Colloidal gold test strip for detecting sheep fasciola hepatica infection circulating antigen

A technology of colloidal gold test paper and Fasciola goat liver fluke, which is applied in the field of immunology, can solve problems such as lack, and achieve the effects of easy portability, rapid detection, and guaranteed specificity

Pending Publication Date: 2022-03-25
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Aiming at the lack of colloidal gold rapid detection kits for Fasciola goat liver fluke infection in my country at present, the present invention establishes a

Method used

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  • Colloidal gold test strip for detecting sheep fasciola hepatica infection circulating antigen
  • Colloidal gold test strip for detecting sheep fasciola hepatica infection circulating antigen
  • Colloidal gold test strip for detecting sheep fasciola hepatica infection circulating antigen

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1 Expression and Purification of Fasciola hepatica Cathepsin L Recombinant Protein

[0033] 1. Inoculate the BL21 positive strain of Fasciola hepatica cathepsin L recombinant protein into LB medium, and activate it by culturing at a constant temperature of 37°C. The activated strain was re-inserted into LB medium, and cultured at a constant temperature of 37°C until the OD of the bacterial solution 600 The value is 0.6, adding inducer, and inducing at 37°C for 8 h. The supernatant was discarded by centrifugation. The pellet was resuspended in PBS and sonicated. The bacterial solution was centrifuged to take the precipitated sample and subjected to SDS-PAGE electrophoresis, and the gel was stained with Coomassie brilliant blue staining solution.

[0034] 2. Introduce the positive strains into a large amount of LB medium in proportion, prepare a large number of protein samples according to the previous step and perform conventional SDS-PAGE electrophoresis, soa...

Embodiment 2

[0038] Preparation and purification of monoclonal antibody against Fasciola hepatica cathepsin L

[0039] 1. Immunize BALB / C mice with purified Fasciola hepatica cathepsin L recombinant protein, immunize 4 times, and detect the antibody titer in the serum after each interval of 1 week, select the mouse with the highest titer before the cell fusion test 3-day booster immunization;

[0040] 2. Blow off the adherent SP2 / 0 cells and transfer to a centrifuge tube. Blood was collected from the eyes of the mouse, killed by neck dislocation, the spleen was removed, crushed, sieved and purified by centrifugation; the thymus was removed and processed in the same way; the processed thymocytes were transferred to a centrifuge tube, added with HAT and HT and placed at a constant temperature of 37°C spare in the box;

[0041] 3. Discard the supernatant of the centrifuged splenocytes, add IMDM to mix, and centrifuge to discard the supernatant. Place the centrifuge tube in a 37°C water bat...

Embodiment 3

[0060] Preparation and purification of polyclonal antibody against Fasciola hepatica cathepsin L

[0061] 1. Use the purified Fasciola hepatica cathepsin L recombinant protein to immunize New Zealand rabbits for 3 times. After the third immunization, the antibody titer in the serum was detected at an interval of 7 days, blood was collected from the heart, and the serum was separated.

[0062] 2. Mix the serum and Tris-HCl buffer evenly, and centrifuge to absorb the supernatant. Equilibrate the purification column with sodium phosphate solution, inject the serum into the purification column, wash thoroughly and elute the antibody with the eluent and collect it into a centrifuge tube. The eluate was put into a dialysis bag and submerged in PBS solution for dialysis at 4°C overnight, and a small amount of sample was taken for SDS-PAGE electrophoresis staining analysis.

[0063] result

[0064] 1. Determination of titer of rabbit polyclonal antibody against Fasciola hepatica ca...

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Abstract

The invention discloses a colloidal gold test strip for detecting a lamb fasciola hepatica infection circulating antigen, and the prepared lamb fasciola hepatica infection colloidal gold test strip can be used for detecting the lamb fasciola hepatica infection circulating antigen and has the advantages of simplicity in operation, rapidness in detection, convenience in carrying and the like in the aspect of use. The detection result is visual, easy to understand and convenient to interpret, and the kit can be used for detecting early infection of fasciola hepatica, can distinguish current infection from previous infection, and is suitable for on-site detection of clinical samples. The anti-fasciola hepatica cathepsin L monoclonal antibody ensures the specificity of the detection method. The rabbit anti-fasciola hepatica cathepsin L polyclonal antibody improves the sensitivity of the detection method.

Description

technical field [0001] The invention relates to a colloidal gold test strip for detection of circulating antigens infected by Fasciola goat liver fluke, which is used for the detection of Fasciola goat liver fluke infection and belongs to the technical field of immunology. Background technique [0002] Fascioliasis is caused by Fascioliasis ( Fasciola hepatica ) caused by an important zoonotic parasitic disease, was a worldwide epidemic. The disease mainly occurs in ruminants, mostly cattle and sheep. Fasciola hepatica parasitizes the hepatic and bile ducts of the host, causing chronic or acute hepatitis and cholangitis. Chronic infection manifests as nutritional disorders and systemic poisoning. Severe infection can cause acute death of young animals and cause huge economic losses to animal husbandry. [0003] Currently, the methods used to detect Fasciola hepatica infection mainly include pathogenic detection methods, molecular biological detection methods and immunologi...

Claims

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Application Information

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IPC IPC(8): G01N33/558G01N33/573G01N33/577G01N33/58G01N33/532
CPCG01N33/558G01N33/577G01N33/587G01N33/532G01N33/573G01N2333/96413
Inventor 张西臣刘少雄于钦磊张楠李建华宫鹏涛王晓岑李新
Owner JILIN UNIV
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