ELISA-Array method for detection of encephalitis viruses, and special kit thereof

A technology of Japanese encephalitis virus and tick-borne encephalitis virus, which is applied in the field of ELISA-Array method and special kit for detecting encephalitis-like viruses, and can solve the problems of infectious disease detection

Inactive Publication Date: 2012-04-25
MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, only ELISA-Array technology has been used in the research reports of tumor

Method used

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  • ELISA-Array method for detection of encephalitis viruses, and special kit thereof
  • ELISA-Array method for detection of encephalitis viruses, and special kit thereof
  • ELISA-Array method for detection of encephalitis viruses, and special kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0070] The conditions of embodiment 1, ELISA-Array are explored

[0071] 1. Optimization of antibody spotting conditions

[0072] 1. Preparation and purification of monoclonal antibodies

[0073] Monoclonal antibodies 4D5, 2B5, 1F1, 2B8, 4F9, 4E11 and 2A10 (all recorded in Application of monoclonal antibody 2A10 in the detection of flaviviruses by protein chip technology, Sun Tingting, Li Yuchang, Liu Hong, Kang Xiaoping, Lin Fang, Zhu Qingyu, Yang Yinhui, Lu Cheng, Chinese Journal of Microbiology and Immunology, 2010, Volume 30, No. 8, 775-778, available to the public from the Institute of Microbial Epidemiology, Academy of Military Medical Sciences of the Chinese People's Liberation Army. Dengue virus monoclonal antibody serum Research on the Nature of Science, Liu Jinglan, Chen Wanrong, Cui Zhenmin, Xin Yanbin, Yang Baoan, Xu Pinfang, Zhu Qingyu, Yan Guozhen, Chinese Journal of Immunology, Issue 02, 1985, pages 3-6, available to the public from Research on Microbial Epidem...

Embodiment 2

[0136] Embodiment 2, the detection of multiple virus mixtures

[0137] In order to determine whether the ELISA-Array technology can be used for the detection of mixed infection viruses, take an appropriate amount of JEV, TBEV, DV2, EEEV, DV4 and SV virus culture supernatant after mixing two or three viruses (in the detection of multiple virus mixtures, The amount of each virus used is 4 times diluted virus culture supernatant), according to the ELISA-Array technology detection method of step 2 of Example 1, add it to the microwell plate for detection,

[0138] The result is as Figure 5 As shown, A is DV-2 and EEEV; B is TBEV and DV2; C is TBEV and EEEV; D is JEV, TBEV and DV-4; E is DV-2, DV-4 and TBEV, using array 3 , as shown in Table 3 and Table 4. It can be seen that all the antibodies targeted by the tested pathogens showed positive signals, indicating that this technology can be used for the detection of mixed infections of various pathogens.

Embodiment 3

[0139] Embodiment 3, the detection of clinical sample inoculation chicken embryo culture

[0140]In order to verify the detection effect of ELISA-Array technology on clinical samples, sera from 3 positive patients with TBEV infection (Mudanjiang Forestry Central Hospital, the patients were informed, and the 3 sera had been tested positive for TBEV infection by immunofluorescence) were inoculated with chicken embryo urine. The cyst cavity was cultured in a 37°C incubator for 3 days, and the liquid in the allantoic cavity was collected. After the virus was inactivated with β-propiolactone, TBEV tick-borne encephalitis virus samples 1, 2, and 3 were obtained, and real-time quantitative RT was performed respectively. -PCR detection (upstream primer 5-GGG CGG TTC TTG TTC TCC-3, downstream primer 5-ACA CAT CAC CTC CTT GTCAGA CT-3, probe sequence 5-FAM-TGA GCC ACC ATC ACC CAG ACA CA-TAMRA-3 , the template is the RNA extract of TBEV tick-borne encephalitis virus sample 1,2,3) and dete...

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Abstract

The present invention discloses an ELISA-Array method for detection of encephalitis viruses, and a special kit thereof. The kit of the present invention comprises six capture antibodies, wherein the solution of each capture antibody is a mixing solution prepared by mixing the capture antibody and a spotting solution, the concentration of each capture antibody in the corresponding capture antibody solution is 0.05 mg/ml. Experimental results of the present invention show that: the six specific encephalitis virus monoclonal antibodies are prepared by the method of the present invention; with adopting the ELISA-Array technology platform, and optimizing the experimental conditions, the ELISA-Array technology capable of concurrent detection of the six encephalitis viruses is established, and can be adopted for detection of the six encephalitis-related viruses; compared to the common ELISA, the ELISA-Array method of the present invention has the following advantages that: the specificity of the ELISA-Array method is the same as the specificity of the common ELISA, the sensitivity is high, the highest sensitivity is more than 10 times the sensitivity of the common ELISA, and the ELISA-Array method has a high clinical application prospect.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an ELISA-Array method for detecting encephalitis-like viruses and a special kit thereof. Background technique [0002] my country is a country where encephalitis-like viruses are seriously harmful. Correct diagnosis of pathogenic bacteria is the basis for effective control of its spread and the key to effective treatment. The encephalitis-like viral pathogens prevalent in my country mainly include: Japanese Encephalitis virus (JEV), Tick borne encephalitis virus (TBE), Eastern equine encephalitis virus (Eastern equine encephalitis virus) Encephalitis virus (EEEV), Sindbis Virus (Sindbis Virus), Dengue virus (Dengue virus, DV), etc., are all insect-borne viruses, of which Japanese Encephalitis virus (JEV), tick Tick ​​borne encephalitis virus (TBE) and Eastern equine encephalitis virus (EEEV) can cause severe encephalitis symptoms and have a high fatality rate, Sindbis virus (Sindbis V...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/569G01N33/543G01N21/76
CPCY02A50/30
Inventor 康晓平李裕昌杨银辉林方范丽魏婧靖户义李靖常国辉祝庆余
Owner MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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