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Detection kit and detection method for equus cryptosporidiosis

A technology of cryptosporidiosis and detection kit, which is applied in the directions of measuring devices, instruments, scientific instruments, etc., can solve the problem of no accurate detection of equine cryptosporidiosis, and achieve convenient and fast detection, high stability and specificity. , the effect of reducing costs

Inactive Publication Date: 2018-09-04
边学静 +3
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no effective and accurate detection method for equine cryptosporidiosis

Method used

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  • Detection kit and detection method for equus cryptosporidiosis
  • Detection kit and detection method for equus cryptosporidiosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Horse peripheral blood samples were obtained from domestic horses infected with Cryptosporidiosis equine and non-infected horses, and the number was 127 samples to be tested.

[0041] 10 mL of blood was collected from each sampled individual with a sodium heparin collection tube, and transported to the laboratory within 6 hours at room temperature.

[0042] The detection steps of each sample to be tested are as follows:

[0043] (1) Separate the PBMC with the separation medium, resuspend the separated cells in serum-free medium, and the cell concentration is 1.5×10 5 Cells / mL; Add the resuspended cells into a PVDF96-well plate, add 100 μL to each reaction well, add each sample to 10 wells, and make 10 repetitions;

[0044] (2) Add 50 μL of a mixture consisting of serum-free medium and 6 polypeptides with a final concentration of 5 μM to each reaction well, mix well and carry out co-incubation; set a positive control and a negative control, and add high Specific mouse an...

Embodiment 2

[0053] Horse peripheral blood samples were obtained from domestic horses infected with Cryptosporidiosis equine and non-infected horses, and the number was 127 samples to be tested.

[0054] 10 mL of blood was collected from each sampled individual with a sodium heparin collection tube, and transported to the laboratory within 6 hours at room temperature.

[0055] The detection steps of each sample to be tested are as follows:

[0056] (1) Separate the PBMC with the separation medium, resuspend the separated cells in serum-free medium, and the cell concentration is 3.5×10 5 Cells / mL; Add the resuspended cells into a PVDF96-well plate, add 100 μL to each reaction well, add each sample to 10 wells, and make 10 repetitions;

[0057] (2) Add 50 μL of a mixture consisting of serum-free medium and 6 polypeptides with a final concentration of 5 μM to each reaction well, mix well and carry out co-incubation; set a positive control and a negative control, and add high Specific mouse ...

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Abstract

The invention provides a detection kit and detection method for equus cryptosporidiosis. The kit comprises 6 polypeptides with amino acid sequences shown as SEQ ID NO. 1 to SEQ ID NO. 6. The detectionmethod comprises the following steps: (1) separating a sample to be detected by utilizing a separation solution and re-suspending separated cells into a serum-free medium; (2) adding the 6 polypeptides with the amino acid sequences shown as SEQ ID NO. 1 to SEQ ID NO. 6 into the re-suspended cells and co-incubating; then adding a high-specificity mouse anti-human IgG antibody into a co-incubatingsystem and carrying out antigen-antibody reaction; (3) washing the system after the antigen-antibody reaction; then adding a color developing substrate and carrying out color developing reaction; counting up the number of spots generated by the reaction. The kit provided by the invention can used for artificially synthesizing complete sequences, is convenient and rapid to produce and has high stability and specificity; the kit is simple to operate and the cost is extremely reduced.

Description

technical field [0001] The invention belongs to the technical field of veterinary medical detection, and in particular relates to a detection kit for equine cryptosporidiosis and a detection method thereof. Background technique [0002] Cryptosporidium (Cryptosporidium) is one of the important food-borne and water-borne zoonotic protozoa. There are many species and genotypes in the genus, and most of them have host adaptability. There are geographic features in the type distribution. Cryptosporidiosis is considered to be one of the six most common diarrheal diseases in humans. It is a self-limiting infection in people with normal immune function, but it can cause acute, persistent and chronic diarrhea in children, HIV-infected people and patients with immunodeficiency diseases. even lead to death. [0003] In recent years, based on molecular biological detection, the species / genotypes of Cryptosporidium infected by horses have shown diversity, with Cryptosporidium parvum a...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56905G01N2333/44
Inventor 边学静殷玉芹夏召莲夏召亮
Owner 边学静
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