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Kit for detecting enzyme activity of PEMT (phosphatidyl ethanolamine N-methyltransferase)

A technology of phosphatidylethanolamine and methyltransferase, which is applied in the fields of biomedicine, clinical diagnosis and detection, can solve the problem that the enzyme activity detection method is not easy to popularize and the like

Inactive Publication Date: 2016-06-22
北京中科卓明生物医学研究所有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] Therefore, based on the defects of the above-mentioned prior art, in order to solve the problem that the enzyme activity detection method of existing phosphatidylethanolamine N-methyltransferase (PEMT, PhosphatidylethanolamineN-methyltransferase, EC2.1.1.17) is not easy to popularize, the purpose of the present invention It is a combination product that provides a rapid, specific, accurate and reliable detection of phosphatidylethanolamine N-methyltransferase (PEMT, PhosphatidylethanolamineN-methyltransferase, EC2.1.1.17) enzyme activity

Method used

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  • Kit for detecting enzyme activity of PEMT (phosphatidyl ethanolamine N-methyltransferase)
  • Kit for detecting enzyme activity of PEMT (phosphatidyl ethanolamine N-methyltransferase)
  • Kit for detecting enzyme activity of PEMT (phosphatidyl ethanolamine N-methyltransferase)

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0085] Example 1: Crude Extraction of Rat Liver Endoplasmic Reticulum

[0086] Weigh 20 g of rat liver tissue from which adipose tissue has been excised, add 200 mL of 0.25 M (containing 0.01 M, pH 8 phosphate buffer) sucrose solution, and homogenize 20 times with a glass homogenizer. Sigma3-30K centrifuge at 2600 Centrifuge at 2800 rpm for 10 minutes, filter through four layers of gauze, and discard the precipitate. The supernatant was centrifuged at 15,000rpm for 30 minutes, and the resulting precipitate was the part containing the endoplasmic reticulum, which was suspended in 0.25M (containing 0.01M, pH8.0 phosphate buffer) sucrose solution, and the endoplasmic reticulum could be stored at -80°C Save it.

Embodiment 2

[0087] Embodiment 2: Assay of rat liver endoplasmic reticulum phosphatidylethanolamine N-methyltransferase enzyme activity (carry out pre-processing)

[0088] The PEMT detection reagent of the present embodiment is three doses of reagents, comprising

[0089] Reagent 1

[0090] Tris-hydrochloric acid buffer (pH8.5) 30mmol / L

[0091] SAM0.1mmol / L

[0092] Reagent 2

[0093] Tris-hydrochloric acid buffer (pH8.5) 30mmol / L

[0094] PE0.2mmol / L

[0095] Reagent 3

[0096] Potassium phosphate buffer (pH8.0) 30mmol / L

[0097] S-adenosyl-L-homocysteine ​​hydrolase 10KU / L

[0098] The PEMT sample to be tested is the extracted endoplasmic reticulum of rat liver tissue. After crushing, add sodium cholate solution to the sample to make the final volume percentage concentration reach 0.5%. After mixing gently, place it on ice for 30 minutes to wait. use. The total protein dosage in the reaction system is 10ug, and the reaction system is 250ul. A total of 5 parallel samples we...

Embodiment 3

[0105] Embodiment 3: Determination of enzyme activity of rat liver endoplasmic reticulum phosphatidylethanolamine N-methyltransferase (contrast sample This is compared with the previous treatment)

[0106] The phosphatidylethanolamine N-methyltransferase detection reagent of this embodiment is three reagents, including

[0107] Reagent 1

[0108] Tris-hydrochloric acid buffer (pH8.5) 30mmol / L

[0109] SAM0.1mmol / L

[0110] Reagent 2

[0111] Tris-hydrochloric acid buffer (pH8.5) 30mmol / L

[0112] PE0.2mmol / L

[0113] Reagent 3

[0114] Potassium phosphate buffer (pH8.0) 30mmol / L

[0115] S-adenosyl-L-homocysteine ​​hydrolase 10KU / L

[0116] The sample of phosphatidylethanolamine N-methyltransferase is the extracted endoplasmic reticulum of rat liver tissue. %, after mixing gently, place it on ice for 30min; add the same amount of sample suspension to another part, mix gently and place it on ice for 30min, the total protein dosage in the reaction system is 10ug, and...

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Abstract

The invention provides a combined product for detecting enzyme activity of PEMT (phosphatidyl ethanolamine N-methyltransferase). The combined product contains S-adenosine-L-methionine or an enzyme reaction system capable of producing S-adenosine-L-methionine, a methyl receptor and S-adenosine-L-homocysteine hydrolase, wherein the enzyme reaction system capable of producing S-adenosine-L-methionine comprises adenosine triphosphate, methionine and S-adenosylmethionine. Preferably, the combined product adopts a kit form, and all components are in self-existent reagent states. The combined product can be used on semi-automatic or full-automatic analysis detection equipment and is high in detection sensitivity, high in specificity and convenient to operate, so that the combined product can be popularized and used actually.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and at the same time belongs to the field of clinical diagnosis and detection. Specifically, the present invention provides a combination product for detecting phosphatidylethanolamine N-methyltransferase (PEMT, PhosphatidylethanolamineN-methyltransferase, EC2.1.1.17) enzyme activity. Background technique [0002] Choline is a strong organic base. It is a component of lecithin and also exists in sphingomyelin. important role in fetal development. Choline has an affinity for fat, which can make fat be transported from the liver through the blood in the form of phospholipids, thereby preventing the formation of fatty liver, effectively protecting and treating liver fibrosis caused by alcohol, and has the effect of anti-hepatic fat peroxidation, thereby reducing alcohol. The apoptosis of liver cells caused by choline can regulate the activity of liver cells, reduce the cytotoxicity of tumor ne...

Claims

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Application Information

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IPC IPC(8): C12Q1/48C12Q1/34
CPCC12Q1/48C12Q1/34G01N2333/91017G01N2333/914
Inventor 菅晓勇王天泽
Owner 北京中科卓明生物医学研究所有限公司
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