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Dot-immunogold filtration reagent kit for livestock schistosomiasis diagnosis, its preparation and application method

A spotted gold diafiltration and schistosomiasis technology, which is applied in the field of domestic animal schistosomiasis spotted gold immune diafiltration diagnostic kit and its preparation, can solve the problems of long processing period, unfavorable industrial development, and high detection cost, and achieve stability and reproducibility are good, conducive to industrial development, the effect of simple production process

Inactive Publication Date: 2011-06-08
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The main problem of this method is that it is necessary to prepare schistosomiasis recombinant protein polyclonal antibody or rabbit anti-human IgG or rabbit anti-schistosome egg polyclonal antibody or GST-PcAb. The preparation of these specific polyclonal antibodies has a long cycle, complicated procedures, and high cost, which limits They are popularized and applied in the census of livestock schistosomiasis
Moreover, when preparing colloidal gold-labeled probes, the amount of polyclonal antibody is usually determined by visual inspection of the test tube. In order to ensure sufficient coating and avoid the interference of excess colloidal gold and non-specific protein binding to the detection result judgment, it is generally added on this basis. 10% to 20% of the polyclonal antibody dosage, and then in order to remove the excess free polyclonal antibody, purification such as chromatography or high-speed centrifugation is required. These treatments either require high equipment conditions, or the processing cycle is long, and the steps Complicated, so the cost is high, which is not conducive to industrial development; third, because the method uses the capture polyclonal antibody as the solid phase, and the serum to be tested and the colloidal gold probe as the mobile phase, one reaction box can only detect blood samples from one person , the detection cost is high and the efficiency is low; at the same time, the test box containing the nitrocellulose membrane coated with antigen or polyclonal antibody needs to be stored at 4-8°C, which increases the difficulty for large-scale application; the fourth is There are many reaction steps, and the interference is relatively increased

Method used

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  • Dot-immunogold filtration reagent kit for livestock schistosomiasis diagnosis, its preparation and application method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Embodiment 1: (diagnostic kit)

[0029] Including box body, built-in reaction box, black reagent bottle, normal saline bottle (with 0.85% normal saline inside), glass capillary (inner diameter 1mm) for spotting blood samples, standard positive blood paper and negative blood paper. The purpose of these two papers is to provide, One is for users to check whether the colloidal gold used for schistosome antigens is effective, and the other is to provide users with reference standards for distinguishing negative reactions and positive reactions; among them, the reaction box is a small plastic box of 3cm×2.5cm×0.6cm, divided into boxes There are two parts, the bottom and the box cover. There is a small round hole of 0.8cm in the center of the box cover. 0.45μ or 0.65μ), close the lid to form a reaction box; the black reagent bottle contains schistosome antigen colloidal gold.

Embodiment 2

[0030] Embodiment 2: (preparation method 1 of colloidal gold)

[0031] The present invention uses 1% chloroauric acid, 1% trisodium citrate, 1% tannic acid, 25mMK 2 CO 3 Colloidal gold was prepared by tannic acid-sodium citrate reduction method with double distilled water as raw material. Sodium citrate is mainly a reducing agent, while tannic acid has the dual functions of reduction and protection, controlling the formation process of "crystal nuclei". The amount of tannic acid determines the size of the colloidal gold particle size, so changing the amount of tannic acid can achieve the purpose of preparing colloidal gold particles with required diameters. This example A liquid, the proportioning of each raw material component of B liquid is shown in Table 1:

[0032] Table 1 Preparation of liquid A and liquid B (in ml)

[0033] Heat the prepared liquid A and liquid B simultaneously in a water bath to 59-61°C, stir liquid A on an electromagnetic stirrer, quickly add liq...

Embodiment 3

[0034] Embodiment 3: (preparation method 2 of colloidal gold)

[0035] Table 2 Preparation of liquid A and liquid B (in ml)

[0036] The preparation and synthesis methods of liquid A and liquid B are the same as in Example 1, and the diameter of colloidal gold particles is 10<D≤11nm.

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Abstract

The invention provides a preparing and application method of diffusion medium box for diagnosing livestock's snail fever spot gold, which is a new immune labeling technique and livestock's snail fever immunology diagnosis technique. The invention labels colloid gold onto Japanese snail antigen protein, and is suit for testing many kinds of animals such as cattle, sheep, pig and rabbit; it spots tested animal's blood dried paper leachate onto pyroxylin film as posited phase, then it droplets snail antibody colloid gold 2-3 drops (100-150mul) and produces naked eye visible red speckle in 2 minutes. Its positive coincidence rate is 100% and its negative coincidence rate is 99.2%.

Description

technical field [0001] The invention relates to the technical field of diagnosis of livestock schistosomiasis, in particular to a spotted gold immunofiltration diagnostic kit for livestock schistosomiasis and a preparation method and application method thereof. Background technique [0002] Domestic animal schistosomiasis not only endangers the health of domestic animals, but also is an important disseminator of the source of the disease. With the in-depth development of my country's schistosomiasis work, although great achievements have been made, there are still more than 100 counties (cities, districts) in my country that have failed to control the spread of the disease. Long-term consolidation and monitoring tasks remain. If the conventional fecal-hatched schistosomiasis method is used to detect schistosomiasis miracidia, it needs to invest a lot of manpower, material resources and funds; the existing serological methods for schistosomiasis diagnosis in livestock include...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569G01N21/78G01N33/532G01N33/53
Inventor 张雪娟卢福庄方兰勇冯尚连项美华程菊芬周永学
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES