GB/T 40220-2021.Determination of soybean agglutinin of plant metabolites-Enzyme-linked immunosorbent assay.
1 Scope
GB/T 40220 specifies the enzyme-linked immunosorbent assay method for soybean agglutinin, a plant metabolite.
GB/T 40220 is applicable to the determination of soybean agglutinin, a plant metabolite in soybean and its products.
2 Normative references
The contents of the following documents constitute the essential provisions of this document through normative references in the text. Among them, for dated references, only the version corresponding to the date applies to this document; for undated references, the latest version (including all amendments) applies to this document.
GB/T 6682 Specifications and test methods for water for analytical laboratories
3 Terms and definitions
There are no terms and definitions that need to be defined in this document.
4 Principle
After the sample is treated, the soybean lectin in the sample solution binds to the soybean lectin antibody (capture antibody) coated in the microplate. The plate is washed to remove other unbound components, and then it binds to the soybean lectin enzyme-labeled antibody. The enzyme-labeled substrate is added for color development. The absorbance value is measured at a wavelength of A50 mm using an enzyme reader to calculate the soybean lectin content.
5 Reagents and materials
Unless otherwise specified, the chemical reagents used in this method are analytically pure, and the water is secondary water specified in GB/T 6682.
5.1 Soybean lectin detection kit: see Appendix A.
5.2 Soybean lectin: purity ≥95%, CAS number 68513-95-1.
5.3 The required solution should be prepared according to the instructions of the kit.
6 Instruments and equipment
6.1 Small grinder.
6.2 Sampling sieve: aperture 0.9 mm.
6.3 Homogenizer.
6.4 Vortex mixer.
6.5 Electronic balance: sensitivity 0.01 g.
6.6 Centrifuge tube with stopper: 15 mL.
This document specifies the enzyme-linked immunosorbent assay method for soybean lectin, a plant metabolite. This document is applicable to the determination of soybean lectin, a plant metabolite, in soybeans and their products.

ICS07.080
CCSA40
National Standard of the People's Republic of China
GB/T40220—2021
Determination of soybean agglutinin of plant metaholitesEnzyme-linked imnunosorbent assay2021-05-21Release
State Administration for Market Regulation
National Standardization Administration
2021-12-01Implementation
People's Republic of China
National Standard
Determination of soybean lectin as a plant metabolite
Enzyme-linked immunosorbent assay
GB/T40220-2021
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GB/T40220—2021
Part 1, Structure and drafting rules for standardization documents" This document was drafted in accordance with the provisions of GB/T1.12020 Guidelines for standardization work.
Please note that some contents of this document may involve patents. The issuing organization of this document does not assume the responsibility for identifying patents. This document is proposed and managed by the National Technical Committee for Standardization of Biochemical Testing (SAC/TC387). The drafting units of this document: Jiangnan University, Greentown Agricultural Science Testing Technology Co., Ltd., Beijing Technology and Business University, Beijing Sambo Technology Co., Ltd.
The main drafters of this document: Gu Chuanlai, Wang Chuanpi, Wang Zhen, Ma Aijin, Wang Zhongxing, Hao Shuai, Zeng Lu, Ma Wei, 1
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1Scope
Determination of soybean lectin, a plant metabolite
Enzyme-linked immunosorbent assay
This document specifies the phenol-linked immunosorbent assay method for soybean lectin, a plant metabolite. This document applies to the determination of the total amount of soybean metabolites in soybean and its products. 2 Normative reference documents
GB/T402202021
The following documents constitute the essential clauses of this document through normative reference in the text. For the referenced documents with a date, only the version corresponding to that date applies to this document. For the referenced documents without a date, the latest version (including all amendments) applies to this document.
GB/T6682
3 Terms and definitions
Specifications and test methods for water for analytical laboratories
There are no terms and definitions that need to be defined in this document. 4 Principle
After the sample is treated, the soybean lectin in the liquid binds to the soybean lectin antibody (capture antibody) coated in the microplate. The unbound other components are removed by washing the plate, and then bind to the soybean lectin antibody. The substrate is added to develop color. The absorbance value is measured at a wavelength of 150 by a sea level analyzer to calculate the soybean lectin content. 5 Reagents and Materials
Unless otherwise specified, the chemical reagents used in this method are analytical pure water, which is secondary water specified in GB/T6682. Soybean lectin detection kit: See Appendix A5.1
Soybean lectin, purity ≥95%, CAS number 68513-95-1. 5.2
The required solution should be prepared according to the instructions of the kit. Instruments and Equipment
6.1 Small pulverizer.
6.2 Sieve: aperture 0.9mm
6.3 Homogenizer.
6.4 Vortex mixer.
6.5 Electronic balance: sensitivity 0.01g.
Stopper centrifuge tube: 15mL.
