HLA-G hypersensitization detecting method by chemiluminescence immune analysis

A technology of chemiluminescence immunity and analysis method, which is applied in the direction of chemiluminescence/bioluminescence, analysis by causing chemical reaction of materials, and analysis of materials, which can solve the problems of low sensitivity and unsuitability for early detection of malignant tumors

Inactive Publication Date: 2009-07-29
GUANGZHOU TIANMEI BIOTECH
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the sensitivity of ELISA to detect HLA-G is relatively low, it is not suitable for detecting early malignant tumors

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The preparation method of the horseradish peroxidase luminescent system: the concentration of the luminescent substrate diacylhydrazine derivative (preferably luminol) is 1.0mMol / L. Compound oxidant: sodium perborate-H 2 o 2 Each is 1.0mMol / L. Composite enhancer: 0.05mMol / L for each of p-hydroxycinnamic acid and p-iodophenol. Adjust the pH with 0.1Mol / L Tris-HCl buffer at pH 8.60.

Embodiment 2

[0028] Establishment of biotin-avidin double monoclonal antibody sandwich one-step method for detection of HLA-G chemiluminescent immunoassay:

[0029] 1. Prepare the complex of monoclonal antibody (1) and biotin and the complex of monoclonal antibody (2) and horseradish peroxidase according to the aforementioned literature, and bind streptavidin to the solid phase carrier.

[0030] 2. To establish a CLIA method for the detection of HLA-G by a one-step sandwich method of avidin-biotin double monoclonal antibody. Add 50 μl each of the serum to be tested (or HLA-G standard), the complex of monoclonal antibody (1) with biotin, and the complex of monoclonal antibody (2) with horseradish peroxidase In the coated Nanc plate wells, incubate at 37°C for 60 minutes, wash 3 times with PBST, add the substrate luminol solution (see Example 1 for the specific preparation), and detect the luminescence value at 425nm with a chemiluminescent immunoassay analyzer. The software calculates the ...

Embodiment 3

[0032]Please measure HLA-G content in normal human blood. 105 cases of normal people (voluntary unpaid blood donors), after testing HLA-G, the content of HLA-G in normal human serum is zero, and no HLA-G is detected in normal human serum.

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Abstract

The invention discloses an ultra-sensitive method for the chemiluminescence immunoassay detection of HLA-G. The method takes artificially synthesized HLA-G antigen fragment as immunogen immune BALB/c mouse for obtaining a hybridoma cell line. By using the combination of an HLA-G monoclonal antibody which is secreted by the hybridoma cell line with a chemiluminescence immunoassay technology, a broad-spectrum tumor diagnostic kit is formed. The broad-spectrum tumor diagnostic kit is used for the clinical diagnosis of various malignant tumors by detecting the HLA-G. The broad-spectrum tumor diagnostic kit detects the HLA-G by a double monoclonal antibody sandwich method or competition method. The chemiluminescence immunoassay technology uses an enzymatic chemiluminescence method or an acridine ester chemiluminescence method.

Description

technical field [0001] The invention belongs to the field of biomedical clinical immune analysis. Establish an early, sensitive and broad-spectrum method for the diagnosis of malignant tumors by detecting HLA-G using chemiluminescence immunoassay. In addition, the present invention can also be used in various researches on physiology, pathology, pharmacology, immunity, reproduction, tumor, organ transplantation and the like. technical background [0002] Tumor is a common disease and frequently-occurring disease, and its incidence rate ranks second. According to the report of the World Health Organization in 1997, the annual death toll of cancer patients in the world is as high as 6.6 million, and by 2020, the number of cancer patients in the world will reach 15 million. Various malignant tumors seriously threaten human health. In the early stage of tumor onset, there are often no subjective symptoms, and once the subjective symptoms are found, it has reached the middle a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/574G01N33/577G01N21/76
Inventor 苏殿杰韩苏魏桂梅
Owner GUANGZHOU TIANMEI BIOTECH
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