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Rapid honey detection test strip, and preparation method and application thereof

A technology for detecting test strips and test strips, which is applied to measurement devices, instruments, scientific instruments, etc., can solve the problems of inability to identify the true and false honey, long time consumption, etc., and achieve the effects of short detection time, low preparation cost, and easy use.

Inactive Publication Date: 2017-12-12
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In July 2015, the State Food and Drug Administration officially responded on its official website: my country's current national standards for honey cannot identify the authenticity of honey, and will actively promote the revision of the national standard for honey food safety and the establishment of supplementary testing methods for honey adulteration
[0023] CN105693858A discloses a preparation method, detection method and kit for MRJP1 antibody pairs in honey, but generally requires personnel with professional background to operate, and takes a long time

Method used

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  • Rapid honey detection test strip, and preparation method and application thereof
  • Rapid honey detection test strip, and preparation method and application thereof
  • Rapid honey detection test strip, and preparation method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0046] Example 1: Preparation and purification of specific polyclonal antibodies SP-1 and SP-2 against MRJP1

[0047] Through the homology analysis of the amino acid sequences of the MRJPs protein family, two specific oligopeptides of MRJP1 that are different from other family members were screened out, namely oligopeptide-1 SGEYDYKNNYPSDID and 360th oligopeptides located at positions 56-70 of the amino acid sequence of MRJP1 - IKEALPHVPIFD oligopeptide-2 at position 372, there is no repeated continuous sequence between these two polypeptides. Rabbits were immunized with synthetic polypeptides of the two sequences to obtain MRJP1-specific antibodies SP-1 and SP-2;

[0048] According to the selected polypeptide sequence, chemical synthesis is carried out respectively. Conjugate 5 mg of the synthetic polypeptide with KLH (the coupling agent is Sulfo-SMCC) as the immune antigen, and couple the other 5 mg of the synthetic polypeptide with BSA (the coupling agent is glutaraldehyde...

Embodiment 2

[0051] Example 2: Determination of the potency of polyclonal antibodies SP-1 and SP-2 against MRJP1

[0052] 1. Determination of titer of polyclonal antibodies SP-1 and SP-2 against MRJP1 The specific operation steps are as follows:

[0053] (1) Coating: MRJP1 was coated with carbonate buffer (CBS, Na 2 CO 3 1.59 g, NaHCO 3 2.93 g,ddH 2 O 950mL, adjust the pH to 9.6, add ddH 2 (dilute to 1000 mL, store at 4°C) and dilute to 1 μg / mL, add to 96-well microtiter plate at 100 μL / well, overnight at 4°C;

[0054] (2) Plate washing: Discard the coating solution, place the microplate plate on the plate washer, wash the plate with PBST (PBS+0.05% Tween-20) washing buffer for 5 times, remove and spin dry;

[0055] (3) Blocking: Add 200 μL of phosphate buffer solution containing 5% skimmed milk powder to each well to block, and react at 37°C for 1.5 h;

[0056] (4) Wash the plate: Discard the blocking solution, and wash the plate with the same method as (2);

[0057] (5) Add the ...

Embodiment 3

[0067] Embodiment 3: Colloidal gold rapid detection test strip of MRJP1 polyclonal antibody in honey

[0068] 1. Preparation and identification of colloidal gold solution

[0069] Take 100 mL of chloroauric acid aqueous solution with a mass concentration of 0.01%, heat it to boiling with a magnetic stirring reflux device, quickly add 1 mL of a mass concentration of 1% trisodium citrate solution at one time, and prepare it under constant temperature reflux reaction under stirring, and continue to boil 8min, until the solution is wine red, and cooled at room temperature, a colloidal gold solution containing colloidal gold particles with a particle size of 40 nm can be prepared, and the colloidal gold solution is stored at 4°C; the fired colloidal gold is purple-red and transparent 1. Clarification: The maximum absorption peak is 532 nm as measured by an ultraviolet spectrum scanner; most particles of colloidal gold fired through electron microscopy are round, without polygons, a...

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Abstract

The invention discloses a rapid honey detection test strip, and a preparation method and application thereof. The preparation method includes (1), preparing and purifying polyclonal antibodies SP1 and SP2 capable of specifically recognizing MRJP1; (2), preparing a collaurum solution; (3), coating a nitrocellulose membrane with a quality control line (line C) and a detection line (line T); (4), assembling the rapid honey double-antibody sandwich detection test strip taking the bee endogenous protein MRJP1 as a biomarker; (5), preparing a honey detection sample. After separated and purified protein in the honey sample reacts with the gold-labeled test strip, authenticity of the honey sample can be judged according to color development conditions of the line T. The gold-labeled honey double-antibody test strip is applicable to detection of fake honey pretend by artificial syrup, and a new reliable rapid detection method is provided for quality and authenticity detection of honey.

Description

technical field [0001] The invention relates to a preparation method of a honey rapid detection test strip, the test strip and its application. Background technique [0002] Honey is a natural sweet substance that is fully brewed by bees after collecting nectar, secretions or honeydew from plants and combining them with their own secretions. As a nutritious natural sweetener and health supplement, honey is loved by consumers around the world, but it is a food that is easily adulterated. In recent years, the adulteration of honey has intensified, and adulteration has become increasingly difficult to be identified, which has seriously disrupted the order of the honey market. The 2014 online honey sampling results announced by the China Bee Products Association showed that 13 of the 20 top-selling honey products on Taobao, Tmall and JD.com were adulterated, resulting in a crisis of product trust and a decline in the overall price and efficiency of the industry. Serious impact...

Claims

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Application Information

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IPC IPC(8): G01N33/558G01N33/543
CPCG01N33/558G01N33/543
Inventor 沈立荣张一帆辛晓璇蒋晨旻王一然
Owner ZHEJIANG UNIV
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