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A kind of quantitative detection kit of almond protein and preparation method thereof

A quantitative detection and almond kernel technology is applied in the field of quantitative detection of almond protein, which can solve the problems of inability to specifically distinguish almond kernels from almonds, hidden dangers of food safety and omission of labels for people allergic to almond kernels, and achieves good anti-matrix interference effect. , convenient screening, good repeatability

Inactive Publication Date: 2016-05-04
SHENZHEN ACAD OF METROLOGY & QUALITY INSPECTION
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, because it is highly similar to almonds, it is often confused with almonds, and merchants often omit or mislabel it on the labels, which poses a great food safety hazard to people allergic to almonds
However, due to the damage of DNA during thermal processing and the close relationship between the two, the current PCR method cannot specifically distinguish almonds and almonds in thermally processed products.

Method used

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  • A kind of quantitative detection kit of almond protein and preparation method thereof
  • A kind of quantitative detection kit of almond protein and preparation method thereof
  • A kind of quantitative detection kit of almond protein and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Preparation of target antigen.

[0024] According to the protein sequence of NCBI almond pruin-1, Blast comparison was carried out, comprehensive specificity, antigenicity analysis, synthetic peptide (amino acid sequence: RQGRQQGRQQQEEGR-Cys) coupled with keyhole limpet hemocyanin as immunogen, coupled with carrier protein bovine serum Albumin was used as the coating source. The coupling method is as follows:

[0025] Dissolve 5.8 mg of 3-(2-pyridyldithiol)propionic acid N-hydroxysuccinimide ester in 1 mL of dimethyl sulfoxide, and gradually add it dropwise to 1 mL of 0.1 g of keyhole limpet hemocyanin or bovine serum albumin. 01MpH7.4PBS, react at room temperature for 12 hours. Free N-hydroxysuccinimide 3-(2-pyridyldimercapto)propionate was removed by dialysis overnight. Add 4 mg of polypeptide to the above-mentioned activated protein solution. After 12 hours of reaction, it was dialyzed overnight and stored in freeze-dried condition.

Embodiment 2

[0026] Example Preparation and purification of secondary anti-almond prunin-1 protein antibody.

[0027] 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.

[0028] 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 bovine serum t...

Embodiment 3

[0035] Example 3 Preparation of almond prunin-1 protein standard.

[0036] After crushing the almond pulp with a grinder, 100 g of the sample was taken, and 8M urea was used for ultrasonic extraction for 6 hours. Take the supernatant by centrifugation, use a liquid phase focuser for primary separation and purification, and intercept components with an isoelectric point of 5-6.

[0037] 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 40kd was cut out with reference to the pre-stained protein standard. After crushing the protein gel with a mortar, 10 mL of 7M urea was added for extraction overnight. After centrifugation, the supernatant was dialyzed and free...

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Abstract

The invention provides a preparation method of a quantitative determination kit for almond protein. The preparation method comprises the following steps: (S1) preparing monoclonal antibodies; (S2) extracting and separating almond pruning-1 protein; (S3) establishing and evaluating by virtue of an enzyme-linked immunosorbent assay; and (S4) assembling the quantitative determination kit. The invention further provides the quantitative determination kit for the almond protein. The quantitative determination kit has the beneficial effect that the almond protein in foods can be quantitatively determined.

Description

technical field [0001] The invention relates to the quantitative detection of almond protein, in particular to a quantitative detection kit of almond protein and a preparation method thereof. Background technique [0002] Almond, commonly known as almond, is a popular nut produced all over the world. It is rich in protein, dietary fiber, vitamin E, and does not contain cholesterol. It is accepted and loved by consumers in China. However, driven by profit, unscrupulous food producers often use additives such as almond essence to make almond protein drinks. The root cause of this confusion and doubt is the lack of quantitative methods for vegetable protein contained in vegetable protein beverages. The national standard GB16322-2003 "Hygienic Standards for Vegetable Protein Beverages" stipulates that the protein content in vegetable protein beverages should not be less than 5g / L. Due to technical limitations, the total protein can only be quantified according to the Kjeldahl ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/531
CPCG01N33/531G01N33/6818G01N2333/415
Inventor 赖心田张世伟黄静敏刘小青杨国武王士峰冯荣虎唐栋陈极锋
Owner SHENZHEN ACAD OF METROLOGY & QUALITY INSPECTION
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