Immunity-chropadography test paper for detecting Yersinia pestis and preparation method thereof

An immunochromatographic test paper, the technology of Yersinia, applied in the field of biochemistry, can solve the problems of low sensitivity, long time, complicated detection methods, etc., and achieve the effect of simple sample processing

Inactive Publication Date: 2010-03-24
SHANGHAI HAITAI JINXIN BIOMOLECULAR DETECTION TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The technical problem to be solved by the present invention is to provide an immunochromatographic test paper for detecting Yersinia pestis and a preparation method thereof, the described immunochromatographic test paper for detecting Yersinia pestis requires Solve the technical problems of low sensitivity, complicated detection method and long time in the method for detecting Yersinia pestis in the prior art

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1 Preparation of antibodies against Yersinia pestis F1 antigen

[0018] Healthy big-eared white rabbits with a body weight of 2Kg were selected and injected subcutaneously with Freund’s complete adjuvant Yersinia pestis F1 antigen 2mg / only (purchased from Changchun Institute of Biological Products), and respectively on the 20th day after the initial immunization, On the 30th and 40th days, an additional shot of immunization solution was added. The dose and route were the same as the initial immunization. The antibody titer in the serum was tested 10 days after the last immunization, and the antibody titer was detected by the agar two-way diffusion method to reach 1:32 or more. Blood collection .

[0019] The anti-Yersinia pestis F1 antigen antibody in the blood was purified by saturated ammonium sulfate salting-out method, and the antibody obtained by agar two-way diffusion method had good specificity and affinity.

Embodiment 2

[0020] Embodiment 2 prepares immune colloidal gold probe

[0021] 1) Preparation of colloidal gold solution: prepare colloidal gold particles by citrate reduction method, the specific method is: HAuCl 4 Configure it as a 0.01% aqueous solution, take 100mL and heat it to boiling, and accurately add 1.6mL of 1% trisodium citrate (Na 3 C 8 h 5 o 7 2H 2 O) aqueous solution, the liquid color is stable into wine red, promptly obtains colloidal gold solution.

[0022] 2) Determine the saturation concentration of the colloidal gold-coupled antibody: use 0.2M K 2 CO 3 Or 0.1M HCl solution to adjust the pH value of the colloidal gold solution to 9.0, dilute the antibody against Yersinia pestis F1 antigen to a concentration of 1mg / mL, add 5μL, 10μL, 15μL, 20μL, 25μL, and 30μL to 1mL colloid In each tube of the gold solution, mix well and place it at room temperature for 5 minutes, add 0.1 mL of 10% NaCl aqueous solution, mix well, let stand, observe the color of the liquid after 1...

Embodiment 3

[0024] Example 3 Preparation of Immunochromatographic Test Paper for Detecting Yersinia pestis

[0025] 1) Dilute the quality control antibody (goat anti-rabbit IgG) with PBS at a concentration of 0.01M and pH7.2 to a final concentration of 2.5-3.0 mg / mL, and the anti-Yersinia pestis F1 antigen prepared in step 1 The antibody was diluted to a final concentration of 2.0-2.26mg / mL, and the diluted goat anti-rabbit IgG was sprayed on the NC membrane with a length of 300mm and a width of 25mm to form a control band with the XYZ3000 spotting instrument of BioDot Company, and the diluted anti-rabbit IgG The antibody of Yersinia pestis F1 antigen was sprayed on another area of ​​the NC membrane to form a test strip, dried at room temperature, and then an absorbent paper pad was pasted on the end of the NC membrane away from the test strip.

[0026] 2) Put the 300mm long and 10mm wide glass fiber membrane into the colloidal gold probe solution prepared in step 2 to obtain the gold sta...

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PUM

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Abstract

The invention provides an immunity-chropadography test paper for detecting Yersinia pestis, comprising a cellulose nitrate film, wherein a test belt and a comparison belt are arranged on the cellulosenitrate film and the Yersinia pestis F1 antigen antibody solution is sprayed on the test belt and the anti-antibody for resisting Yersinia pestis F1 antigen antibody is sprayed on the comparison beltand a water absorption pad is arranged on the cellulose nitrate film near to one side of the comparison belt and a gold-label pad is arranged on the cellulose nitrate film near to one side of the test belt and the immune colloidal gold probe solution for resisting the Yersinia pestis F1 antigen antibody is arranged in the gold-label pad and and a sample pad is arranged at one side of the gold-label pad apart from the test belt. The invention also provides a preparation method of the immunity-chropadography test paper for detecting Yersinia pestis. The immunity-chropadography test paper has same specificity and sensitivity with reverse heamagglutination and is simpler and more quick as an effective method for detecting plague epidemic and early detection.

Description

Technical field: [0001] The invention relates to the field of biochemistry, in particular to an immunochromatographic test paper, in particular to an immunochromatographic test paper for detecting Yersinia pestis and a preparation method thereof. Background technique: [0002] The pathogen of plague, Yersinia pestis (Yersinia pestis), is a well-known virulent pathogenic microorganism. It has the characteristics of easy acquisition, strong pathogenicity, high lethality, and strong resistance to the natural environment. People of all ages and ages are susceptible groups, and thus are recognized as classic biological warfare agents. After the "9.11" incident, the U.S. national anti-terrorism plan listed Yersinia pestis as a potent pathogenic microorganism in the bioterrorism red alert (the highest level), and the rapid detection of Yersinia pestis in the environment has now become an anti-biological Horror essentials. [0003] The classic detection procedure of Yersinia pesti...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/558G01N33/532
Inventor 葛海鹏
Owner SHANGHAI HAITAI JINXIN BIOMOLECULAR DETECTION TECH
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