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Kit for early diagnosis of glioma

A technology for glioma and its application, which is applied in the direction of disease diagnosis, biological testing, material inspection products, etc., can solve the problems of differential diagnosis depending on pathology and difficulty in early diagnosis of glioma, and achieve high clinical application value, high specificity, The effect of high sensitivity

Active Publication Date: 2014-10-01
AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0008] Another object of the present invention is to provide a kit for the early diagnosis of glioma. In the present invention, the B7-H4 glioma tumor marker is used for glioma diagnosis, screening and evaluation of prognosis, which can solve the problem of At present, the early diagnosis of glioma is difficult, and the differential diagnosis is too dependent on pathology, and it should be applied to relevant treatment options and prognosis assessment

Method used

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  • Kit for early diagnosis of glioma
  • Kit for early diagnosis of glioma
  • Kit for early diagnosis of glioma

Examples

Experimental program
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Effect test

Embodiment 1

[0023] 1. Western blot detection of B7-H4 expression in normal brain tissue and different grades of glioma:

[0024] Preparation of protein samples: Mince brain tissue, add homogenization buffer, mechanically or ultrasonically homogenize at room temperature for 0.5-1min. Then centrifuge at 12,000 g for 15 min at 4°C. Take the supernatant as a sample;

[0025] Electrophoresis: prepare SDS-PAGE electrophoresis gel, add sample buffer to the sample, and after cooling to room temperature, load the protein sample to the gel for electrophoresis; transfer membrane: cut the gel strip to a suitable size after electrophoresis Membrane buffer equilibration, 5min×3 times. Pre-cut the filter paper and NC membrane with the same size as the rubber strip, and soaked in the transfer buffer for 10 minutes; the transfer device is sequentially arranged as anode carbon plate, 24-layer filter paper, NC membrane, gel, 24-layer filter paper, cathode carbon The order of the plates is placed, the fil...

Embodiment 2

[0039] Embodiment 2, preparation kit for the early diagnosis of glioma

[0040] The kit includes the following components:

[0041] 1. Solid-phase 96-well enzyme plate;

[0042] 2. Mouse anti-human B7-H4 purified monoclonal antibody (eBioscience);

[0043] 3. Mouse anti-human B7-H4 biotinylated monoclonal antibody (eBioscience);

[0044] 4. Streptavidin-peroxidase working solution (Abcam);

[0045] 5. Standard product: B7-H4 protein (R&D Systems);

[0046] 6. ABTS substrate: ABTS1mg+0.1M, pH5.0 citrate buffer 2ml+3%H2O24ul;

[0047] 7. Washing solution: 0.05% Tween20-PBS;

[0048] 8. Diluent: 0.1% BSA-PBS;

[0049] 9. Stop solution: 2M H2SO4.

[0050] The detection method of the kit includes:

[0051] Standard product preparation: Dissolve B7-H4 standard products with different contents in the diluent to reach concentrations of 0.1, 0.5, 2, 10, 25, 50, 100 ng / ml;

[0052] Coating and blocking: the mouse anti-human B7-H4 purified monoclonal antibody was diluted to 2ug / ...

Embodiment 3

[0058] Example 3. Sensitivity and specificity of the kit for detecting the concentration of B7-H4 in cerebrospinal fluid for diagnosing glioma

[0059] Collection of specimens: Firstly, 56 patients (5 cases of traumatic brain injury, 1 case of meningioma, 2 cases of metastatic tumor, 16 cases of low-grade glioma, 32 cases of high-grade glioma) were collected from the Department of Neurosurgery of Huashan Hospital from 2011 to 2012. After the consent of the tumor), lumbar puncture was performed on them, and 5ml of cerebrospinal fluid was collected from each person and stored in a -80°C refrigerator;

[0060] Detection of B7-H4 concentration: thaw the cerebrospinal fluid sample in an ice bath for 30-60 min, centrifuge at 1500 rpm×5 min, take the supernatant, follow the method in Example 2, use the kit, prepare standard products, coat and seal, Sample addition, primary antibody incubation, enzyme-labeled antibody incubation, color development, microplate reader detection and othe...

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Abstract

The invention belongs to the technical field of biology, and relates to a kit for tumor diagnosis, in particular to a kit for early diagnosis of glioma. The kit comprises a mouse anti-human B7-H4 biotin labeling monoclonal antibody and streptavidin-peroxidase working liquid, a collected cerebrospinal fluid sample is detected, and corresponding diagnosis can be made by detecting B7-H4 molecules in the cerebrospinal fluid. The kit can be used for quantitatively detecting the B7-H4 protein expression quantity in the cerebrospinal fluid, so that the glioma infection risk can be predicted, people at high risk can be screened, the early diagnosis of glioma can be realized, and glioma and other intracranial space occupying diseases can be distinguished. The kit has the characteristics that the sensitivity is 70.8 percent, the specificity is 100 percent, the sensitivity for distinguishing low-grade and top-grade glioma is 87.5 percent, and the specificity for distinguishing low-grade and top-grade glioma is 75.0 percent. The kit is free of cross reaction with other cell factors, so that the powerful technical support is provided for the accurate diagnosis and prevention of glioma.

Description

technical field [0001] The invention belongs to the field of biotechnology and relates to a tumor diagnosis kit, in particular to a novel kit for the early diagnosis of glioma. The kit can not only screen high-risk groups by detecting B7-H4 molecules in cerebrospinal fluid It can also be used to guide tumor staging, which provides a basis for finding sensitive and specific glioma cerebrospinal fluid markers. Background technique [0002] Studies have shown that among primary intracranial tumors, glioma has the highest incidence rate, close to 40.5%. Gliomas account for 20% of childhood tumors and are the leading cause of death in children with solid tumors. According to the survey, the peak incidence of glioma is between 30 and 40 years old, and it has the characteristics of short survival period, high recurrence rate and mortality rate. Especially for glioblastoma, the 2-year survival rate is 27.2%, and the 5-year survival rate is only 9.8%. Although the comprehensive tre...

Claims

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Application Information

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IPC IPC(8): G01N33/68
CPCG01N33/57407G01N2800/52G01N2800/7028
Inventor 周良辅毛颖姚瑜岳琪叶红星阿普拉吉塔
Owner AFFILIATED HUSN HOSPITAL OF FUDAN UNIV
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