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Microfluidic chip for detecting microcystins in water and detection method

A microcystin and microfluidic chip technology, applied in the direction of analytical materials, instruments, etc., can solve the problems of limited routine monitoring times, complicated preparation process, high detection cost, etc., and achieve fast detection speed, simple operation and high degree of automation Effect

Active Publication Date: 2018-05-25
杭州绿洁科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] At present, the research on the detection technology of algal toxins in water is still relatively backward
Most algal toxins are detected by expensive and bulky HPLC-MS or GC-MS. The sample preparation process is complicated and time-consuming, requiring professional and technical personnel to operate, and the detection cost is high.
thus limiting the number of routine monitoring

Method used

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  • Microfluidic chip for detecting microcystins in water and detection method
  • Microfluidic chip for detecting microcystins in water and detection method

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Embodiment 1 (preparation of microfluidic chip)

[0036] The glass substrate was sequentially treated with concentrated sulfuric acid, a mixed solution of isopropanol and acetone (1:1) and ethanol, rinsed with deionized water, blown dry with a nitrogen gun, and removed the remaining water in an oven. Coat a layer of Cr on the surface of the glass substrate, and evenly cover a layer of photoresist with a gluing machine; spin-coat RZJ-340 positive photoresist on the treated substrate, and bake it on a hot plate to evaporate it. Residual solvent in photoresist. The substrate is subjected to ultraviolet exposure, development and corrosion. Before etching, immerse the glass piece in a chromium etching solution for etching and harden the film to improve the adhesion between the photoresist and the sacrificial layer. The wet etching is carried out at room temperature, and the etching solution contains HF and HNO 3 of etchant. In order to prolong the resistance time of the p...

Embodiment 2

[0038] Embodiment 2 (coupling of microcystin)

[0039] In this example, the microcystin on the surface of the magnetic beads is coupled by coating the original MC-LR-OVA (a conjugate of MC-LR (microcystin-LR) and OVA (ovalbumin)) . The preparation method of MC-LR-OVA comprises the following steps: take EDC (1-(3-dimethylaminopropyl)-3-ethylcarbodiimide) 100mg, and use 10mmol / L PBS solution 2.5ml of pH=7.0 Make it fully dissolved; take 10mg of MC-LR, dissolve it with 2ml deionized water, take 25mg of blood ovalbumin, dissolve it in PBS solution with pH=7.0, 10mmol / L, mix MC-LR solution with ovalbumin solution , add 2ml of EDC solution dropwise under magnetic stirring, stir at room temperature for 1 hour in the dark, then add the remaining EDC solution dropwise, stir at 4°C for 12 hours and let it stand for 10 hours, then fully dialyze it with distilled water for about 48 hours to obtain MC -LR-OVA.

[0040] The prepared MC-LR-OVA is coupled with immunomagnetic beads, and the...

Embodiment 3

[0042] Embodiment 3 (preparation of microcystin antibody)

[0043] The preparation method of microcystin antibody comprises the following steps: (1) dissolving 0.5mg MC-LR in 2mL, 0.1mol / L carbonate buffer solution of pH=9; weighing 170.4mg β-mercaptoethylamine and adding to the above solution , fully shake and shake, 50 ℃ water bath for 1-1.5h. After the reaction is completed, the solution temperature drops to room temperature, and an acetic acid solution equimolar with β-mercaptoethylamine is added to terminate the reaction;

[0044] (2) Dissolve H2N-MC-LR in 3mL 0.01mol / L PBS with pH=7.4, add 500μL of 1.25% glutaraldehyde solution, and shake vigorously in the dark for more than 3h; according to BSA: H2N-et-MC-LR =10:1 Add 0.5mL of 10mg / mL BSA, shake and mix thoroughly, and shake overnight at 4°C to complete the reaction; filter with an ultrafiltration tube with a molecular weight of 30,000Da, the supernatant solution is the prepared complete antigen, and the conjugated Th...

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Abstract

The invention discloses a microfluidic chip for detecting microcystins in water. The microfluidic chip comprises a substrate made of organic glass and a cover sheet, wherein the cover sheet is thermally bonded on the substrate; the substrate is carved with a micro flow path; the micro flow path comprises a reagent channel, a circular detection pool and a waste liquid draining channel in sequence;the reagent channel comprises an immunomagnetic bead channel, a PBST (Phosphate Buffered Saline Tween-20) buffer solution channel, a microcystins antibody channel, a fluorescein-marked second antibodychannel, a blank sample channel, a water sample channel and an antigen antibody dissociating agent channel; immunomagnetic beads are fixed in the detection pool through a micro circuit and magnets; the surfaces of the immunomagnetic heads are coupled with the microcystins. The invention further discloses a detection method for detecting the microcystins in the water by the microfluidic chip. Theanalyzing method has the advantages of high detection speed, high sensitivity, low reagent consumption, high system concentration degree, high automation degree, easiness in operation, low cost and the like.

Description

technical field [0001] The invention relates to the field of environmental monitoring, in particular to a microfluidic chip for detecting microcystin in water and a detection method. Background technique [0002] With the acceleration of social industrialization, human beings discharge a large amount of pollutants containing nitrogen and phosphorus into water bodies in industrial and agricultural production and daily life, which accelerates the eutrophication of lakes and algae Rich nutrition and mass reproduction. [0003] In the 1980s, a comprehensive investigation of the water quality of water sources nationwide was conducted, and the results showed that more than half of the lakes in the 34 lakes were in a state of eutrophication. Entering the 1990s, the eutrophication of freshwater bodies in the country has become increasingly serious, and the scope of involvement has continued to expand. [0004] 25% to 70% of the cyanobacterial bloom pollution in the world can produ...

Claims

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Application Information

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IPC IPC(8): G01N35/00
CPCG01N35/00029G01N2035/00158
Inventor 崔海松毛芳芳魏峰张立鹏周剑伟
Owner 杭州绿洁科技股份有限公司
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