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Immunofluorescence reagent applied to detection of adenovirus IgM antibody and application of immunofluorescence reagent

A technology of adenovirus and antibody, applied in virus/bacteriophage, measuring device, and microorganism-based method, etc., can solve the problems of low sensitivity and low accuracy of IgM antibody, and achieve the effect of favorable price, high sensitivity and strong specificity

Inactive Publication Date: 2015-02-18
北京英诺特生物技术股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the colloidal gold method for detecting IgM antibodies has the disadvantage of low sensitivity and relatively low accuracy, so it is usually only used as an auxiliary diagnostic reagent

Method used

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  • Immunofluorescence reagent applied to detection of adenovirus IgM antibody and application of immunofluorescence reagent
  • Immunofluorescence reagent applied to detection of adenovirus IgM antibody and application of immunofluorescence reagent
  • Immunofluorescence reagent applied to detection of adenovirus IgM antibody and application of immunofluorescence reagent

Examples

Experimental program
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Effect test

Embodiment 1

[0021] The preparation of embodiment 1 adenovirus antigen

[0022] 1.1 Preparation of adenovirus seeds

[0023] The adenovirus type 55 used in this application was isolated from a throat swab of a respiratory patient in Taiyuan, Shanxi with A549 cells in 2011, amplified and sequenced by nucleic acid methods, and the gene sequence is GenBank accession number KF911353, such as SEQ ID No.1.

[0024] Specifically, the A549 cells (human lung adenocarcinoma cells) introduced from the Institute of Microbial Epidemiology, Academy of Military Medical Sciences were cultured with DMEM containing 10 wt% fetal bovine serum, and the well-grown monolayer cells were selected, and the culture medium was poured out and added Digest the cells with 0.25% trypsin, and when dots appear on the cells, pour off the digestion solution, add the cell growth solution, blow and beat the cells to completely disperse the cells, and count with moss blue, according to the number of cells 6×10 5 ~8×10 5 Each...

Embodiment 2

[0041]Preparation of Example 2 Adenovirus IgM Antibody Detection Kit (Indirect Immunofluorescence Method)

[0042] 2.1 Preparation of adenovirus antigen sheet

[0043] (1) Selection of the degree of infection of cells prepared by adenovirus antigen sheet

[0044] Take out the adenovirus seed from -70 ℃, and dilute the virus to 10 with DMEM (cell maintenance medium) containing 2% fetal bovine serum. -8 . Select A549 well-grown monolayer cells, pour off the cell growth medium, and inoculate 10 cells per bottle. -8 0.2 mL of virus, 37 °C CO 2 Adsorb in the incubator for 1 hour, add 6 mL of cell maintenance solution, and place in a 37°C CO2 incubator.

[0045] When the cytopathy was 15%, 25%, and 35%, respectively, the cells were digested with 0.25% trypsin. Pipette the cells with a certain amount of cell growth medium to prepare a cell suspension.

[0046] The above-mentioned three different degrees of cytopathic (15%, 25%, 35%) cell suspensions were dropped onto the 10-wel...

Embodiment 3

[0075] Performance verification of embodiment 3 adenovirus IgM antibody detection kit (indirect immunofluorescence method)

[0076] The adenovirus antigen prepared in Example 1 was used to prepare an antigen sheet according to the conditions of Example 2, and the ADV-IgM antibody was determined by indirect immunofluorescence.

[0077] 3.1 The principle and components of the kit are as follows:

[0078] (1) Kit principle: such as figure 1 As shown, this kit adopts the principle of indirect immunofluorescence (IFA). The adenovirus-specific IgM antibody in the sample to be tested reacts with the adenovirus antigen adsorbed on the slide, and the IgM immunoglobulin not bound to the antigen removed in the wash step. In the next step, the antigen-antibody complexes are reacted with fluorescein-labeled anti-human IgM globulin, and the results are visualized by immunofluorescence microscopy.

[0079] (2) The main components of the kit:

[0080] 1) Glass slide: 2 pieces (1×10 wells)...

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Abstract

The invention relates to an adenovirus antigen prepared by culturing an adenovirus strain with the preservation number of CGMCC No.9596. The invention also relates to an adenovirus IgM antibody detecting kit. The kit comprises the adenovirus antigen. Compared with the prior art, the diagnostic kit applied to preparation of the adenovirus antigen has the advantages of high specificity, high sensitivity, low cost and the like; the requirements on clinical diagnosis of adenovirus infection are well met.

Description

【Technical field】 [0001] The invention relates to the field of biological detection, in particular to an adenovirus antigen, a kit containing the antigen, a preparation method thereof, and an application of the kit in detecting adenovirus IgM. 【Background technique】 [0002] Adenovirus (adenovirus) is one of the main pathogens of viral pneumonia. Infection with adenovirus in older children or young adults only causes upper respiratory tract disease, and can cause severe acute adenoviral pneumonia in infants and young children. In 1958, Chang et al. first reported a case of type 7A adenovirus pneumonia, which was later reported in various countries. my country also began to study adenovirus infection in 1958. It was found that the pathogens causing adenovirus pneumonia in infants and young children are mainly adenovirus types 3 and 7. The prognosis of mild and older children is better, while severe infection and adenovirus pneumonia are less severe. Poor prognosis and high mo...

Claims

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Application Information

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IPC IPC(8): C12N7/00G01N33/569C12R1/93
Inventor 张晓刚吴纯赵建宏陈廷友吴丽霞华艳
Owner 北京英诺特生物技术股份有限公司
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