Method for rapidly identifying thyroid papillary carcinoma lymphonodi cervicales metastasis in operation

A technology for lymph node metastasis and papillary carcinoma, which is applied in measurement devices, instruments, scientific instruments, etc., can solve the problems of low contrast between color strips and background, inability to achieve quantitative detection of detected objects, and difficulty in realizing multiple inspections and joint inspections, etc. Achieve the effect of reducing external conditions and background, shortening hospital stay, improving detection sensitivity and result confidence

Active Publication Date: 2019-09-24
JIANGSU INST OF NUCLEAR MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] (1) The colloidal gold labeling process is an electrostatic adsorption process, which is a physical adsorption, so the stability in the liquid phase is poor, often causing the labeled protein molecules to fall off again
[0007] (2) The test result is judged by displaying a single purple-red strip, and the color is single, so it is difficult to realize multiple inspection and joint inspection
[0008] (3) Only when the gold particles gather to a certain amount, the human naked eye can observe the purple-red band, and the color band has little contrast with the background, thus limiting the detection sensitivity
[0009] (4) The matrix effect of different materials is obvious, and the background interference is very large
[0010] (5) The detection sensitivity is low
[0011] (6) Quantitative detection cannot be realized
[0012] At present, there is also a detection method that uses quantum dots to label rapid immunochromatographic test strips, but this method cannot achieve quantitative detection of the test substance.
[0013] In addition, in the immunoassay methods for detecting Tg levels, there is still a lack of combinations that can be detected with high sensitivity and high specificity in double-antibody sandwich assays. Finding more suitable immunogenic Tg epitope peptides, preparing specific Sexual Tg antigens and antibodies are also the key points to be solved

Method used

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  • Method for rapidly identifying thyroid papillary carcinoma lymphonodi cervicales metastasis in operation
  • Method for rapidly identifying thyroid papillary carcinoma lymphonodi cervicales metastasis in operation
  • Method for rapidly identifying thyroid papillary carcinoma lymphonodi cervicales metastasis in operation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1: Screening of Human Tg Antigen Peptides

[0046] The Tg described in this application is known in the art, and the complete Tg is composed of a single polypeptide chain containing 2768 amino acids, with a molecular weight of about 660000 Daltons. Its amino acid sequence is known in the art and can be found in professional databases such as NCBI. The specific sequence is as follows:

[0047] Human Tg(1-2768):

[0048] MALVLEIFTLLASICWVSANIFEYQVDAQPLRPCELQRETAFLKQADYVPQCAED GSFQTVQCQNDGRSCWCVGANGSEVLGSRQPGRPVACLSFCQLQKQQILLSGYINSTDT SYLPQCQDSGDYAPVQCDVQQVQCWCVDAEGMEVYGTRQLGRPKRCPRSCEIRNRRLLH GVGDKSPPQCSAEGEFMPVQCKFVNTTDMMIFDLVHSYNRFPDAFVTFSSFQRRFPEVS GYCHCADSQGRELAETGLELLLDEIYDTIFAGLDLPSTFTETTLYRILQRRFLAVQSVISGRFRCPTKCEVERFTATSFGHPYVPSCRRNGDYQAVQCQTEGPCWCVDAQGKEMHGTR QQGEPPSCAEGQSCASERQQALSRLYFGTSGYFSQHDLFSSPEKRWASPRVARFATSCP PTIKELFVDSGLLRPMVEGQSQQFSVSENLLKEAIRAIFPSRGLARLALQFTTNPKRLQ QNLFGGKFLVNVGQFNLSGALGTRGTFNFSQFFQQLGLASFLNGGRQEDLAKPLSVGLD SNSSTGTPEAAKKDGTMN...

Embodiment 2

[0053] Embodiment 2: Preparation of TG antibody

[0054] The TG antigenic epitope peptides (1) and (2) obtained in Example 1 were linked to the carrier protein to prepare the antigens (1) and (2) for immunization respectively, and the animals were immunized with the obtained antigens (1) and (2), respectively, Antigen (1) is thus used to prepare specific monoclonal antibodies and polyclonal antibodies, and antigen (2) is used to prepare specific monoclonal antibodies and polyclonal antibodies.

[0055] 1. Antigen preparation: Tg peptides were linked with carrier protein BSA to prepare TG antigens. Take each 5.0 mg of the two antigenic peptides described in this application, dissolve them in 1 mL DMF, add 5 mg EDC (dissolved in 50 μL H 2 (0), after stirring for 20 min, the reaction was added dropwise to 5 mg BSA (dissolved in 1 mL of PBS buffer), immediately added 5 mg NHS, and stirred overnight at room temperature. Put the reaction solution into the treated dialysis bag, pla...

Embodiment 3

[0067] Example 3: Specific identification of human TG antibodies (1) and (2)

[0068] Detection was performed by ELISA. The ELISA plates were coated with human Tg protein, GAPDH protein, and neuron-specific enolase NSE as detection antigens, and the specific reactions of the prepared TG monoclonal antibodies (1) and (2) with different proteins were detected by ELISA , normal BALB / c mouse serum was used as negative control, and PBS solution was used as blank control.

[0069] Result: Tg monoclonal antibody (1) and (2) only react positively with Tg respectively (P / N>2.1), and are negative with GAPDH protein, neuron-specific enolase NSE reaction, illustrate and utilize the present invention Monoclonal antibodies (1) and (2) prepared from Tg epitope peptides (1) and (2) both have specificity.

[0070] Identification of polyclonal antibodies was performed using the same method as described above for identifying the specificity of monoclonal antibodies.

[0071] Result shows: Tg ...

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Abstract

The invention discloses fluorescence immunochromatographic test paper for rapidly detecting human thyroglobulin (Tg) so as to rapidly identify thyroid papillary carcinoma lymphonodi cervicales metastasis. According to the test paper, a double-antibody sandwich method and a fluorescence immunochromatography technology are adopted to detect human Tg; and antibodies are prepared through specific antigen epitope peptides. With the method adopted, human Tg in an object to be detected can be rapidly and accurately detected. The test paper has the advantages of simple, convenient and rapid operation, wide detection range, high specificity and high sensitivity.

Description

technical field [0001] The invention belongs to the field of immune detection, and in particular relates to a human thyroglobulin (Tg) epitope peptide, a Tg-specific antigen prepared by using the epitope peptide and a corresponding monoclonal antibody or polyclonal antibody, and the antibody is used in the preparation of human Use of the in vitro diagnostic kit for cervical lymph node metastasis of thyroid papillary carcinoma, a related in vitro diagnostic kit, and a fluorescent immunochromatographic test paper for rapidly identifying cervical lymph node metastasis of papillary thyroid carcinoma during operation and a preparation method thereof. Background technique [0002] Thyroid cancer is the most common malignant tumor among endocrine tumors and head and neck tumors, and its incidence is the fastest growing solid malignant tumor in the past 20 years. The rapid increase in the incidence of thyroid cancer is mainly attributed to the rapid increase in the incidence of diff...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/533G01N33/543G01N33/558
CPCG01N33/533G01N33/54313G01N33/558
Inventor 邹贤周彬李文新施龙顺包建东朱国华高芸王国瑞李秀龙杨克勤
Owner JIANGSU INST OF NUCLEAR MEDICINE
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