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HCV Nucleic acid aptamer and application thereof in preparing HCV-cAg detection kit

A nucleic acid aptamer and detection kit technology, which is applied in measuring devices, recombinant DNA technology, DNA/RNA fragments, etc., can solve the problems of inability to monitor the curative effect of hepatitis C patients, high technical and environmental requirements, and reduced sensitivity. Conducive to serological diagnosis and blood screening, improved sensitivity, and easy operation

Active Publication Date: 2012-10-03
山东莱博生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

[0003] The two main indicators currently used in the diagnosis of HCV infection are anti-HCV detection and HCV RNA detection. There is a "window period" of about 70 days in anti-HCV detection (that is, the period from the beginning of virus infection to the detection of the virus standard) ), this period has strong infectivity, and blood transfusion, injection, etc. during this period are very likely to cause HCV infection, thereby causing the spread of the disease. In addition, this method cannot be used for monitoring the efficacy of hepatitis C patients; and HCVRNA in The content in patients with advanced liver disease is very small and often cannot be detected, and the method has high technical and environmental requirements, and its application is greatly restricted
In 2001, Ortho Company launched the ELISA kit for detecting HCV core antigen. In 2005, Hunan Jingda Pharmaceutical Co., Ltd. released the HCV free core antigen detection kit. The kit is easy to operate and requires relatively simple personnel and instruments. When antibodies are produced, the antigens and antibodies in the serum form antigen-antibody complexes, and the sensitivity of detection will be greatly reduced

Method used

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  • HCV Nucleic acid aptamer and application thereof in preparing HCV-cAg detection kit
  • HCV Nucleic acid aptamer and application thereof in preparing HCV-cAg detection kit
  • HCV Nucleic acid aptamer and application thereof in preparing HCV-cAg detection kit

Examples

Experimental program
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Effect test

Embodiment 1

[0022] Example 1 Screening and determination of hepatitis C virus core antigen (HCV-cAg) nucleic acid aptamer

[0023] According to the published amino acid sequence of the hepatitis C virus core antigen protein (focusing on the 2-180 amino acids of the HCV core region), a random nucleotide library was designed by a conventional method, and the nucleic acid aptamer of the hepatitis C virus core antigen (HCV-cAg) was The screening and optimization of final determination of hepatitis C virus core antigen (HCV-cAg) nucleic acid aptamer oligonucleotide sequence is as follows (shown in SEQ ID No.1):

[0024] agcagcatga agtcatcgtt gatgttgctg ttatggtagc atgttgcagg tggtatcaac 60

[0025] ggagtagctg cacacatgtc acctactgca acc 93

[0026] Further, the above-mentioned hepatitis C virus core antigen (HCV-cAg) nucleic acid aptamer oligonucleotide molecule was analyzed with DNAsis software, and its secondary structure was predicted as figure 2 shown.

Embodiment 2

[0027] Example 2 Application of hepatitis C virus core antigen (HCV-cAg) nucleic acid aptamer in the preparation of HCV core antigen detection kit

[0028] The composition of the HCV core antigen detection kit:

[0029] 1 piece of anti-HCV-cAg monoclonal antibody ELISA plate for 96 people, 1 bottle of negative control, 1 bottle of positive control, 1 bottle of sample diluent, biotin-labeled HCV-cAg oligonucleotide aptamer (Bio-HCV- cAg aptamer) diluent 1 bottle, horseradish peroxidase-labeled avidin (Avi-HRP) diluent 1 bottle, washing solution 1 bottle, chromogenic reagent A 1 bottle, chromogenic reagent B 1 bottle, stop solution 1 bottle, 1 instruction manual, 3 self-adhesive seals.

[0030] Preparation and practical application of the above-mentioned hepatitis C virus core antigen detection kit:

[0031] 1. Select a polystyrene 96-well microtiter plate, coated with anti-HCV-cAg monoclonal antibody to make a detection microtiter plate:

[0032] The best preparation conditi...

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Abstract

The invention discloses a hepatitis C virus core antigen nucleic acid aptamer, wherein the nucleotide sequence of the aptamer is as indicated in SEQ ID No.1. The invention also discloses the application of the hepatitis C virus core antigen nucleic acid aptamer in preparing a hepatitis C virus (HCV) core antigen detection kit. The aptamer and the HCV core antigen detection kit prepared therefrom in the invention are capable of detecting the core antigen at pg / ml level; the sensitivity of the hepatitis C virus core antigen is greatly improved; therefore, the HCV nucleic acid aptamer and the HCV-cAg detection kit has important clinic diagnosis meaning and wide application prospect for early diagnosis of window-phase infected patients.

Description

technical field [0001] The present invention relates to a nucleic acid aptamer of hepatitis C virus and its application, in particular to a nucleic acid aptamer of hepatitis C virus core antigen (HCV-cAg) and its detection in the preparation of hepatitis C virus core antigen (HCV-cAg). application in the kit. Background technique [0002] Hepatitis C is caused by hepatitis C virus (Hepatitis C Virus, HCV) infection and has become a serious global health problem and health problem. HCV is a hepatotropic chronic virus, and the clinical symptoms of patients infected with HCV are extremely atypical, which easily leads to misdiagnosis. Compared with hepatitis B, hepatitis C is more prone to chronicity, early liver cirrhosis, liver cancer, and higher mortality. Therefore, the detection of HCV has great significance for the early diagnosis of hepatitis C virus infection and the guidance of clinical treatment. [0003] The two main indicators currently used in the diagnosis of HCV...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/115G01N33/577
Inventor 王佳颖朱之炜郑祖惠丁兴龙韩娟张增丽李夫东王恒江长林
Owner 山东莱博生物科技有限公司
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