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Kit for detecting thyroglobulin antibody and subtype thereof

A technology of thyroglobulin and kits, applied in the field of biomedicine, can solve the problems of not being included in clinical routine testing, narrow linear range, and toxic media

Active Publication Date: 2022-03-18
JIANGSU PROVINCE HOSPITAL THE FIRST AFFILIATED HOSPITAL WITH NANJING MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the specific distribution of TGAb IgG subtypes in different patients with Hashimoto's thyroiditis and the relationship between the subtype distribution and the pathophysiological process of Hashimoto's thyroiditis and the progress of thyroid function damage are few related reports at home and abroad. Its related tests have not yet been included in clinical routine tests, and related test kits have not yet been listed
[0004] At present, there are still few studies on TGAb IgG subtypes, and the detection method is mainly enzyme-linked immunoassay (ELISA), but the medium used in ELISA operation is toxic or carcinogenic, the sensitivity, repeatability and specificity are poor, and the linear range is narrow. and often lead to conflicting results

Method used

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  • Kit for detecting thyroglobulin antibody and subtype thereof
  • Kit for detecting thyroglobulin antibody and subtype thereof
  • Kit for detecting thyroglobulin antibody and subtype thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] 1. Detection of thyroglobulin antibody and its subtypes

[0029] 1. Determination of serum TGAb total IgG:

[0030] (1) Antigen dilution: Take a 15mL centrifuge tube and dilute with 6mL of pH 7.4 antigen buffer 125 I-TG to 20000CPM / 60μL;

[0031] (2) Incubate the labeled antigen with the serum overnight: Take a 96-well plate, add 5 μL of the serum to be tested in each well, and then add 60 μL of the diluted labeled antigen to each well. The CPM value of each well should be ≥20000. Each sample and The quality control serum is double-well, the labeled antigen and the serum are shaken and mixed at room temperature for 1 hour, and then overnight at 4°C;

[0032] (3) Incubate 96-well PVDF plate: Take a 96-well PVDF micropore filter plate, add 150 μL of antigen buffer to each well, and overnight at 4°C;

[0033] (4) Precipitate antigen-antibody complexes: Discard the incubation solution of the 96-well PVDF plate, add 25 μL of protein A / G-agarose to each well, and transfer ...

Embodiment 2

[0062] Comparison of agreement and correlation with electrochemiluminescence kits

[0063] 99 human serum samples were detected simultaneously with MODMLAR ANALYTICS E170 automatic electrochemiluminescence immunoassay system and supporting kits (Roche Diagnostics, Germany). The TGAb test results of 99 outpatients were classified into negative and positive results, as shown in Table 4.

[0064] Table 4 The results of two methods for measuring TGAb

[0065]

[0066] The kit of the present invention detects a positive coincidence rate of 100%, a negative coincidence rate of 79.2%, a total coincidence rate of 94.9%, and a Kappa value of 0.852, suggesting that the test results of the kit of the present invention have a high degree of consistency with the ECLIA method, and there is no significant difference Significance (paired chi-square test, P>0.05). Carry out Spearman correlation analysis to the TGAb measurement value in 80 parts of detection limit scope (10-4000IU / mL), it ...

Embodiment 3

[0068] Determination of each IgG subtype of TGAb

[0069] Detection of IgG subtypes of TGAb in serum of patients with Hashimoto's thyroiditis. According to the thyroid function status at the first diagnosis, they were divided into 3 groups: hypothyroid group, subclinical hypothyroid group (subclinical hypothyroid group), normal thyroid function group (normal thyroid function group).

[0070] figure 2 It is the distribution map of TGAb IgG subtype in the serum of patients with Hashimoto's thyroiditis (84 cases). figure 2 The results showed that in patients with Hashimoto's thyroiditis, there were statistical differences in the distribution of IgG subtypes of TGAb, and IgG1 was the main one (IgG1%: IgG2%: IgG3%: IgG4% = 42.42: 17.76: 18.44: 16.92).

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Abstract

The invention discloses a kit for detecting a thyroglobulin antibody and a subtype thereof, and belongs to the technical field of biological medicines. The kit comprises a < 125 > I labeled human thyroglobulin tracer agent, a streptavidin agarose gel reagent, a biotinylated anti-human IgG subtype monoclonal antibody, protein A agarose and protein G agarose. On the basis of an existing radioactive ligand detection method, core steps are replaced by combining upstream and downstream principles, the streptavidin agarose gel reagent and the biotinylated anti-human IgG subtype monoclonal antibody are introduced for the first time, a core technical platform is created again for subtype typing of the antibody, and the method has the advantages of high specificity, high sensitivity and high sensitivity. And then the biotinylated anti-human IgG subtype monoclonal antibody is combined with the TGAb IgG subtype, so that the detection of the TGAb IgG subtype is realized, and the upgrading of a technical platform and the breakthrough of a TGAb IgG subtype detection technology are successfully realized.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a kit for detecting thyroglobulin antibody and its subtype. Background technique [0002] Hashimoto thyroiditis (HT) is a very common autoimmune disease, with insidious onset, variable speed of progression, many atypical symptoms, complex and diverse clinical manifestations, and about 20% of the first diagnosis showed thyroid function 5% is mild hyperthyroidism, and most of them are euthyroid, but due to gland atrophy and destruction, they will eventually develop hypothyroidism. The probability of hypothyroidism in HT gradually increases with the prolongation of the disease course. Existing studies have shown that the incidence of hypothyroidism increases by 5%-7% per year. [0003] Thyroglobulin (TG) is a homodimeric glycoprotein secreted by thyroid follicular epithelial cells consisting of 5496 amino acids and a molecular weight of 660KD. TG is synthesized in ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/564G01N33/577G01N33/60
CPCG01N33/564G01N33/577G01N33/60G01N2800/046G01N2333/47
Inventor 陈恒顾愹陈逗逗王知笑郑旭琴蔡赟赵成程杨涛
Owner JIANGSU PROVINCE HOSPITAL THE FIRST AFFILIATED HOSPITAL WITH NANJING MEDICAL UNIV
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