Microfluidic chip for multiple loop-mediated isothermal amplification (LAMP) detection and preparation method thereof

A microfluidic chip and chip technology, which is applied in the field of temperature amplification, can solve the problem of not being able to detect multiple targets at the same time, and achieve the effects of being conducive to popularization and application, satisfying effective and rapid diagnosis, and intuitive results.

Inactive Publication Date: 2011-09-28
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the "isothermal mode" of the LAMP reaction and its complex nucleic acid probes make it impossible to realize the simultaneous detection of multiple targets in a sample, that is, multiple detection (ie, the simultaneous detection of multiple targets in a sample)

Method used

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  • Microfluidic chip for multiple loop-mediated isothermal amplification (LAMP) detection and preparation method thereof
  • Microfluidic chip for multiple loop-mediated isothermal amplification (LAMP) detection and preparation method thereof
  • Microfluidic chip for multiple loop-mediated isothermal amplification (LAMP) detection and preparation method thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0027] Example 1 Rapid typing detection of influenza A virus (Flu A virus), seasonal influenza A virus (Seasonal flu A virus) and 2009 swine-derived influenza A virus (Pandemic flu A virus).

[0028] The specific design size of the chip: the size of the amplification pool is (length: 10 mm, width: 0.6 mm, depth: 0.8 mm); the size of the capillary channel is (length: 2 mm, width: 0.1 mm, depth: 0.1 mm).

[0029] The specific LAMP probes of the three influenza viruses were respectively coated on the 1st-4th amplification pool 1(1'), 2(2'), 3(3'); the amplification pool 4(4') A positive control LAMP probe was applied; the fifth amplification pool 5 (5') was not coated with a probe, and served as a blank control. We use this functional chip to detect and type three important influenza viruses.

[0030] Take 4 μL of the three virus standard solutions and clinical reference products respectively, add them to the sample wells of the chip, then add 46 μL of LAMP amplification solutio...

Embodiment 2

[0036] Embodiment 2, the typing detection of eight kinds of important swine disease viruses

[0037] The specific size of the chip is the same as above, LAMP multi-chip amplification pools 1-8 are respectively coated for porcine foot-and-mouth disease virus (FMDV), swine fever virus (CSFV), porcine blue ear disease virus (PRRSV), porcine transmissible gastroenteritis Virus (TGEV), porcine pseudorabies virus (PRV), porcine parvovirus (PPV), porcine circovirus (PCV), swine influenza virus (SIV) LAMP-specific nucleic acid probes for eight important porcine disease viruses, forming an eight Functional typing chip of swine disease virus; No. 9 amplified pool was coated with other probes and used as a negative control; No. 10 amplified pool was not coated with probes and used as a blank control.

[0038] Add sample 1 (FMDV), sample 2 (CSFV), sample 3 (PRRSV), sample 4 (TGEV), sample 5 (PRV), sample 6 (PPV), sample 7 (PCV), Sample 8 (SIV), Sample 9 (FMDV, CSFV), Sample 10 (FMDV, CSF...

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Abstract

The invention belongs to the technical field of nucleic acid isothermal amplification, and particularly relates to a microfluidic chip for multiple loop-mediated isothermal amplification (LAMP) detection and a preparation method thereof. The chip is made of a high polymer, and is prepared by a micro-electromechanical system (MEMS) method. The chip mainly comprises amplification pools which are spatially and sequentially arranged, capillary channels, and connecting pipelines, wherein the amplification pools realize the effective discrimination of multiple LAMP signals through the spatial discrimination of the signals; the capillary channels are used for preventing the intersection and mixing of LAMP primers, amplification products, byproducts and the like among different amplification pools; and the amplification pools and the capillary channels are communicated through the connecting pipelines, so that fluid steadily and uniformly flows into the amplification pools from the capillary channels. The chip is easy to manufacture and operate; and an effective scheme is provided for the realization of the synchronous clinical detection of various pathogens by an LAMP method.

Description

technical field [0001] The invention belongs to the technical field of nucleic acid isothermal amplification, and in particular relates to a microfluidic chip for multiple LAMP detection and its preparation method and application. technical background [0002] Isothermal nucleic acid amplification is relative to polymerase chain reaction (Polymerase chain reaction, PCR) variable temperature nucleic acid amplification. This technology can amplify a large amount of nucleic acid at a constant temperature without relying on expensive instruments such as a temperature cycler. In recent years, it has been highly valued by academia and industry, and various types of isothermal nucleic acid amplification have been developed one after another. Loop-mediated isothermal nucleic acid amplification (Loop-mediated isothermal amplification, LAMP) was first invented by Japanese scientists. This technology has the advantages of high sensitivity, good specificity, fast response speed, and the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12M1/34C12Q1/68
Inventor 方雪恩陈惠孔继烈
Owner FUDAN UNIV
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