Helicobacter pylori CagL protein detection kit and detection method thereof

A technology for detection of Helicobacter pylori and protein, which is applied in the field of medicine and biology, can solve problems such as difficult acceptance by patients and family members, and great pain for patients, and achieve the effects of rapid diagnosis, convenient operation, and wide application prospects

Pending Publication Date: 2019-11-19
ZHENJIANG DEWEI LEPU ENERGY ENVIRONMENTAL PROTECTION TECH CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] There are many methods for laboratory diagnosis and detection of Helicobacter pylori infection, including traditional direct microscopic examination of pathological specimens, bacterial culture of living tissue and urease test, etc., but these methods are all invasive and traumatic examinations, which cause great pain to patients. , not easily accepted by patients and their families

Method used

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  • Helicobacter pylori CagL protein detection kit and detection method thereof
  • Helicobacter pylori CagL protein detection kit and detection method thereof
  • Helicobacter pylori CagL protein detection kit and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Helicobacter pylori CagL protein detection kit, coated with mouse monoclonal CagL antibody on a microwell plate, combined with rabbit polyclonal CagL antibody and HRP-labeled goat anti-rabbit IgG antibody, using the principle of double antibody sandwich method for detection CagL protein in samples.

[0044] in:

[0045] The preparation method of rabbit polyclonal CagL antibody is as follows: New Zealand white rabbits are used for subcutaneous injection, and the CagL protein of 0.5 mg / ml is mixed with an equal volume of complete Freund's adjuvant for the first immunization, and the back is subcutaneously injected at multiple points, each 2ml; 14 days later. For the second immunization, take 0.5mg / ml CagL protein mixed with incomplete Freund's adjuvant, and inject subcutaneously on the back, 2ml each; 21 days later, carry out the third immunization, the method is the same as the second immunization, but reduce the injection volume, 1ml each; After 28 days of booster immu...

Embodiment 2

[0066] Based on the kit of Example 1, as a further improvement to the present invention, the detection method of the kit is provided:

[0067] The detection method of the kit comprises the following steps:

[0068] 1) Coating: Dilute the CagL monoclonal antibody to 1:32000 with coating buffer, add 100ul per well to a 96-well microplate, overnight at 4°C, wash the plate 3 times with PBST for 5min each time, and finally shoot Dry;

[0069] 2) Blocking: add 5% skimmed milk powder, 200 μL per well, and incubate at 37°C for 2 hours;

[0070] 3) Washing: Discard the blocking solution, wash the plate 3 times with PBST buffer, each time for 5 minutes, and finally pat dry;

[0071] 4) Add the sample to be tested: add 100ul of the sample to be tested to each well, and add 100ul each of the negative control substance and the positive control substance at the same time, and incubate at 37°C for 1h;

[0072] 5) Washing: Discard the liquid, wash the plate 3 times with PBST buffer, each t...

Embodiment 3

[0083] Based on the detection method in Embodiment 2, in this embodiment, the detection sample of the kit is Helicobacter pylori, and the OD value at a wavelength of 450 / 620 nm is measured by a microplate reader.

[0084] The judgment method is: OD4 detected by the sample 50 / 620nm When the value>0.0536, it can be judged as positive.

[0085] The preparation method is as follows: select well-grown Helicobacter pylori strains, add physiological saline to adjust to suspend bacteria liquid, and adjust the minimum to OD of 1.0.

[0086] The detection values ​​of 20 negative samples are shown in the table below:

[0087]

[0088] Detection value of different concentrations of bacterial liquid

[0089]

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Abstract

The invention proposes a helicobacter pylori CagL protein detection kit and a detection method thereof. In the kit, murine monoclonal antibody CagL antibody is coated on a micropore plate, rabbit polyclonal CaGl antibody and HRP-marked goat anti-rabbit IgG antibody are matched, and CagL protein in a to-be-detected sample is detected by a dual-antibody sandwiched method principle. A convenient andeffective tool is provided for rapid diagnosis and comprehensive treatment of helicobacter pylori, and the kit has relatively wide application prospect and is worth further clinical research.

Description

technical field [0001] The invention relates to a nano reagent kit, in particular to a Helicobacter pylori CagL protein detection kit and a detection method thereof, belonging to the field of medical biotechnology. Background technique [0002] Helicobacter pylori (Helicobacter pylor, Hp) is an important pathogenic bacterium of human gastric diseases, with high infectivity, the global infection rate has reached 50%; Helicobacter pylori is a Gram-negative helical bacterium, Colonize and multiply in gastric epithelial cells. Helicobacter pylori infection is not only the main pathogen of chronic gastritis and peptic ulcer, but also a risk factor for gastric cancer, and it is also related to many diseases outside the stomach. Therefore, the diagnosis of Helicobacter pylori infection has increasingly attracted the attention of many countries and regions. [0003] There are many methods for laboratory diagnosis and detection of Helicobacter pylori infection, including traditiona...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/68G01N33/569
CPCG01N33/68G01N33/56911
Inventor 李冠霖邵世和徐驰王晔
Owner ZHENJIANG DEWEI LEPU ENERGY ENVIRONMENTAL PROTECTION TECH CO LTD
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