Kit for enzyme-linked immunosorbent assay of cordyceps sinensis and preparation method of kit

An enzyme-linked immunoassay and cordyceps sinensis technology, which is applied in the direction of biological testing, measuring devices, material inspection products, etc., can solve the problems of inability to identify, weight gain, etc., and achieve the effects of wide application range, good accuracy and high sensitivity

Inactive Publication Date: 2015-04-29
SHENZHEN ACAD OF METROLOGY & QUALITY INSPECTION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This led to the emergence of various pseudo-Cordyceps in the market, including weight gain, using bamboo sticks as shoddy products, and some using similar Cordyceps instead of Cordyceps, making it difficult to identify

Method used

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  • Kit for enzyme-linked immunosorbent assay of cordyceps sinensis and preparation method of kit
  • Kit for enzyme-linked immunosorbent assay of cordyceps sinensis and preparation method of kit
  • Kit for enzyme-linked immunosorbent assay of cordyceps sinensis and preparation method of kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] Example 1 Preparation of target antigen.

[0022] After grinding the body part of Cordyceps sinensis under liquid nitrogen conditions, 10 g of the sample was taken, and 100 ml of 0.1M PBS buffer (pH7.4, containing 0.5% mercaptoethanol, 0.1% Tween80) was ultrasonically extracted for 12 hours in an ice bath. The supernatant was collected by centrifugation and precipitated with 60% saturation ammonium sulfate. 8M urea was used to dissolve the precipitate, centrifuged, and the supernatant was dialyzed overnight to obtain a crudely separated sample.

[0023] The primary separated sample was repurified using a vertical electrophoresis apparatus: the primary separated sample was subjected to SDS-PAGE electrophoresis using a 4% stacking gel and a 12% separating gel, and a prestained protein standard was used as a molecular weight reference. After electrophoresis at 80v for 2 hours, a protein gel band with a molecular weight of 70kd was cut out with reference to the prestained ...

Embodiment 2

[0024] Example 2 Preparation and purification of anti-Cordyceps sinensis antibody.

[0025] The immunogen prepared in Example 1 was used to immunize female balb / c mice for 6 weeks respectively, 3 mice in each group. For the first immunization injection, 100 μL of 100 μg / mL immune antigen was fully emulsified with the same amount of complete Freund’s adjuvant, and injected directly into the intraperitoneal cavity. After an interval of two weeks, take the sampled antigen, emulsify it with 100 μL of incomplete adjuvant, and inject it in the same way.

[0026] One day before cell fusion or on the same day, the Kunming rats were killed by pulling the neck, soaked in 70% alcohol, and disinfected the body surface; the Kunming rats were fixed on the wax board with pins, the abdomen was cut open on the ultra-clean workbench, and the peritoneum was picked up with small tweezers. Inject 5 mL of RPMI-1640 complete culture solution (obtained by adding 15% fetal calf serum to GIBICO RPMI-1...

Embodiment 3

[0033] Example 3 Preparation of standard substance of Cordyceps sinensis.

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Abstract

The invention provides a preparation method of a kit for enzyme-linked immunosorbent assay of cordyceps sinensis. The preparation method comprises the following steps: 1, extracting and separating cordyceps sinensis 70kDa glycoprotein; 2, preparing a monoclonal antibody; 3, establishing and evaluating an enzyme-linked immunosorbent assay method; and 4, assembling the kit. The invention also provides the kit for enzyme-linked immunosorbent assay of cordyceps sinensis. The kit provided by the invention has the beneficial effect that the true and false identification of cordyceps sinensis can be realized by using the kit for the enzyme-linked immunosorbent assay of cordyceps sinensis.

Description

technical field [0001] The invention relates to the identification of Cordyceps sinensis, in particular to a Cordyceps ELISA detection kit and a preparation method thereof. Background technique [0002] "Chinese Pharmacopoeia" (2010 edition) stipulates that the authentic product of Cordyceps sinensis is "the complex of the larvae and the larvae of the fungus Cordyceps sinensis (Berk.) Sacc. of the family Ergotaceae". It has the effects of nourishing the kidney and lung, stopping bleeding and resolving phlegm, and is known as the three major Chinese tonics together with ginseng and deer antler. Cordyceps sinensis has a special growth environment, scarce resources, and high prices. There are a large number of adulterated and confused products on the market. How to quickly and effectively identify the authenticity is the primary problem to be solved to ensure the safety and effectiveness of the clinical use of Cordyceps sinensis. Counterfeit products of Cordyceps sinensis main...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/577G01N33/531
CPCG01N33/531G01N33/577G01N33/68
Inventor 赖晓芳张世伟赖心田伍聪杨国武李碧芳冯荣虎王士峰马广东王珍妮
Owner SHENZHEN ACAD OF METROLOGY & QUALITY INSPECTION
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