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Chemiluminescent test kit for testing activity of MGMT (O6-Methylguanine DNA-Methyltransferase) and test method

A chemiluminescence detection and kit technology, used in measurement devices, scientific instruments, instruments, etc., can solve the problems of multi-step and time-consuming solid phase separation and reaction, false positives, etc., and achieve high specificity, simple components, and high ratio. The effect of surface area

Active Publication Date: 2015-05-20
上海羽帛科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The MGMT activity assay based on isotope-labeled substrates is only suitable for laboratory research and is difficult to be used in routine clinical tests; in addition, the protein precipitation step used in this method is easy to introduce non-specific binding of proteins such as melanin (Melanin), cause false positive
Although some people use non-isotope-labeled substrates and enzyme-linked immunoassay methods to measure MGMT activity, but this method requires multiple steps and time-consuming solid phase separation and reaction
This is an obvious disadvantage from the point of view of clinical application and detection efficiency (sensitivity)

Method used

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  • Chemiluminescent test kit for testing activity of MGMT (O6-Methylguanine DNA-Methyltransferase) and test method
  • Chemiluminescent test kit for testing activity of MGMT (O6-Methylguanine DNA-Methyltransferase) and test method
  • Chemiluminescent test kit for testing activity of MGMT (O6-Methylguanine DNA-Methyltransferase) and test method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1: Detection of MGMT activity in human glioma cell MGR-1 samples

[0042] The chemiluminescent ELISA kit for detecting MGMT activity in human glioma cell MGR-1 samples includes:

[0043] (1) Streptavidin magnetic beads: particle size 1~3μm, concentration 10mg / mL, 10mL / bottle, 1 bottle;

[0044] (2) Biotinylated O 6 -The substrate of methylguanine-DNA methyltransferase: the concentration is 5μg / mL, 5mL / bottle, 1 bottle; it is used after diluting 1000 times with diluent, which contains 1% BSA and 0.2g / L of sodium azide preservative solution.

[0045] (3)O 6 -Enzyme marker for methylguanine-DNA methyltransferase-specific antibody: the marker enzyme is alkaline phosphatase, the concentration is 5 μg / mL, 5 mL / bottle, 1 bottle; use after diluting 2000 times with diluent , the diluent is a thimerosal preservative solution containing 0.05% glycerin and 0.2g / L.

[0046] (4)O 6 -Methylguanine-DNA methyltransferase standard solution: the concentrations are 600, 150,...

Embodiment 2

[0069] Example 2: Detection of MGMT activity in human breast cancer tissue samples

[0070] Chemiluminescent ELISA kits for detecting MGMT activity in human breast cancer tissue samples include:

[0071] (1) Streptavidin magnetic beads: particle size 1~3μm, concentration 10mg / mL, 10mL / bottle, 1 bottle;

[0072] (2) Biotinylated O 6 -The substrate of methylguanine-DNA methyltransferase: the concentration is 5μg / mL, 5mL / bottle, 1 bottle; it is used after diluting 1000 times with diluent, which contains 1% BSA and 0.2g / L of sodium azide preservative solution.

[0073] (3)O 6 - Enzyme marker for methylguanine-DNA methyltransferase-specific antibody: the marker enzyme is horseradish peroxidase, the concentration is 5 μg / mL, 5 mL / bottle, 1 bottle; dilute 2000 times with diluent when used After use, the diluent is a thimerosal preservative solution containing 0.05% glycerin and 0.2g / L.

[0074] (4)O 6 -Methylguanine-DNA methyltransferase standard solution: the concentration is...

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Abstract

The invention provides a chemiluminescent test kit for testing activity of MGMT (O6-Methylguanine DNA-Methyltransferase), which comprises a streptavidin magnetic bead, a biotinylated O6-benzyl guanine, an enzyme label for MGMT specific antibody anti-MGMT, a standard substance solution of the MGMT, a luminescent substrate fluid and a concentrated cleaning solution. The invention also discloses a test method for applying the test kit to MGMT. The test method comprises the following steps: firstly, extracting total protein from a sample; testing by using the test kit; and lastly analyzing a test result. The invention provides the chemiluminescent test kit and the test method for testing the activity of MGMT in human tissues or cells; the chemiluminescent test kit and the test method have the advantages of convenience and high speed in operation, high flexibility of test result, strong specificity, and the like; and the chemiluminescent test kit and the test method have wide application prospects in the aspect of clinical test for the activity of MGMT.

Description

(1) Technical field [0001] The present invention detects O 6 - A chemiluminescent detection kit for methylguanine-DNA methyltransferase activity and a detection method thereof. (2) Background technology [0002] o 6 -Methylguanine-DNA methyltransferase (O 6 -Methylguanine DNA-Methyltransferase, MGMT) is an efficient DNA repair enzyme that can repair the damage caused by alkylating agents in chemotherapy to the DNA of tumor cells. If the MGMT activity in tumor cells reaches a certain level, it may lead to drug resistance Therefore, the level of MGMT activity provides direct information for drug resistance, and the detection of MGMT activity is of great significance for evaluating the effect of chemotherapy and reducing drug side effects. Unfortunately, there is no simple and reliable method that can be applied to routine clinical tests to measure the activity of MGMT, which has important tumor value. There have been a large number of literatures describing the methylation...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/577
Inventor 梁媛媛温汉华何睿焦艳华陈灿玉黄志坚
Owner 上海羽帛科技有限公司
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