若松甘敏王登远杨栋刘琚珥许恒毅胡海宁
Owner NANCHANG UNIV
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