Indirectly racing ELISA detecting method for gonyatoxine GTX2,3

A technology for the detection of gonyal toxins and detection methods, which is applied in the field of indirect competition ELISA, can solve the problems of high prices, and achieve the effect of low cost and guaranteed benefits

Inactive Publication Date: 2008-06-11
JINAN UNIVERSITY
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0011] At present, there are commercialized PSP enzyme immunology detection kits on the market in foreign countries, but they are expensive. T

Method used

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  • Indirectly racing ELISA detecting method for gonyatoxine GTX2,3
  • Indirectly racing ELISA detecting method for gonyatoxine GTX2,3
  • Indirectly racing ELISA detecting method for gonyatoxine GTX2,3

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Embodiment 1

[0040] Utilize anti-GTX2, the 3 monoclonal antibody detects GTX2, the indirect competitive ELISA method of 3, comprises the following steps:

[0041] (A) GTX2, 3 was coupled to hemocyanin (KLH) by the formaldehyde method, mixed with Freund's incomplete adjuvant to immunize BALB / c mice, and the splenocytes of the immunized mice were fused with SP2 / 0 myeloma cells, The positive cell line stably secreting anti-GTX2,3 monoclonal antibody (MAb-GTX2,3) was screened out and expanded for culture, the cells were injected into mice to induce ascites, and the protein A column was purified to obtain anti-GTX2,3 monoclonal antibody.

[0042] (B) GTX2,3-glucose oxidase was obtained by coupling with periodate oxidation method.

[0043] (C) Preparation of antigen pre-coated strips:

[0044] (a) Coating: Dilute GTX2, 3-glucose oxidase (GTX2, 3-GOX) (Glucose Oxidase, glucose oxidase) to 5 μg / mL, 100 μL per well was added to the microwells of a polystyrene microplate, and coated overnight at ...

Embodiment 2

[0066] Utilize anti-GTX2, the 3 monoclonal antibody detects GTX2, the indirect competitive ELISA method of 3, comprises the following steps:

[0067] (A) GTX2, 3 was coupled to hemocyanin (KLH) by the formaldehyde method, mixed with Freund's incomplete adjuvant to immunize BALB / c mice, and the splenocytes of the immunized mice were fused with SP2 / 0 myeloma cells, The positive cell line stably secreting anti-GTX2,3 monoclonal antibody (MAb-GTX2,3) was screened out and expanded for culture, the cells were injected into mice to induce ascites, and the protein A column was purified to obtain anti-GTX2,3 monoclonal antibody.

[0068] (B) GTX2,3-glucose oxidase was obtained by coupling with periodate oxidation method.

[0069] (C) Preparation of antigen pre-coated strips:

[0070] (a) Coating: Dilute GTX2, 3-glucose oxidase (GTX2, 3-GOX) (Glucose Oxidase, glucose oxidase) to 5 μg / mL, 100 μL per well was added to the microwells of a polystyrene microplate, and coated at 37°C for 3...

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Abstract

The invention discloses an ELISA method by indirect competition for testing the Gonyautoxin GTX2 and GTX3, which comprises the preparation method of antigen pre-peridium strips and the test method of Gonyautoxin. By utilizing the gained secretory positive cells of the monoclonal antibody with the function of anti-GTX2 and GTX3, the method provided in the invention can prepare in mass production the monoclonal antibody with the function of anti-GTX2 and GTX3, which costs low and can fast, conveniently and sensitively test the content of the GTX2 and GTX3 of the samples. The invention can make fast and sensitive test on the PSP (Paralytic shellfish poison) left in the seafood like fishes and shellfishes on a large scale.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to an indirect competitive ELISA method for detecting the gonylin GTX2,3 by using a monoclonal antibody against the gonyellin GTX2,3. Background technique [0002] Paralytic shellfish poison is a low-molecular-weight, highly toxic shellfish poison that lives in marine microalgae and inhibits nerve conduction mainly through the influence of sodium ion channels. It is a kind of neurotoxin. Gonyautoxin (GTX) is one of the important members of the Paralytic Shellfish Posoning (PSP) family. It is a kind of biguanide-containing tricyclic alkaloid compound. Water is relatively stable under weak acid and low temperature conditions. The survey showed that there are red tide algae that can produce paralytic shellfish poisoning in the north and south sea areas of my country, and a variety of shellfish containing paralytic shellfish poisoning were found in multiple sampling points. It...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/543
Inventor 向军俭贺文妃唐勇罗辉武唐琦邓宁王宏杨红宇
Owner JINAN UNIVERSITY
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