Reagent kit for LgG immunoblotting diagnosis of SARS coronavirus antibody

A diagnostic kit, coronavirus technology, applied in the direction of measuring devices, instruments, scientific instruments, etc., can solve problems such as inappropriate rapid detection of diseases, incorrect test results, complicated experimental techniques, etc., to achieve suitable mass production, high sensitivity, Ease of use

Inactive Publication Date: 2003-11-05
GUANGZHOU WONDFO BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] ②The test result is incorrect (false negative)
[0027] Cell culture is a kind of aseptic operation technology, which requires high experimental working environment and conditions (aseptic operation room, ultra-clean workbench, operation room, etc.), and requires a large number of instruments and equipment (incubator, centrifuge, microscope, centrifuge, etc.) Machines, etc.), requires a large number of professional experimental equipment (various culture vessels), the experimental technology is complex, requires experienced professional and technical personnel to operate, and the experimental cycle is very long (several months), not suitable for rapid detection of diseases

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] In this embodiment, the purified SARS coronavirus whole virus lysate antigen is electrophoresed and transferred to a nitrocellulose membrane, and an anti-human IgG line is also used as a quality control line on the membrane to check whether the membrane strip fails. Horseradish peroxidase (HRP)-labeled rabbit anti-human IgG and corresponding chromogenic substrates were used for detection. The manufacturing method of the present embodiment is as follows: a. Antigen preparation Purified SARS coronavirus whole virus antigen uses 0.02M pH7.6 PBS to regulate protein concentration to 10mg / ml. The lysate was added at a volume of 1:2, and lysed at 100°C for 5 minutes. b. Antigen separation (gel electrophoresis)

[0046] 1. Prepare 12% polyacrylamide gel as separating gel and 2.5% polyacrylamide gel as stacking gel.

[0047] 2. Exhaust under negative pressure for 5 minutes.

[0048] 3. Pour plate: first pour the separation glue, the area is 15×15cm 2 , after the separating g...

Embodiment 2

[0066] In this embodiment, the purified SARS coronavirus whole virus lysate antigen is electrophoresed and transferred to a nitrocellulose membrane, and an anti-human IgG line is also used as a quality control line on the membrane to check whether the membrane strip fails. Alkaline phosphatase (ALP)-labeled goat-anti-human IgG antibody and corresponding chromogenic substrate were used for detection. The manufacturing method of this embodiment is basically the same as that of Embodiment 1, the differences are as follows:

[0067] a. Preparation of enzyme-labeled antibody

[0068] Choose imported high-purity alkaline phosphatase (ALP)-labeled goat-anti-human IgG antibody 1mg, according to the drop-mixing results, dilute to the planned volume with 0.02M PBS containing 1.0% BSA, pH7.4, and store at 4°C.

[0069] b. Chromogenic substrate:

[0070] Tris 12.1g

[0071] MgCl 2 0.475g

[0072] NaCl 5.8g

[0073] BCIP 0.4g

[0074] ...

Embodiment 3

[0079] In this example, the purified SARS coronavirus recombinant antigen mixture was electrophoresed and transferred to a nitrocellulose membrane, and an anti-human IgG line was also used as a quality control line on the membrane to check whether the membrane strip failed. Horseradish peroxidase (HRP)-labeled rabbit anti-human IgG and corresponding chromogenic substrates were used for detection. The manufacturing method of this example is the same as that of Example 1 as follows.

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Abstract

The present invention relates reagent kit for IgG immunoblotting diagnosis of SARS coronavirus antibody. The reagent kit consists of solid film, antiantibody of enzyme label and corresponding chromogenic substrate. The solid film is obtained through purifying SARS coronary totivirus lysate liquid antibody or recombining mixed antigen, gel electrophoresis to separate and transferring to film. The antiantibody of enzyme label is compounded with rabbet's antihuman IgG labeled with horseradish peroxidase or sheep's antihuman IgG labeled with alkaline phosphatase, 0.01 % concentration thimerosal as protecting agent, ox serum protein in 1 wt% and PBS in pH 7.4 in 0.01 M. The present invention may be used in detecting several kinds of antigen and antibody in clinical test department and sanitary epidemic prevention department.

Description

technical field [0001] The invention relates to a kit for detecting human anti-SARS coronavirus antibody (IgG) by immunoblotting method, and carrying out epidemiological investigation and clinical diagnosis of atypical pneumonia. Background technique [0002] Atypical pneumonia (also known as Severe Acute Respiratory Syndrome, Severe Acute Respiratory Syndrome, abbreviated as SARS) has been raging around the world for nearly five months. On April 16, the World Health Organization officially announced a new type of coronavirus that has never been found in humans before. It is the pathogen of SARS. Coronavirus (Corona Virus) was discovered by Almeida et al. in 1968. It was named "coronavirus" because of its shape. Under the electron microscope, the virus envelope has spikes similar to the corona. Coronaviruses are widely classified and can infect humans, poultry and livestock, and can cause poultry infectious bronchitis, mouse hepatitis, porcine encephalomyelitis, feline infe...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/535G01N33/566G01N33/569
Inventor 王继华
Owner GUANGZHOU WONDFO BIOTECH
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