Human milk transferrin and application of detection kit thereof in nasopharyngeal carcinoma diagnosis

A lactotransferrin and detection kit technology, applied in the field of lactotransferrin detection kits, can solve the problem of no sensitive and effective early and non-invasive detection methods for nasopharyngeal carcinoma, and achieve great economic and social value and high Sensitivity, fast detection effect

Inactive Publication Date: 2008-07-30
CENT SOUTH UNIV
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0005] Another object of the present invention is to propose the application of the human lactotransferrin ELISA detection kit in the early diagnosis, screening and risk prediction of nasopharyngeal carcinoma, so as to solve the early and non-invasive method that is not sensitive and effective for nasopharyngeal carcinoma at present. check method problem

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  • Human milk transferrin and application of detection kit thereof in nasopharyngeal carcinoma diagnosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Embodiment 1: the ELISA kit of preparation LTF

[0025] (1) Preparation of LTF antibody

[0026] 1. Preparation of mouse anti-human LTF protein polypeptide antibody

[0027] (1) Input the amino acid sequence of LTF into DNAStar software to obtain a polypeptide of 21 amino acids with the strongest immunogenicity of the protein. The sequence of the polypeptide is: NYKSQQSSPDDPNCCVDRPVEG. The polypeptide is chemically synthesized, and the polypeptide is combined with hemocyanin ( Keyhole limpet hemocyanin, KLH) was cross-linked to increase immunogenicity.

[0028] (2) Dissolve the polypeptide in phosphate buffered saline (PBS) and inject it into mice. The first dose is 300ug / kg, and the booster dose is about 1 / 4 of the first dose. Booster immunization every 2 to 3 weeks, a total of 4 immunizations, and then take mouse spleen B lymphocytes to fuse with myeloma cells, and carry out selective culture in HAT medium. After clonal selection, specific monoclonal antibodies can ...

Embodiment 2

[0052] Example 2: The steps of using the ELISA kit of LTF and the detection of specificity and stability

[0053] Take the nasopharyngeal secretion to be tested. The specific method is: after washing the nasal cavity with normal saline, insert a thin cotton swab into the nasopharynx (the surface of the suspected tumor) under the rhinoscope, leave it for 5-8 minutes, and quickly take out the cotton swab. Put the cotton ball into a small centrifuge tube with a hollow bottom, cover the centrifuge tube with a slightly larger centrifuge collection tube, and centrifuge at 7500Xg for 1 minute at 4°C to obtain nasopharyngeal secretions. This method is non-invasive and can be generally accepted.

[0054] Purify and quantify the LTF gene recombinant protein as a standard, dilute the antigen (nasopharyngeal secretion) at 1:10, add it to the previously coated microtiter plate, incubate at 37°C for 2 hours, wash the plate with PBST Unbound antigen was washed away and residual liquid was b...

Embodiment 3

[0059] Embodiment 3: the clinical application of the ELISA kit of LTF

[0060] The nasopharynx of a patient suspected of nasopharyngeal carcinoma was rinsed with normal saline, and 40ul of the patient's nasopharyngeal secretions were collected according to the above method, and a pathological biopsy was performed on the patient at the same time. Dilute the obtained nasopharyngeal secretions to 400ul with PBS, and begin to use this kit to quantitatively detect the LTF protein in the secretions;

[0061] Take out the coated ELISA plate from the refrigerator, place it at room temperature for 20 minutes, and dilute the standard at the following concentrations: 12.8ug / ul, 6.4ug / ml, 3.2ug / ml, 1.6ug / ml, 0.8ug / ml ml, 0.4ug / ml, 0.2ug / ml, 0ug / ml (blank control), take 100ul of each diluted standard product according to the order in Table 2 (the 96 cells in the table below are the 96 cells in the microtiter plate) hole) was added to the microtiter plate, repeated 3 times;

[0062] Add 5...

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Abstract

The invention discloses the application of human lactoferrin (hLTF) and its ELISA detection kit in diagnosis of nasopharyngeal carcinoma. The invention proves the specific down-regulation of hLTF gene in nasopharyngeal carcinoma tissue by experiments so as to provide the use of hLTF in diagnosis, screening and prognosis of nasopharyngeal carcinoma. The invention further provides an anti-hLTF antibody and an hLTF ELISA detection kit for diagnosis of nasopharyngeal carcinoma by double-antibody sandwich method. The expression level of LTF in nasopharyngeal secretion from different people can be quantitatively detected by collecting secretion from nasopharynx with ELISA method, so as to predict the risk of nasopharyngeal carcinoma, screen susceptible persons, and perform early and rapid non-invasive diagnosis to nasopharyngeal carcinoma patients.

Description

technical field [0001] The invention relates to the application of lactotransferrin in early diagnosis of nasopharyngeal carcinoma and a lactotransferrin detection kit for early diagnosis of nasopharyngeal carcinoma. Background technique [0002] Nasopharyngeal carcinoma (Nasopharyngeal Carcinoma, NPC) is a common malignant tumor in southern my country. It is a carcinoma of the nasopharyngeal mucosa epithelium. Most of them are poorly differentiated squamous cell carcinomas. Those with crypts and the top of the nasopharynx have no obvious early symptoms, so it is difficult to detect early, and the rate of misdiagnosis and treatment is high. The five-year survival rate of nasopharyngeal carcinoma has been hovering around 50-60% in recent years, and the five-year survival rate of early-diagnosed nasopharyngeal carcinoma patients can reach more than 90%. Therefore, screening and early diagnosis of high-risk groups for nasopharyngeal carcinoma, In order to carry out early treatm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/574G01N33/531C12N5/18
Inventor 李桂源熊炜曾朝阳周艳宏张文玲李小玲周厚德范松青肖岚周鸣沈守荣
Owner CENT SOUTH UNIV
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