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Micro-fluidic chip for visual instant detection of pathogen nucleic acid as well as preparation method and detection method thereof

A technology of microfluidic chips and pathogens, applied in biochemical equipment and methods, methods based on microorganisms, microorganisms, etc., can solve the problems of not reaching POCT

Pending Publication Date: 2018-04-13
RENJI HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, for the nucleic acid detection of respiratory pathogens, DNA or RNA needs to be extracted from clinical samples, and corresponding detection systems are required, such as PCR analyzers, centrifuges or gel electrophoresis instruments, etc., which are limited by fixed detection sites and cannot reach POCT. requirements

Method used

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  • Micro-fluidic chip for visual instant detection of pathogen nucleic acid as well as preparation method and detection method thereof
  • Micro-fluidic chip for visual instant detection of pathogen nucleic acid as well as preparation method and detection method thereof
  • Micro-fluidic chip for visual instant detection of pathogen nucleic acid as well as preparation method and detection method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0063] This embodiment is a preferred form of microfluidic chip.

[0064] Such as figure 1and image 3 As shown, the microfluidic chip described in this embodiment includes an upper substrate 1 of PDMS material and a lower substrate 2 of glass silicon wafer material sealed with each other, and the shapes of the upper substrate 1 and the lower substrate 2 are four square. A microfluidic channel 6 is set on the upper substrate 1 , specifically, a sample processing and DNA extraction area 3 , a fluid control area 4 , and a visual detection area 5 connected sequentially through the fluid channel.

[0065] The above-mentioned sample processing and DNA extraction area 3 is a circular channel area, which communicates with the sample inlet 32, the sample waste liquid outlet 31 and the fluid control area 4 respectively through the fluid channel, and the sample inlet 32 ​​and the sample waste liquid outlet 31 are both arranged in the On the opposite side of the fluid control area 4 ,...

Embodiment 2

[0071] This embodiment is the preparation method of the microfluidic chip described in Embodiment 1, which includes the following steps:

[0072] Step 1) Use photoresist to make the male mold of the microfluidic chip: the chip is made using soft lithography technology and micromachining technology, and the AZ-50 photoresist process is used to make the chip microchannel male mold, and laser cutting is used to make the diameter PMMA polymethyl Methyl acrylate microcolumns (diameter 10mm, height 2mm) and PMMA microcolumns (diameter 3mm, height 2mm), PMMA microcolumns were pasted on the AZ-50 positive mold with shadowless glue to make the microfluidic chip positive mold ;

[0073] Step 2) Pour the PDMS casting solution on the male mold of the microfluidic chip prepared in step 1), and pre-embed a colorless and transparent PMMA screw valve (5mm in diameter, 5mm in height, and 0.7mm in pitch) in the uncured casting solution Among them, the vertical distance d between the bottom of ...

Embodiment 3

[0077] This embodiment is another preferred microfluidic chip, which is used for simultaneous detection of Mycoplasma pneumoniae and Streptococcus pneumoniae.

[0078] The rest of the structure of the microfluidic chip in this embodiment is the same as that of the microfluidic chip in Embodiment 1, and there are only differences in the following aspects: figure 2 As shown, in this embodiment, the microfluidic chip is provided with 8 detection pools 54, which are divided into 2 groups, each group has 4 detection pools 54, and each group detects different nucleic acids of pathogens, specifically 1 on the left. In the detection pool of the group, the detection pool (1' and 1") coated with the isothermal amplification primer chromatographic paper of Mycoplasma pneumoniae, the detection pool coated with the specific primer of Mycoplasma pneumoniae and the DNA chromatography paper of Mycoplasma pneumoniae (positive control P1), and the detection pool without Detection pools coated ...

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Abstract

The invention relates to a micro-fluidic chip for visual instant detection of pathogen nucleic acid. The micro-fluidic chip comprises an upper layer substrate and a lower layer substrate, wherein theupper layer substrate is provided with a micro-fluidic passage; the lower layer substrate is sealed with the upper layer substrate; the upper layer substrate is provided with a sample processing and DNA extraction region, a fluid control region and a visual detection region which are sequentially communicated; the visual detection region comprises a detection pool; the detection pool is provided with chromatography paper for covering specific primers or DNA; the fluid control region is provided with a screw valve used for performing fluid control. The invention also relates to a preparation method and a detection method of the micro-fluidic chip. The invention provides the micro-fluidic chip integrating sample treatment, pathogen nucleic acid extraction, amplification and visual detection;the simple, fast and visual beside pathogen nucleic acid detection can be realized; the detection method using the micro-fluidic chip has the characteristics of high speed, simplicity and no need ofspecial instruments; the micro-fluidic chip and the method can be applied to the instant inspection of various kinds of pathogen nucleic acids in respiratory tracts and genital tract samples.

Description

technical field [0001] The invention relates to the fields of integrated microfluidic chip manufacturing technology and gene diagnosis, in particular to a microfluidic chip for visually detecting pathogenic nucleic acid in real time and a preparation method thereof, and a method for detecting pathogenic nucleic acid using a microfluidic chip. Background technique [0002] The history of human health and medical development is a history of fighting against various infectious diseases. The emergence of each new infectious disease has caused serious social panic and economic losses. As a common and frequently-occurring disease in pediatrics, respiratory tract infection is the first cause of death in children under 5 years of age worldwide. Its clinical manifestations are easy to recognize, but the etiology of respiratory tract infection is difficult to clarify. [0003] Laboratory detection of respiratory pathogens can clarify the cause of infection, facilitate timely diagnosi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12M1/34C12M1/00C12Q1/6806C12Q1/6844C12Q1/14C12R1/46
CPCC12Q1/6806C12Q1/6844C12Q2563/143C12Q2563/149C12Q2531/119
Inventor 李敏汪骅刘倩徐燕萍秦娟秀
Owner RENJI HOSPITAL AFFILIATED TO SHANGHAI JIAO TONG UNIV SCHOOL OF MEDICINE
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