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Kit for detecting helicobacter pylori antibody by using latex immunoturbidimetry assay

A technology for Helicobacter pylori and latex immunization, which is applied in measurement devices, color/spectral property measurement, instruments, etc., can solve the problems of large differences between batches of natural antigens, long cycle, and cumbersome preparation process, and is not easy to interfere and operate. Simple, broad-spectrum effects

Active Publication Date: 2013-01-02
WUHAN CHANGLI BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to overcome the shortcomings of using natural antigens to prepare latex reagents at present. There are large differences between batches of natural antigens, and the preparation process is cumbersome and the cycle is long. It provides a latex immune kit using genetic recombinant antigens to measure pylorus. Helicobacter antibody and method for using the reagent to detect clinical samples

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  • Kit for detecting helicobacter pylori antibody by using latex immunoturbidimetry assay
  • Kit for detecting helicobacter pylori antibody by using latex immunoturbidimetry assay
  • Kit for detecting helicobacter pylori antibody by using latex immunoturbidimetry assay

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Embodiment 1

[0033] In order to make the present invention easier to understand, the present invention will be further described below with reference to specific embodiments, which are only used to illustrate the present invention and not to limit the scope of the present invention. Example 1: Preparation of Helicobacter pylori gene recombinant antigen (taking urease B antigen as an example)

[0034] (a) Protein expression

[0035] Using the nucleic acid sequence (AAU21200.1 GI:51989332) published by Helicobacter pylori urease B in the GeneBank database as the template, the C-terminal nucleotide sequence rich in antigenic determinants was retained by referring to the protein sequence analysis software, and the N-terminal was truncated by 450 Then, the synthesized gene sequence (sequence shown in SEQ ID No. 1) was inserted into the expression vector pET30a with 6X His Tag, and the expression vector was used to transform Escherichia coli BL21. When the cultures reached an OD600 value of 80,...

Embodiment 2

[0039] Example 2: Preparation of Helicobacter pylori antibody latex immune kit

[0040] 1. Preparation of reagent R1

[0041] First dissolve NaCl (7.0-9.0g) with double distilled water, then add Tris-HCl buffer, and finally add double distilled water to 1000ml to make the final concentration of Tris 0.05mol / L, shake well, then add a small amount of PEG 6000 and Tween80, mix well.

[0042] Those skilled in the art can also choose other conventional buffers, such as one or more of phosphate buffer, HEPES buffer, glycine buffer, and barbiturate buffer.

[0043] 2. Preparation of reagent R2

[0044] Disperse the latex with pH=5 MES solution to make its concentration 1%, add EDC and mix evenly, shake for 12-15 minutes, and activate the latex with a particle size of 90nm-300nm and a carboxyl functional group. 14,000 rpm, centrifuge for 10 minutes to collect latex precipitate, shake or ultrasonically disperse the latex with 1XPBS, and adjust the latex concentration to 2%. Add the...

Embodiment 3

[0048] Example 3: Assay method for Helicobacter pylori antibody detection reagent

[0049] This kit is suitable for automatic or semi-automatic biochemical analyzers of Beckman, Hitachi, Olympus, Toshiba, Roche, Abbott, Siemens, Mindray and other brands. The determination method is as follows:

[0050] 1. Measurement conditions

[0051] Assay conditions of the kit of the present invention

[0052] Temperature: 37℃

[0053] Dominant wavelength: 548nm

[0054] Secondary wavelength: 800nm

[0055] Sample volume: 5μL

[0056] R1 dosage: 200μL

[0057] R2 dosage: 50μL

[0058] Analysis type: Two-point endpoint method

[0059] 2. The measurement method is as follows:

[0060] To measure the blank absorbance, add 5 μL of sample to 200ul R1 reagent, pre-incubate for 5 minutes, after measuring the blank absorbance, add 50 μL of R2 reagent, incubate for 5 minutes, and measure the reaction absorbance.

[0061] The sample volume, R1 dosage and R2 dosage can be adjusted according ...

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PUM

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Abstract

The invention relates to the technical field of biology and concretely discloses a kit for detecting a helicobacter pylori antibody by using a latex immunoturbidimetry assay. The kit provided by the invention comprises a helicobacter pylori antibody calibrator, a buffer solution with the pH value of 6.5-8.5 and a helicobacter pylori gene recombination antigen latex reagent. The kit provided by the invention is high in sensitivity up to 0.10mu g / ml due to the adoption of the latex immunoturbidimetry assay for detecting the content of helicobacter pylori antibodies in a detected sample, good in stability, simple and rapid in operation, strong in specificity, difficult to be interfered, accurate in quantifying and wide in application prospect.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a latex immune reagent prepared by using a gene recombinant antigen and a kit for detecting Helicobacter pylori antibodies by a latex immune turbidimetric method containing the reagent. Background technique [0002] Helicobacter pylori, or Hp for short, was first discovered by Barry J. Marshall and J. Robin Warren, who won the 2005 Nobel Prize in Physiology or Medicine prize. Helicobacter pylori is a unipolar, polyflagellar, blunt-ended, spiral-curved bacterium, 2.5-4.0 μm long and 0.5-1.0 μm wide. The surface of gastric mucosal epithelial cells often presents a typical spiral or arc shape. [0003] Helicobacter pylori infection is the main pathogenic factor of chronic active gastritis, peptic gastric ulcer and gastric mucosa-associated lymphoid tissue (MALT) lymphoma, and is closely related to the occurrence of gastric cancer. WHO / IARC) has classified Helicobacter pylori as a cla...

Claims

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Application Information

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IPC IPC(8): G01N33/569G01N33/531G01N33/536G01N21/31
Inventor 张英伟
Owner WUHAN CHANGLI BIOLOGICAL TECH CO LTD
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