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Serum polypeptide combined marker, detection kit and method for early warning and/or diagnosing diabetes

A technology for markers and diabetes, applied in biological testing, chemical instruments and methods, disease diagnosis, etc., can solve the problems of low specificity, lack of sensitivity of markers, etc., and achieve high diagnostic specificity, high sensitivity, and good stability Effect

Active Publication Date: 2021-06-08
DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to solve the problems of lack of markers for early warning and diagnosis of diabetes, low sensitivity and low specificity, and to provide a combination of polypeptide markers by using non-targeted peptidomics to discover differential polypeptides and targeted peptidomics verification. Its application and kit in early warning / diagnosis of diabetes, and also provides a method for the quantitative analysis and detection of the above-mentioned combined polypeptide markers

Method used

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  • Serum polypeptide combined marker, detection kit and method for early warning and/or diagnosing diabetes
  • Serum polypeptide combined marker, detection kit and method for early warning and/or diagnosing diabetes
  • Serum polypeptide combined marker, detection kit and method for early warning and/or diagnosing diabetes

Examples

Experimental program
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Effect test

Embodiment 1

[0045] Example 1: Screening of Differential Polypeptides by Standard-Free Quantitative Method

[0046] Clinical samples: 23 human serum samples (including 7 healthy subjects, 7 prediabetic patients and 9 type 2 diabetic patients) were obtained from Shanghai Sixth People's Hospital. Patients with prediabetes and type 2 diabetes were diagnosed or excluded by experts, and the three groups of subjects were matched for age, body mass index (BMI), blood sugar and other indicators. Blood samples were approved by the ethics committee of the local hospital, and all subjects signed informed consent. Blood samples were stored in a -80°C refrigerator after collection.

[0047] Extraction of serum endogenous polypeptides: Ultrafiltration and centrifugation. Specific operation procedure: Take out the blood sample from the -80°C refrigerator and thaw it in an ice bath, vortex and mix well, and centrifuge at 20,000g for 5 minutes to discard lipids. Take 25 μL of each serum sample, add 125 ...

Embodiment 2

[0055] Example 2: Dimethyl-labeled polypeptide and targeted MRM validation

[0056] Clinical samples: 18 human serum samples (including 9 serum samples from healthy subjects and 9 serum samples from patients with type 2 diabetes) were obtained from Shanghai Sixth People's Hospital. Patients with type 2 diabetes were diagnosed or excluded by experts, and the age, body mass index (BMI), blood sugar and other indicators of the three groups of subjects were matched. None of the diabetic patients had complications such as hypertension and hyperglycemia. Blood samples were approved by the ethics committee of the local hospital, and all subjects signed informed consent. Blood samples were stored in a -80°C refrigerator after collection.

[0057] Extraction of serum endogenous polypeptide: same as Example 1.

[0058] Synthesis of peptides: According to the information in Table 3 above, the structure of the synthesized standard peptides was reconfirmed by comparing their MS spectra....

Embodiment 3

[0080] Example 3: Determination of early warning / diagnostic markers

[0081] From the above quantitative results, it can be seen that in the two groups of samples of healthy people and diabetic patients, peptides GVLSSRQLGLGPGPDVPDHAAYHPF (245), GVLSSRQLGLGPGPDVPDHAA (244), GLPGPPDVPDHAAYHPF (251), SRQLGLGPGPPDVPDHAA (256), DIQMTQSPSSVSASVGD (254), DEAGSEADHEGFF (252), GVND 248), the content of GSEMVVAGKLQ (261) changed significantly.

[0082] Using SPSS statistical software, through binary logistic regression analysis, 8 polypeptides were regressed as combined marker variables, and the ROC (receiver operating characteristic) curve was used to evaluate the sensitivity and specificity of the combined markers ( figure 2 ). Candidate markers were identified with an AUC greater than 0.8.

[0083] The 8 polypeptides are regressed as the combined marker variable X, and the binary logistic regression equation is:

[0084] X=712.913-4.476×C(245)+1704×C(244)-2637×(251)-94.36×C(256)...

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Abstract

The invention relates to the field of medical biological detection, in particular to novel application of a polypeptide combined marker in preparation of a diabetes diagnosis reagent or kit. The invention provides a method and a kit for quantitatively detecting endogenous polypeptides in serum by using liquid chromatography-mass spectrometry multiple reaction monitoring (MRM). The reagent and the method can be quantitatively detecting the contents of eight endogenous polypeptide markers in human serum, and calculating the combined marker variable and the determined intercept point value based on a binary logistic regression equation for early warning and / or diagnosis of diabetes mellitus. The kit and the detection method disclosed by the invention are good in stability and high in specificity, and have clinical popularization potential.

Description

technical field [0001] The invention relates to the field of medical biological detection technology and the diagnosis, prevention and treatment of metabolic diseases, specifically, it is based on the method of quantitatively measuring endogenous polypeptides in human plasma and the combination markers of polypeptides in the preparation of reagents for early warning and / or diagnosis of diabetes boxes and methods. Background technique [0002] According to the data of the International Diabetes Federation in 2017, there are 415 million diabetic patients worldwide, of which type 2 diabetes (type 2diabetes, T2DM) accounts for more than 95%. It is estimated that by 2030, the number of people with diabetes worldwide will increase to 552 million. Diabetes is the third major disease that endangers human health after cardiovascular disease and tumors. At present, although the diagnostic criteria for type 2 diabetes are clear, and there are gold standards such as oral glucose toler...

Claims

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Application Information

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IPC IPC(8): G01N33/68C07K14/47C07K7/08C07K7/06
CPCG01N33/6848G01N33/6893C07K14/47C07K7/06C07K7/08G01N2800/042G01N2800/50
Inventor 吴仁安王莉牛欢
Owner DALIAN INST OF CHEM PHYSICS CHINESE ACAD OF SCI
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