Preparation method of immunogold rapid test paper for vibrio parahaemolyticus

A hemolytic vibrio and detection test paper technology, which is applied in the detection field, can solve the problems of prolonged product transportation and storage time, affecting the work efficiency of inspection institutions, and cumbersome inspection methods, etc., and achieves the effects of bright color, convenient customs clearance speed, and low price

Active Publication Date: 2012-03-28
厦门佰能检验技术服务有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The inspection method is cumbersome, time-consuming and labor-intensive, which not only affects the work efficiency of the inspection agency, but also prolongs the product transportation and storage time of the production department and increases the production cost

Method used

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Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0032] 2. Preparation of rabbit anti-Vibrio parahaemolyticus antibody

[0033] The inactivated bacterial solution was immunized with intravenous injection and Freund's adjuvant, and the specific immunization procedure was as follows:

[0034] 1) Intravenous injection immunization: once every 10 days, 1 mL for the first time, 1.5 mL for the second time, 2 mL for the third time, 3 mL for the fourth time, bloodletting when the titer reaches 1:1280 or 1:2560;

[0035] 2) Multi-point subcutaneous injection of Freund's adjuvant for immunization: After Freund's complete adjuvant was mixed with an equal volume of the bacterial solution, 4 mL was injected subcutaneously at multiple points on the back of the rabbit's neck, and after 2 weeks, the Freund's incomplete adjuvant was mixed with an equal volume of the bacterial solution Inject 4 mL subcutaneously at multiple points on the back of the neck, inject 4 mL intravenously 5-7 days before blood collection after 2 weeks, and let blood ...

Embodiment 1

[0044] Rabbit anti-Vibrio parahaemolyticus antibody labeling, take the required amount of colloidal gold solution, and use 0.1 mol / L K 2 CO 3 and 0.1 mol / L HCl to adjust its pH to 8.5. Then add the purified rabbit anti-Vibrio parahaemolyticus antibody drop by drop under magnetic stirring (the rabbit antibody immunized with adjuvant is labeled with gold, add 16.7ug of rabbit anti-Vibrio parahaemolyticus antibody per mL of colloidal gold solution), and continue magnetic stirring for 30 min Then add 5% PEG (MW 6000) to a final concentration of 0.1%, and continue to stir for 15 min, and finally place it in a 4°C refrigerator for overnight. The next day, the labeled rabbit anti-Vibrio parahaemolyticus antibody colloidal gold was centrifuged at 4000 r / min for 10 min, and the supernatant was centrifuged at 15000 r / min for 30 min, and the supernatant was discarded carefully. Resuspend the pellet with resuspension buffer (containing 1% casein, 0.05% PEG6000, 0.05% Tween-20, 0.01 mol / ...

Embodiment 2

[0046] The composition of the reaction test strip is: the size is about 5.0 cm~6.0cm×0.4cm~0.6 cm. The specific composition is: the bottom layer is PVC rubber sheet, the upper end of the test strip is absorbent paper, and the lower end is glass combined with colloidal gold. Fiber membrane and sample pad, the middle is a nitrocellulose membrane (NC membrane), the upper end is sprayed with quality control reagents, such as goat anti-rabbit IgG or SPA, the lower end is sprayed with rabbit anti-2, and the surface of the test strip is covered A layer of film is used for protection.

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PUM

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Abstract

The invention discloses a preparation method of immunogold rapid test paper for vibrio parahaemolyticus. The method comprises the following steps of: (a) preparing a vibrio parahaemolyticus antigen; (b) preparing a vibrio parahaemolyticus inactivated vaccine; (c) preparing a polyvalent antiserum with a vaccine immune experimental rabbit; (d) purifying the antiserum; (e) marking with colloidal gold; and (f) preparing an immunogold test strip. A detection technology in which the immunogold rapid test paper for vibrio parahaemolyticus is adopted has the advantages of easiness, sensitivity, rapidness, specificity and the like; the customs clearance speed of port imported and exported goods can be increased greatly; and meanwhile, the test paper has the advantages of low price, no need of instrument, easiness and rapidness for operationing and easiness in judging results.

Description

technical field [0001] The invention relates to the technical field of detection, in particular to a method for preparing a rapid detection test paper for Vibrio parahaemolyticus immunogold. Background technique [0002] Vibrio parahaemolyticus belongs to the genus Vibrio in the family Vibrio, and is a pathogenic bacterium transmitted through food and water. Vibrio parahaemolyticus has similar biological characteristics and the same pathogenicity, and both can cause diarrhea and food poisoning. [0003] Vibrio parahaemolyticus was first isolated from a food poisoning patient in Japan by Fujino et al. in 1953. It is also called halophilic bacteria, because it can grow in a medium containing more salt (3%~4%) , so named because they cannot reproduce in a medium without salt. Vibrio parahaemolyticus is distributed all over the world, and usually exists in bays and coastline areas and seafood. Many people cause food poisoning due to eating seafood contaminated with this bacter...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/531
Inventor 孔繁德徐淑菲刘阳陈信忠龚艳清杨俊萍郭树林
Owner 厦门佰能检验技术服务有限公司
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