Deoxynivalenol immune affinity column and preparation method and use thereof

A technology of deoxynivalenol and immunophile, which is applied in the direction of chemical instruments and methods, separation methods, preparation of test samples, etc., can solve the problems affecting the efficiency of deoxynivalenol and the linearity of the standard curve. Good, uncontrollable directionality and other problems, to save time and cost, easy to operate, and improve purification efficiency

Active Publication Date: 2014-10-29
山东美正生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the following problems exist in GC chromatography: poor linearity of the standard curve, response drift, retention and memory effects of the last injected sample, large coefficient of variation in repeated injections during MS detection, and matrix interference.
However, since the coupling is non-specific, any part of the antibody may be coupled to the carrier, and the directionality is uncontrollable
It is bound to affect the efficiency of affinity purification column to purify deoxynivalenol

Method used

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  • Deoxynivalenol immune affinity column and preparation method and use thereof
  • Deoxynivalenol immune affinity column and preparation method and use thereof
  • Deoxynivalenol immune affinity column and preparation method and use thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0043] Example 1: Preparation of deoxynivalenol immunoaffinity purification column

Embodiment approach

[0044] A preferred embodiment of the present invention to prepare deoxynivalenol immunoaffinity purification column is as follows:

[0045] 1. Agarose Gel Activation

[0046] Take 2% agarose gel sepharose 2B and wash it with 20 times the volume of distilled water to wash away the remaining ethanol. Use a funnel to filter out the water. Weigh 5 grams of the wet gel after filtering off the water, add 7.5 milliliters of 0.8M NaOH, 2 milliliters of 30% epichlorohydrin, 2 milliliters of sodium borohydride NaBH4, 5 milliliters, shake the table at 25 ° C React for 8 hours.

[0047] After the reaction, wash with 50 ml of distilled water to remove the mixed epichlorohydrin in the gel.

[0048] 2. Coupling of Protein G to Activated Sepharose

[0049] Take 1 gram of activated agarose gel, and use coupling buffer (0.1M NaHCO 3 , 0.8M NaCl, pH8.9) and washed 3 times. Add 20ml of 2mg / ml protein G, and couple at room temperature for 4 hours.

[0050] The coupled agarose carrier was wa...

Embodiment 2

[0062] Example 2: Purification and detection of deoxynivalenol in flour using a deoxynivalenol immunoaffinity column

[0063] 1. Flour sample processing:

[0064] ----Weigh 25g of flour sample, 5g of polyethylene glycol, add 100ml of distilled water;

[0065] ---- Vigorously oscillate on the shaker for 20 minutes to make it completely mixed;

[0066] ----Filter with microfiber filter paper and collect the filtrate;

[0067] ----Take 1mL of filtrate for sample loading detection.

[0068] 2. Equilibrate the deoxynivalenol immunoaffinity purification column at room temperature for 30 minutes.

[0069] 3. Take out the immunoaffinity column, connect the injection port with the syringe barrel, and connect the syringe to the air-controlled operating frame.

[0070] 4. Wash the affinity column 3 times with deionized water, 10 ml each time, and adjust the air pump pressure of the stomata operation frame to make the liquid

[0071] The body flows out at a flow rate of 3 drops / secon...

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Abstract

The invention relates to a deoxynivalenol immune affinity column and a preparation method and use thereof. The immune affinity purifying column is coupled to a sepharose supporter by protein G, and a deoxynivalenol antibody is coupled to the protein G on agarose; a crosslinking agent is used for crosslinking the deoxynivalenol antibody, the protein G and the sepharose supporter, and then the crosslinked deoxynivalenol antibody-protein G-sepharose supporter fills the affinity column. The immune affinity column is mainly used for purifying deoxynivalenol in food, feed, seasonings, wine, beverages and other various samples.

Description

Technical field: [0001] The invention relates to an immunoaffinity column for deoxynivalenol and its preparation method and application. It belongs to the field of food safety testing. Background technique: [0002] Deoxynivalenol (DON), also known as vomitoxin (VT), is mainly one of the secondary metabolites produced by certain Fusarium fungi - a trichothecene toxin. DON is the most common type of trichothecene toxins. Studies have shown that although DON has no obvious carcinogenicity and mutagenicity, it has a wide range of toxicity, especially has a significant impact on immune function. According to the dose of DON and Depending on the duration of exposure, it can cause immunosuppression or immunostimulation. Animals can be poisoned by eating feed containing mycotoxins. Pigs are most sensitive to the emetic effect of DON, and the minimum amount of oral vomiting is 0.1-0.2 mg / kg. The main toxin-producing bacteria are Fusarium graminearum and Fusarium nivalis Bacteria ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01D15/38B01J20/281G01N1/34
Inventor 张彦明
Owner 山东美正生物科技有限公司
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