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Method and device for detecting viral antigen of C type hepatitis

A technology for hepatitis C virus and antigen, which is applied in the field of hepatitis C virus detection, can solve the problems of difficult vaccine, difficult hepatitis C detection, and no protective effect, and achieves high specificity, good application prospects, and low false negatives. rate and false positive rate

Inactive Publication Date: 2004-11-17
KUNMING KELAISEN BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The reason for this many subtypes and serotypes is the polymorphism and variability of the hepatitis C virus gene. For example, the membrane protein (i.e. E protein) as the main antigenic component of the hepatitis C virus has a variability of up to 30%. This characteristic of hepatitis C virus determines its difficulty in vaccine research, that is, the immune response obtained by one type of hepatitis C virus infection basically has no protective effect on infection by other types of hepatitis C virus. However, it is difficult to make a meaningful diagnosis of another type of hepatitis C virus infection by using an immunodiagnostic reagent prepared by one type of hepatitis C virus
Also, HCV cannot so far grow on in vitro tissue cell cultures
Therefore, traditional vaccine development research is difficult to carry out, and vaccines developed using genetic engineering technology, mainly structural and non-structural peptides, are affected by genetic variability
Therefore, since the hepatitis C virus was first reported in 1989, there has been no vaccine available so far.
At the same time, only antibody detection reagents are available for its diagnostic reagents. This technical limitation makes the clinical diagnosis of hepatitis C virus infection only provide whether the patient has the experience of hepatitis C virus infection, that is, the existence of antibodies, but cannot confirm which period of time Whether there is hepatitis C virus replication and proliferation in the patient's body, that is, whether the antigen exists in the patient's body
These technical obstacles make the detection of hepatitis C caused by the hepatitis C virus more difficult

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Detecting hepatitis C virus antigen method, described method comprises the following steps: (1) antibody coating: I-hepatitis C virus-IgG; (2) the antigen in the sample is combined with coating antibody: I-hepatitis C virus -IgG-hepatitis C virus; (3) the hepatitis C virus antibody of enzyme label is combined with antigen: I-hepatitis C virus-IgG-hepatitis C virus-IgG-HRP; (4) add peroxidase ( HRP) substrate solution color development; (5) with 2mol / L of H 2 SO 4 Terminate the reaction; (6) microplate reader detection.

Embodiment 2

[0023] The preparation method of hepatitis C virus antigen detection kit:

[0024] (1) Antigen for immunization: The antigen used in the preparation of the antibody in the detection kit is a highly purified Hepatitis C virus recombinant gene containing 10 highly conserved regions of hepatitis C virus expressed by Escherichia coli using molecular biology techniques. Hepatitis virus multi-epitope complex antigen. The antigen has good immunogenicity and can induce animals to produce high-titer specific antibody responses against the hepatitis C virus. Due to the highly conserved nature of the immunogen, the prepared antibody can recognize different types of hepatitis C virus.

[0025] (2) Preparation of specific antibody: use clean grade Japanese white rabbits, weighing 1.8-2.0Kg, inject 1 mg / ml compound antigen and the same amount of incomplete Freund's adjuvant, fully emulsify and mix, and inject into muscles at multiple points. The immunization procedure was 3 times in 0, 1,...

Embodiment 3

[0031] Sensitivity and specificity test of hepatitis C virus antigen detection kit: based on the review of hepatitis C virus-RNA detection. Detect the negative and positive samples of hepatitis C antigen (HCAg) with the kit described in Example 2, extract hepatitis C virus-RNA with a small amount of virus RNA / DNA extraction kit, and then use RT-PCR detection kit to carry out HCV-RNA detection to determine its sensitivity (true positive rate), specificity (true negative rate), false positive rate, false negative rate, crude consistency, adjusted consistency, Youden index, positive predictive value and Negative predictive value. A total of 400 samples were compared, and the results are shown in Table 1.

[0032] Table 1.

[0033] RT-PCR method detection results

[0034] Kit HCAg test results Total

[0035] Positive Negative

[0036] Positive 34 3 37

[0037] Female 5 258 363

[0038] Total 39 361 400

[0039] in:

[...

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Abstract

The invention discloses a method and device for detecting virus antigen of type-C hepatitis. The invention designs a compound antigen for type-C hepatitis virus made up of ten specificity antigens in epi-position, the reagent for detecting the type-C hepatitis virus antigen constructed by the inducer and by using the antigens, it can identifies the type-C hepatitis antigen in serum and plasma infected by the type-C hepatitis virus through the detection. The reagent box has a high specificity and low false negative rate and false positive rate.

Description

technical field [0001] The invention relates to a detection method of virus antigen and a device for realizing the method. The methods and devices of the invention are particularly suitable for the detection of hepatitis C virus. Background technique [0002] Hepatitis C virus (HCV) is a human hepatitis virus identified after hepatitis A virus (HAV) and hepatitis B virus (HBV). Like the previous two viruses, it is a pathogen that causes infectious hepatitis in humans. The analysis of etiology, epidemiology and epidemiology of hepatitis C virus has shown that its infection often leads to chronic persistent hepatitis, and about 50% of patients with chronic hepatitis may develop liver cirrhosis and liver cancer. Therefore, it is another important infectious disease faced by human beings after the viral infection caused by HAV and HBV has been effectively controlled by vaccines. The treatment of hepatitis C for more than ten years shows that there is n...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/535G01N33/576
Inventor 谢忠平董少忠王群董少华李琦涵
Owner KUNMING KELAISEN BIOLOGICAL TECH
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