Theileria equi immunoblotting detecting method and method for preparing kit

A technology of equine worm disease and detection kit, which is applied in the detection of equine worm disease serum antibody by immunoblotting test, and in the field of equine worm blot detection (Babesiosis equine), which can solve the problem of equine Theileria diagnostic kit and other issues, to achieve the effect of saving consumption, shortening the cycle, and reducing the amount of reagents

Inactive Publication Date: 2011-07-20
中华人民共和国天津出入境检验检疫局
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] At present, in China, there are few studies on the detection of animal diseases by immunoblotting, and there is no research report on the detection method and diagnostic kit of equine theilesiosis by immunoblotting hybridization in China

Method used

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  • Theileria equi immunoblotting detecting method and method for preparing kit
  • Theileria equi immunoblotting detecting method and method for preparing kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0056] Example 1 :Preparation of Western Blot Diagnosis Kit for Martell's Disease

[0057] Reagent preparation:

[0058] 1. 1x VYM’s buffer:

[0059] CaCl 2 .2H 2 O 0.016 g

[0060] KCl 0.400 g

[0061] KH 2 PO 4 1.415 g

[0062] MgSO 4 .7H 2 O 0.154 g

[0063] Na 2 HPO 4 0.077 g

[0064] NaCl 7.077 g

[0065] Glucose 20.500 g

[0066] ddH 2 O 1 L

[0067] After stirring and dissolving, add Adenine 0.0423 g and Guanosine 0.0708 g, adjust the pH to 7.0-7.2, vacuum filter (0.22 μm), and store at 4°C for later use.

[0068] 2. HL2A-FBS culture medium:

[0069] HL-1 medium 35 mL

[0070] Fetal bovine serum 10 mL

[0071] HB101 supplement 1 mL

[0072] Hepes 0.238 g

[0073] L-glutamine(100x) 1 mL

[0074] Hypoxanthine(20mmol) 0.5 mL

[0075] Gentamicin 1 mL

[0076] Antibiotic-antimycotic(100x) 1 mL

[0077] Stir magnetically for 5 min, adjust the pH to 7.2, vacuum filter (0.22 μm), and store at 4°C for later use.

[0078] 3. PI buffer:

[0079] Tris 3.03 g

[0080] Deionized water 500mL

[0081] S...

Embodiment 2

[0106] Example 2 : Western Blot Detection of Matele's Disease

[0107] Reagent preparation:

[0108] 1. 10x running buffer:

[0109] Tris 121.1 g

[0110] Glycine 570 g

[0111] Distilled water 4 L

[0112] 2. Transfer buffer:

[0113] 10x running buffer 100 mL

[0114] Methanol 200 mL

[0115] Distilled water 700 mL

[0116] 3、 Blocking fluid ： Phosphate buffer containing 0.2% Tween-20, 10% skimmed milk powder

[0117] 4. Washing solution: Phosphate buffer containing 0.2% Tween-20

[0118] Take the NuPAGE 4%-12% Bis-Tris gel plate, wash the gel wells with 1 x SDS Running Buffer three times; place the gel plate face forward in the core rack of the Mini-Cell vertical electrophoresis system and lock it tightly. Add 200 mL 1 x SDS Running Buffer and 500 μL NuPAGE Antioxidant to the electrophoresis tank and mix well. Add 600 mL 1 x SDS Running Buffer to the lower tank. Add 7 μL to 10 μL of protein molecular weight standard marker (Marker) and the antigen sample obtained in Example 1 into the ...

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Abstract

The invention discloses a theileria equi immunoblotting detecting method and a method for preparing a kit, wherein the theileria equi immunoblotting detecting method in the invention is established on the basis of acquiring diagnostic antigen and positive control serum via Cochlearia officinalis hyperoxide enzyme label and chemical light emitting prime. The solvent for the method in the inventionis little in amount, low in cost, high in specificity, high in sensitivity, high and fast in operation, free of toxicity and harm, low in required conditions, easy for judging the test result, long-time for storage, and stronger in specificity than ELISA. The method in the invention detects no false positive and false negative result.

Description

technical field [0001] The present invention relates to an immunoblotting detection method and a kit preparation method for equine theilesiosis (babesia equine), in particular to a method for detecting serum antibodies against equine theilesiosis by immunoblotting test and a diagnostic reagent thereof The invention discloses a method for preparing a box, belonging to the field of animal epidemic detection. Background technique [0002] Theileria equi or Babesia equi is a blood protozoan disease caused by the parasitic horse Theileria equi or Babesia equi in the erythrocytes of equine animals, and is transmitted by hard ticks. After infection, it mainly parasitizes in animal blood cells. The clinical manifestations are fever, anemia, jaundice, hemoglobinuria, splenomegaly and other symptoms. The sick horse was dead or dying when found. [0003] Marthelliasis has a worldwide distribution and its prevalence is directly related to the distribution of transmissible ticks. The ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/545
Inventor 王玉玲柴宏森姜红左锋柴铭骏赵林立侯艳梅赵祥平
Owner 中华人民共和国天津出入境检验检疫局
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