Method and kit for detecting respiratory infectious disease viruses based on liquid chip

A liquid-phase chip detection and liquid-phase chip technology, which is applied in the field of detection of respiratory infectious disease viruses, can solve the problems of false negative results, more materials required for detection, and low sensitivity

Active Publication Date: 2021-03-26
湖北新纵科病毒疾病工程技术有限公司
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Problems solved by technology

The pathogen culture and isolation detection method is a traditional detection technology, the detection result is accurate and reliable, but the sensitivity is low, the detection cycle is long, which is not conducive to the early diagnosis of viral infection; the sensitivity and specificity of immunological detection are strong, but The detection requires more materials and the results of the determination are related to the source and affinity of the antibody; the nucleic acid PCR detection method is a detection method with high sensitivity at present, and the detection rate is higher than that of the pathogen isolation method, but when the PCR target sequence mutation occurs prone to false negative results

Method used

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  • Method and kit for detecting respiratory infectious disease viruses based on liquid chip
  • Method and kit for detecting respiratory infectious disease viruses based on liquid chip
  • Method and kit for detecting respiratory infectious disease viruses based on liquid chip

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preparation example Construction

[0104] (2) Preparation of the liquid phase chip: coupling the fluorescence-encoded microspheres with the tag probes in step (1) and the double-binding probe described in the step (1) to obtain the liquid phase chip.

[0105] (3) Specimen testing:

[0106] Use the PCR primers described in step (1) to amplify the target gene to obtain the amplified product, and then hybridize with the fluorescent reporter and the liquid chip obtained in step (2) to obtain the hybridization product, which is passed through the flow cytometer Read out the test result.

[0107] The above PCR amplified sequence, Tag probe and double binding probe sequence are respectively specific conserved sequences for different viruses. The present invention is based on parainfluenza virus type 1 (PIV1), parainfluenza virus type 2 (PIV2), parainfluenza virus type 1 (PIV2), novel coronavirus (N gene and Rd gene of 2019-nCoV), influenza A virus (INFA), influenza B virus (INFB) and GAPDH were human reference genes...

Embodiment 1

[0126] 1. Liquid phase chip preparation

[0127] Take out 40u1 (1×10 5 1) The surface carboxyl-modified NovaStar magnetic fluorescent microspheres developed by Wuhan Xinzongke Virus Disease Engineering Technology Co., Ltd., centrifuged at 12000rpm for 2min, discarded the supernatant, and added 5u1 0.1M, pH 4.5 MES solution (2-(N -morpholin generation) ethanesulfonic acid), and mix to obtain a coupling system. Dilute the Tag probe solution to 0.1 mM, and add 1 ul to the coupling system. Then add 2.5u1 l0mg / ml EDC (dichloroethane), mix well and place it in the dark for 30min. Add 2.5u1 l0mg / ml EDC again, mix well and place in the dark for 30min. Wash once with 0.2m1 volume percentage of 0.02% Tween-20 and 0.1% mass volume ratio of SDS sodium dodecyl sulfate) solution, and finally resuspend the microspheres in 10u1 1×TE (pH8.0, the substance in it Composition is 1M Tris-HCl, 1mL; 0.5mM EDTA, 0.2mL, add double-distilled water 100mL) solution, get the fluorescent microspheres c...

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Abstract

The invention relates to a method and a kit for detecting respiratory infectious disease viruses based on a liquid chip system. A Tag probe and a double-binding probe for detecting eight human respiratory infectious disease viruses are designed, the probes are respectively cross-linked and mixed with fluorescent coding microspheres with different colors to obtain the liquid chip for detecting theviruses, and the double-binding probe is combined with a target fragment to realize fluorescence detection, so that clinical virus detection and diagnosis time can be greatly shortened, diagnosis efficiency and diagnosis accuracy are improved, in addition, the flux is high, and the market potential is large.

Description

technical field [0001] The invention relates to the technical field of detecting respiratory infectious disease viruses, in particular to a method and a kit for detecting respiratory infectious disease viruses based on a liquid phase chip. Background technique [0002] At present, the conventional methods for clinical virus detection mainly include: cultivation and isolation of pathogens, immunological detection and nucleic acid PCR detection. The pathogen culture and isolation detection method is a traditional detection technology, the detection result is accurate and reliable, but the sensitivity is low, the detection cycle is long, which is not conducive to the early diagnosis of viral infection; the sensitivity and specificity of immunological detection are strong, but The detection requires more materials and the results of the determination are related to the source and affinity of the antibody; the nucleic acid PCR detection method is a detection method with high sens...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/686C12Q1/6837C12R1/93
CPCC12Q1/701C12Q1/686C12Q1/6837C12Q2600/16C12Q2600/166C12Q2537/143C12Q2545/101C12Q2563/107Y02A50/30
Inventor 王华林
Owner 湖北新纵科病毒疾病工程技术有限公司
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