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A method for determining the value of c-reactive protein in serum for the purpose of non-disease diagnosis

A technology for reactive protein and disease diagnosis, applied in disease diagnosis, instruments, analytical materials, etc., can solve problems such as high price, low total protein yield, time-consuming and labor-intensive

Active Publication Date: 2020-08-18
NAT INST OF METROLOGY CHINA
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the yield of isotope-labeled whole protein is low, time-consuming and labor-intensive, and the price is extremely expensive, and there are slight differences in the tertiary structure of natural proteins and their isotope-labeled analogs

Method used

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  • A method for determining the value of c-reactive protein in serum for the purpose of non-disease diagnosis
  • A method for determining the value of c-reactive protein in serum for the purpose of non-disease diagnosis
  • A method for determining the value of c-reactive protein in serum for the purpose of non-disease diagnosis

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Experimental program
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Embodiment Construction

[0037] Reagent:

[0038] Acetonitrile was purchased from German Merck company product;

[0039] Ultrapure water is purified by Millipore pure water system;

[0040] Trifluoroacetic acid was purchased from Sigma-Aldirich, USA;

[0041] Formic acid was purchased from Fisher Scientific, USA;

[0042] Dithiothreitol (DTT) was purchased from U.S. inalco company;

[0043] Trypsin was purchased from Promega, USA;

[0044] Tris, urea (Urea) were purchased from American Amresco company;

[0045] Iodoacetamide (Iodoacetamide, IAM) was purchased from U.S. Inalco Company;

[0046] Magnetic beads were purchased from Invitrogen;

[0047] EK, GK, RK, L-EK, L-GK, and L-RK were synthesized by Nanjing GenScript Biotechnology Co., Ltd. (using

[0048] MALDI-TOF-MS analyzes the digested CRP, selects three peptides EK, GK, and RK with high mass spectrometry response and can be separated; L-EK, L-GK, and L-RK are isotope labels for these three peptides peptides).

[0049] Human serum samples...

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Abstract

The invention discloses a value setting method for C reactive proteins in serum. The method comprises the following steps of (1) purification and enrichment of C reactive proteins (CRP)in serum; (2) purity detection of the C reactive proteins; (3) digestion; and (4) quantitative analysis of the C reactive proteins. The method does not need to consider the factors such as protein extraction rate, digestion efficiency and instrument stability, and only needs to consider the correctness of pure CRP concentration in the whole experiment; the used pure CRP solution is a national primary standard substance (developed by the National Institute of Measurement) which is reliable in numerical value and can be traced to an SI unit; and the raw material is nature CRP which does not need to consider the differences of structures and is more excellent than the isotope labeled whole proteins.

Description

technical field [0001] The invention relates to the technical field of protein content determination, in particular to a method for determining the value of C-reactive protein in serum. Background technique [0002] C-reactive protein is a pentameric serum protein released by the liver, which can be used to monitor cardiovascular disease and systemic inflammation, and is one of the important clinical markers. Because of its good stability and the commercialization of monoclonal antibodies, it has clinical application value. The commonly used detection methods include enzyme-linked immunosorbent assay, immune projection turbidimetry, and immunoluminescence. However, due to the development of methods and reagents for different targets by various manufacturers, the measurement deviation is very large. Therefore, it is urgent to develop a reference measurement method for CRP. [0003] Isotope dilution mass spectrometry has the characteristics of high sensitivity, high quantita...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/577G01N33/543G01N27/447G01N30/72
CPCG01N27/447G01N33/54326G01N33/577G01N33/6848G01N33/6893G01N2333/4737G01N2800/32G01N2800/7095
Inventor 宋德伟朱文陈鸿飞李红梅孙浩峰肖鹏刘建仪马凌云王馨雪
Owner NAT INST OF METROLOGY CHINA
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