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Fusion protein for detecting rubella virus antibody and preparation method of fusion virus protein

A technology of fusion protein and rubella virus, which is applied in the field of genetic engineering technology and diagnostic reagents, can solve the problems of poor diagnostic sensitivity and specificity, difficulty in excluding cell protein components in cell culture antigens, and low antigen purity, so as to improve yield and specificity, The effect of improving the clinical missed detection rate

Inactive Publication Date: 2017-08-29
WEIFANG HIGHTOP BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The diagnosis of rubella virus infection is mainly based on serological diagnosis. The purification of cell culture antigens is difficult to exclude cell protein components. The low purity of antigens leads to poor diagnostic sensitivity and specificity. Therefore, screening highly sensitive and specific recombinant antigens is the key to developing rubella. The key to virus diagnosis

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] A fusion protein for detecting rubella virus antibody, the fusion protein is a fusion protein comprising rubella virus E1 protein fragment I, rubella virus E1 protein fragment II and rubella virus C protein fragment. Among them, E1 protein fragment I is distributed at the amino terminal of the fusion protein, E1 protein fragment II is distributed in the middle, and C protein fragment is distributed at the C-terminal of the fusion protein, and E1 protein fragment I, E1 protein fragment II and C protein fragment are connected by glycine - Glycine-serine linkage. Among them, the E1 protein fragment I is the 9th amino acid to the 116th amino acid in the E1 protein fragment, the E1 protein fragment II is the 208th amino acid to the 293rd amino acid in the E1 protein fragment, and the C protein fragment is the 212th amino acid in the C protein fragment Amino acids to amino acid 243.

Embodiment 2

[0036] A preparation method for detecting a fusion protein of rubella virus antibody, comprising the following steps:

[0037] (1) Screening of epitopes and chemical synthesis of gene fragments

[0038] Using online analysis tools such as ExPasy, TMHMM, SignalIP4.1, etc., the full-length amino acid sequences of rubella virus E1 protein and C protein were screened out, and the rubella virus E1 protein fragment I (the ninth one containing a strong antigenic determinant in the E1 protein was screened out) amino acid to the 116th amino acid), E1 protein fragment II (the 208th amino acid to the 293rd amino acid containing a strong epitope in the E1 protein), and the rubella virus C protein fragment (the 208th amino acid containing a strong epitope in the C protein 212 amino acids to the 243rd amino acid); chemically synthesized rubella virus E1 protein fragment I, E1 protein fragment II and C protein fragment tandem gene sequence;

[0039] (2) Construction of fusion protein recomb...

Embodiment 3

[0049] Application of Purified Recombinant Protein in Detecting Rubella Virus Antibody

[0050] The purified recombinant protein was diluted with carbonate buffer (50mM, pH 9.6) and coated with 100ul / well, 100ng / well protein on the enzyme-linked plate, overnight at 4°C. After washing, 200ul / well of 10% calf serum in PBS was added to block for 2 hours in a 37°C water bath, and washed at 4°C for later use.

[0051] Indirect enzyme-linked immunoassay was used to detect anti-rubella virus positive serum and normal human serum. The results showed that the fusion protein could react with anti-rubella virus positive serum but not with normal human serum, indicating that the fusion protein had good antigenicity and specificity.

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Abstract

The invention belongs to the technical field of biological detection and particularly provides fusion protein for detecting rubella virus antibody. The fusion protein includes rubella virus E1 protein fragment I, rubella virus E1 protein fragment II and rubella virus E1 protein fragment III. The invention further provides a preparation method of the fusion protein. The fusion protein has high sensitivity and specificity, and detection rate can be greatly improved by using the fusion protein to detect the rubella virus antibody.

Description

technical field [0001] The invention relates to the fields of genetic engineering technology and diagnostic reagents, in particular to a fusion protein for detecting rubella virus antibodies and a preparation method thereof. Background technique [0002] Rubella virus (rubella virus, RV) is the only member of the Rubellavirus genus in the Togaviridae family, and its only natural host is humans, but it can infect wild and experimental animals such as monkeys and mice in the laboratory. Rubella is a very mild disease that usually causes only a rash, swollen lymph nodes under the jaw, and other mild signs. But for children and pregnant women, the consequences of rubella can be serious. Infection during the first 12 weeks of pregnancy can lead to congenital rubella infection of the fetus and miscarriage in 80-90% of pregnant women. Congenital infection of the fetus can lead to congenital rubella syndrome, involving multiple organs and systems, and there is a long-term active i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/70
CPCC07K14/005C07K2319/00C12N15/70C12N2770/36222
Inventor 杨致亭高洪元丁兆明刘泽军王婷
Owner WEIFANG HIGHTOP BIOTECH
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