Immunochromatographic test paper for testing abrin and preparation method thereof

An immunochromatographic test paper and abrinia toxin technology, applied in biological testing, measuring devices, analytical materials, etc., can solve problems such as unsuitable for on-site detection, cumbersome operation process, radioactive pollution, etc., easy to achieve results, simple and convenient to operate, highly specific effect

Inactive Publication Date: 2010-06-16
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Its disadvantage is that there is a problem of radioactive pollution
ELISA method for detection of abrin toxin has high sensitivity and...

Method used

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  • Immunochromatographic test paper for testing abrin and preparation method thereof
  • Immunochromatographic test paper for testing abrin and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Preparation and identification of abrin toxin antibody

[0030] 1. Preparation of abrin toxin

[0031] The crude toxin was extracted with 5% acetic acid, and the precipitate was discarded after high-speed centrifugation (10,000g, 20min). The extract was further separated by Sepharose 6B or Sepharose 4B agarose affinity chromatography column, DEAE-Sepharose FF anion exchange column and Hiload 26 / 60Superdex75 gel filtration chromatography prepacked column, and the molecular weight was analyzed by SDS-PAGE electrophoresis. The purity was determined by gel TLC scanning, N-terminal amino acid sequence determination and mass spectrum peptide fingerprint analysis and identification. The results showed that high-purity abrin toxin was prepared.

[0032] 2. Preparation of mouse monoclonal antibody specifically binding to abrin toxin A chain

[0033]After abrin toxin was attenuated by 1% formaldehyde in phosphate buffer for 24 hours, Balb / C mice were immunized to prepare monocl...

Embodiment 2

[0037] Preparation of colloidal gold probes

[0038] 1. Preparation of colloidal gold particles

[0039] Take a siliconized 250mL triangular flask, add 100mL deionized water and 1mL 1% chloroauric acid, heat to boil; add 1.5mL 1% trisodium citrate accurately while stirring, continue to boil for 15min, and return to the original state with deionized water after cooling. Colloidal gold particles with a particle size of 25nm can be obtained. Colloidal gold particles with a particle diameter of 15-40 nm can be prepared by adjusting the amount of adding 1-2 mL of 1% trisodium citrate.

[0040] 2. Colloidal gold-labeled mouse monoclonal antibody that specifically binds to the A chain of abrin toxin

[0041] Take a siliconized 250mL triangular flask, add 100mL of colloidal gold with a particle size of 25nm; add an appropriate amount of 0.2mol / L K2CO3 to adjust the pH to 8.7, and slowly add a mouse-derived monoclonal antibody that specifically binds to the A chain of abrin toxin to ...

Embodiment 3

[0045] Assembly and testing of immunochromatographic test paper for detection of abrin toxin

[0046] 1. Preparation of detection line and quality control line on nitrocellulose membrane

[0047] The purified rabbit-derived anti-abbrin polyclonal antibody and goat anti-mouse IgG polyclonal antibody were diluted with PBS (0.01mol / mL.pH7.5) solution to a final concentration of 1mg / mL and 1.5mg / mL, respectively. Put the diluted rabbit-derived anti-abbrin toxin polyclonal antibody solution into the nozzle 2 of the BIODOT film scriber, and fix it at a position 1.1 cm away from the lower edge of the nitrocellulose membrane, and put the diluted goat anti-mouse IgG polyclonal antibody into The nozzle 1 of the BIODOT membrane machine is fixed at a position 1.6cm away from the lower edge of the nitrocellulose membrane. The parameters are 1.0μL / cm sprayed on the nitrocellulose membrane. Vacuum-dry the sprayed nitrocellulose membrane for 2 hours (at 24°C relative humidity below 40%). Af...

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Abstract

The invention discloses immunochromatographic test paper for testing abrin and a preparation method thereof. The preparation method comprises the following steps of: preparing the gold colloid pad of the immunochromatographic test paper by marking colloidal gold by a mouse source monoclonal antibody which is specifically bound with the chain A of the abrin; coating a rabbit source anti-abrin polyclonal antibody on a nitrocellulose membrane serving as a testing line (T line); and assembling the T line and the gold colloid pad to form the immunochromatographic test paper. The test paper is suitable for an on-site test for a clinical test and emergency and an epidemiological survey, has the advantages of simple operation, high sensitivity, good stability, convenience, rapidness and the like, and has significance and an actual application value for the differential diagnosis of abrin toxication and timely cure.

Description

technical field [0001] The invention belongs to the technical field of biological detection, and in particular relates to an immunochromatographic test paper for detecting abrinia toxin and a preparation method thereof. Background technique [0002] Abrin is a poisonous protein present in the seeds of the leguminous plant Abrus Precatorius L. It is the most toxic plant toxin found so far, and its toxicity is a common chemical weapon The lethal dose taken by an adult is 5.0-7.0 μg / Kg. You only need to chew and swallow a seed to be poisoned to death, and when symptoms appear, serious organic damage has been caused to the body. The treatment and prevention of poisoning bring great difficulties, and it is a potential biotoxin weapon. The abrin toxin is composed of two polypeptide chains A and B connected by a disulfide chain, in which the abrin toxin A chain has N-glycosidase activity, which can specifically hydrolyze the N-C glycosidic bond of the 4324th adenosine of ribosome ...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/531
Inventor 李小兵刘国文孔涛王哲宋文学杨威孙佳张燚刘磊唐佳
Owner JILIN UNIV
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