Bigeminal colloidal gold test strip for detecting toxoplasma gondii and neospora caninum antibodies

A technology of colloidal gold test paper and neospora, which is applied in the field of serological detection, can solve the problems of diagnosis of toxoplasma gondii and neospora infection, etc., and achieves small environmental factors, strong specificity and sensitivity, and requires a large amount of samples little effect

Inactive Publication Date: 2016-07-20
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] There is currently no test that can diagn

Method used

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  • Bigeminal colloidal gold test strip for detecting toxoplasma gondii and neospora caninum antibodies
  • Bigeminal colloidal gold test strip for detecting toxoplasma gondii and neospora caninum antibodies
  • Bigeminal colloidal gold test strip for detecting toxoplasma gondii and neospora caninum antibodies

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0052] Embodiment 1, preparation of TgGRA1 protein and NcSRS2 protein

[0053] TgGRA1 protein and NcSRS2 protein were prepared by prokaryotic expression respectively, the specific method is as follows:

[0054] 1. Cloning of TgGRA1 and NcSRS2 genes

[0055] Primers were designed to amplify the TgGRA1 / NcSRS2 gene, and the primers are shown in Table 1 below. The two ends of the primers contain enzyme cutting sites, which are indicated in lowercase. The 5' end of the primer has added protective bases, and the 3' end is the sequence of TgGRA1 / NcSRS2 gene.

[0056]Table 1 is the primers used to amplify TgGRA1 and NcSRS2 genes

[0057]

[0058] Using the cDNA of the Toxoplasma gondii RH strain as a template, PCR amplification was performed with the upstream and downstream primers of the TgGRA1 gene to obtain a 495bp PCR product, which was the TgGRA1 gene. After sequencing, the nucleotide sequence of the TgGRA1 gene was sequence 1 in the sequence table, TgGRA1 The protein TgGRA...

Embodiment 2

[0090] Embodiment 2, the preparation of double colloidal gold test paper

[0091] 1. Preparation of nitrocellulose membrane containing detection line T1, detection line T2 and quality control line C

[0092] 1) Preparation of coated antigen

[0093] The NcSRS2 protein solution is to dissolve the tagged NcSRS2 protein prepared in Example 1 above in PB, and the concentration of the tagged NcSRS2 protein is 1 mg / ml, which is the NcSRS2 antigen.

[0094] TgGRA1 protein solution is to dissolve the tagged TgGRA1 protein prepared in Example 1 above in PB (PB preparation: weigh 5.37 g of disodium hydrogen phosphate, 0.2 g of potassium chloride, and 0.2 g of potassium dihydrogen phosphate, add ultrapure water to 1000ml, mix well and set aside.), and the concentration of the tagged TgGRA1 protein is 2mg / ml, which is the TgGRA1 antigen.

[0095] Pig IgG protein solution is to dissolve the above-mentioned pig IgG protein (purchased from Zhongke Maichen (Beijing) Technology Co., Ltd., ca...

Embodiment 3

[0125] Embodiment 3, the detection effect of test strip

[0126] 1. Specificity and sensitivity

[0127] The colloidal gold test strip (DualGICA) and indirect immunofluorescence test (IFAT) two methods prepared by above-mentioned embodiment 2 detect identical rabbit, dog, cat serum, compare the coincidence rate of these two kinds of method results, calculate test strip Based on the specificity and sensitivity of IFAT.

[0128] 1) Colloidal gold test strip detection

[0129] Add 100 μl of 20-fold diluted rabbit, dog, and cat serum samples to the sample pad of the colloidal gold test strip prepared in 3 above, and judge the result within 15 minutes. according to Figure 4 Judgment method to determine whether toxoplasma gondii or neospora infection.

[0130] 2) The operation steps of the IFAT method are as follows

[0131] A. Collect the freshly released Toxoplasma gondii or Neospora, suspend them with an appropriate amount of PBS, and drop them on the glass slides with adso...

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Abstract

The invention discloses a bigeminal colloidal gold test strip for detecting toxoplasma gondii and neospora caninum antibodies, and provides a bigeminal colloidal gold test strip for detecting toxoplasma gondii and/or neospora caninum. The bigeminal colloidal gold test strip comprises a bottom plate, as well as a sample pad, a colloidal gold pad, a nitrocellulose membrane and an absorbent pad, which are arranged on the bottom plate, wherein the colloidal gold pad is coated with a colloidal gold protein; the nitrocellulose membrane contains a detection line T1, a detection line T2 and a quality control line C. Experiments show that the prepared bigeminal colloidal gold test strip can be used for simultaneously detecting plasma antibodies of toxoplasma gondii and neospora caninum in an animal body and is convenient to operate and applicable to detection of animals such as pigs, dogs and cats, differential diagnosis of infections caused by the two animal protozoans is facilitated and the cost is reduced.

Description

technical field [0001] The invention belongs to the field of serological detection, and relates to a double colloidal gold test strip for detecting antibodies to toxoplasma gondii and neospora. Background technique [0002] Toxoplasmosis is caused by the obligate intracellular parasitic protozoan Toxoplasmagondii (T. Toxoplasma gondii can infect a variety of animals and humans, and the occurrence, symptoms and pathogenicity of the infection are closely related to the genotype of Toxoplasma gondii, the amount of infection, the genetic background of the host and the immune status. Under normal circumstances, the symptoms of animal infection are mild. If the body's immunity is low, the symptoms will be obvious and death may occur. Another hazard of toxoplasmosis is that it causes reproductive disorders such as abortion and stillbirth in pregnant animals. [0003] Neosporosis (Neosporosis) is a parasitic disease caused by a variety of animals caused by Neospora caninum (Neospo...

Claims

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Application Information

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IPC IPC(8): G01N33/68G01N33/569G01N33/558
CPCG01N33/68G01N33/558G01N33/56905G01N2469/20
Inventor 刘群南汇珠刘晶
Owner CHINA AGRI UNIV
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