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GB/T40220—2021
6.7 Microplate reader: with 450nm filter. 6.8 Instruments required by the test kit
7 Preparation and preservation of samples
7.1 Preparation of solid samples
Weigh 100g of sample and crush it with a small crusher. The crushed sample passes through a 0.9mm sieve. Put it into a clean sealer and mark it for testing.
Preparation of semi-solid samples
Take 100g of sample, mix it thoroughly with a homogenizer, put it into a clean sealed container, mark it, and get the test. 7.3
Preparation of liquid samples
Weigh 100g of sample, mix thoroughly with a vortex mixer, put into a clean sealed container, and mark it for testing. 8 Test steps
8.1 Sample pretreatment
Weigh 1.00g of the prepared sample, place it in a 15mL stoppered centrifuge tube, add the extract required by the kit, and dilute and test according to the method described in the kit manual.
2 Determination
Perform quantitative detection of the sample (liquid) to be tested according to the operating steps of the enzyme-linked immunosorbent assay kit. 8.3
3 Parallel test
According to the above steps, the same standard solution and the same sample solution should be tested in half a row. 8.4 Blank test
Except for not weighing the sample, all the above steps are carried out 5 Positive quality control
According to the content of human bean agglutinin in the sample to be tested, select a suitable quality control standard to determine the accuracy of the test process. 9 Experimental data processing
9.1 Drawing of standard working curve for quantitative detection of enzyme-linked immunosorbent assay kit According to the calculation method or software provided in the kit manual, draw the standard working curve according to the relationship between the mass concentration of the standard and the absorbance change.
9.2 Calculation of mass concentration of the test solution
According to the calculation method and computer software provided in the kit manual, substitute the absorbance of the test solution into the standard working curve obtained in 9.1 to calculate the mass concentration (o) of the test solution. rKaeerkAca-
9.3 Calculation of results
The content of soybean agglutinin in the sample is calculated according to formula (1): X-exVxax1000
mX1000
X—the content of soybean agglutinin in the sample, in micrograms per gram (ug/kg): GB/T402202021
ot9reeepeeReeep(1)
The mass concentration of soybean agglutinin in the sample obtained from the standard working curve, in nanograms per milliliter (ng/mL): VThe volume of sample extract, in milliliters (mL), a
The dilution multiple in the pretreatment process;
The weight of the sample, in grams (g). The calculation result is expressed as the arithmetic mean of two independent determination results obtained under repeatability conditions, retaining 3 significant mathematical digits. 10 Precision
The absolute difference between two independent determination results obtained under repeatability conditions shall not exceed 20% of the arithmetic mean. 11 Accuracy
When the concentration of the soybean lectin solution spiked with the ELISA kit for testing is 100ng/mL, the average recovery rate is 76.2%~120.9%; when the concentration of the soybean lectin solution spiked with the ELISA kit for testing is 1000ng/mL, the average recovery rate is 82.7%~111.9%; when the concentration of the soybean lectin solution spiked with the ELISA kit for testing is 5000ng/ml, the average recovery rate is 89.4%~107.6%.
12 Others
When 1.00g of solid and flat samples such as soybeans and soybean paste are weighed, the detection limit of soybean lectin is 300ug/kg and the quantification limit is 500μg/kg; when 1.00g of liquid samples such as soy milk and soy milk are weighed, the detection limit of soybean lectin is 300ug/kg and the quantification limit is 500μg/kg.At c0g, the detection limit of soybean lectin is 30ug/kg, and the quantification limit is 50μg/kg.
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GB/T40220-2021bZxz.net
Appendix A
(Informative)
Soybean lectin detection kit
A, 1 Enzyme label plate coated with human soybean lectin antibody (capture antibody), sample extract: 0.01mol/LpH7.4 phosphate buffer solution. A.2
Sample diluent: 0.01mol/LpH7.4 phosphate buffer solution. A.3
Soybean lectin standard solution, 0ng/mL.20ng/mL50ng/mL100ng/mL.500ng/mL, 1000ng/mL, A.4
2000ng/ml5000ng/ml. and 10000ng/mlA5Soybean lectin enzyme-labeled antibody: soybean lectin antibody-horseradish peroxidase crosslinked. Washing liquid concentrate: can be used after dilution with water. A.6
Substrate solution.
A.8Reaction stop solution.
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