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GB/T 38475-2020 English PDF (GBT38475-2020)

GB/T 38475-2020 English PDF (GBT38475-2020)

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GB/T 38475-2020: Detection of mycotoxins in pigment -- Rapid quantitative method of colloidal gold strip test

This Standard specifies a method for the detection of mycotoxins in natural pigment products by rapid quantitative method of colloidal gold strip test. This Standard is applicable to the rapid quantitative detection of aflatoxin B1, citrinin, zearalenone and vomitoxin in natural pigment products based on natural product extraction and fermentation.
GB/T 38475-2020
NATIONAL STANDARD OF THE
PEOPLE REPUBLIC OF CHINA
ICS 07.080
A 21
Detection of Mycotoxins in Pigment ?€? Rapid Quantitative
Method of Colloidal Gold Strip Test
ISSUED ON: NOVEMBER 19, 2020
IMPLEMENTED ON: NOVEMBER 19, 2020
Issued by: State Administration for Market Regulation;
Standardization Administration of the PEOPLE Republic of China.
Table of Contents
Foreword ... 3
1 Scope ... 4
2 Normative References ... 4
3 Principle ... 4
4 Reagents and Materials ... 4
5 Apparatus ... 6
6 Preparation of Sample ... 6
7 Determination ... 7
8 Analysis of Results ... 7
9 Repeatability ... 8
10 Others ... 8
Appendix A (Normative) Performance Requirements for Colloidal Gold Rapid Detection Test Strip ... 9
Detection of Mycotoxins in Pigment ?€? Rapid Quantitative
Method of Colloidal Gold Strip Test
1 Scope
This Standard specifies a method for the detection of mycotoxins in natural pigment products by rapid quantitative method of colloidal gold strip test.
This Standard is applicable to the rapid quantitative detection of aflatoxin B1, citrinin, zearalenone and vomitoxin in natural pigment products based on natural product extraction and fermentation.
2 Normative References
The following documents are essential to the application of this document. For the dated documents, only the versions with the dates indicated are applicable to this document; for the undated documents, only the latest version (including all the amendments) is applicable to this document.
GB/T 5491 Inspection of Grain and Oilseeds - Methods for Sampling and Sample Reduction
GB/T 6682 Water for Analytical Laboratory Use - Specification and Test Methods 3 Principle
The biotoxin in the specimen extract competes to combine with the specific antibody labeled with colloidal gold particles in the detection reagent strip during the chromatography process, and a color reaction occurs; and the color depth is related to the content of the biotoxin in the specimen. Use the reader to verify the color depth of the detection line and the quality control line on the test strip; and automatically calculate the biotoxin content in the specimen according to the color depth and the built-in curve of the reader.
4 Reagents and Materials
Unless otherwise specified, all the used reagents are analytically pure. 4.1 Water
GB/T 6682, Class-II water.
4.2 Aflatoxin B1
Purity???98%.
4.3 Citrinin
Purity???98%.
4.4 Zearalenone
Purity???98%.
4.5 Vomitoxin
Purity???98%.
4.6 Mixture of benzene + acetonitrile
Take 98.0 mL of benzene, add 2.0 mL of acetonitrile, and mix well.
4.7 Dilution buffer
Take 2.4228g of Tris(hydroxymethyl)aminomethane; add water to dissolve; adjust pH=8.0. Add 9.0g of sodium chloride, 5mL of TritonX-100; make constant volume to 1000mL by water; or use similar commercial products.
4.8 70% (volume fraction) methanol solution extract
Take 70.0 mL of methanol and add 30.0 mL of water, and mix well.
4.9 PBS buffer solution
Take 8.0 g of sodium chloride, 1.2 g of disodium hydrogen phosphate, 0.2 g of potassium dihydrogen phosphate, and 0.2 g of potassium chloride; dissolve in water; adjust the pH to 7.0; and make constant volume to 1000mL by water.
4.10 0.1% (volume fraction) Tween 20-PBS solution
Pipette 1 mL of Tween 20 and make constant volume to 1000 mL by PBS buffer solution. 4.11 0.1% (volume fraction) phosphoric acid solution
Pipette 1mL of phosphoric acid accurately; add water to make constant volume to 1000mL. 4.12 0.1mg/L biotoxin standard stock solution
filter, which is the solution to be tested.
6.2.2 Liquid samples
The liquid sample to be tested is treated with a homogenizer for no less than 2 min. After fully mixing, 1.0 mL is pipetted respectively with a micropipette and added to two 2.0 mL centrifuge tubes. One portion is sealed and frozen, and the other portion is used for detection. Pipette 200??L of centrifuged liquid sample from the centrifuge tube to be tested and put it in another centrifuge tube; add 1.8 mL of dilution buffer; and mix thoroughly for testing. 6.2.3 Purification
Connect the immunoaffinity column under the 10 mL glass syringe; accurately pipette 10.0 mL of the above sample filtrate through the immunoaffinity column; and pass it through the affinity column at a flow rate of 1 drop/s~2 drops/s. Add 10 mL of 0.1 % Tween 20-PBS solution; rinse the column at a flow rate of 1 drop/s~2 drops/s until the air enters the affinity column; discard all the effluent. Accurately add 1.0 mL of eluent for elution; and the elution flow rate is 1 drop/s~2 drops/s. Collect all the eluates in a centrifuge tube for later use. NOTE: Purification treatment is only for pigment products such as monascus red that cause serious background interference to the colloidal gold reaction results.
7 Determination
7.1 After taking out the test strip stored at low temperature, place it at room temperature for 20min~30min.
7.2 Colloidal gold reagent strip: Immerse the test strip in a centrifuge tube containing 300??L to 500??L of the solution to be tested; and incubate at room temperature for 5 min. 7.3 Colloidal gold detection card: Add 60??L to 80??L of the solution to be tested into the detection port; and incubate at room temperature for 5 min.
7.4 It should choose the colloidal gold test strip or the reader of the test card manufacturer to be used together.
8 Analysis of Results
8.1 After the reaction, take out the colloidal gold immunochromatography test strip or test card to observe the color development of C line (quality control line) and T line (test line). If the C line does not appear, or the C line appears but is diffused or severely inhomogeneous, or the C line appears but the T line is diffused or severely inhomogeneous, it is regarded as an invalid test.

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