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Nosocomial infection multiple causative agent parallel detection gene chip and preparation method and application thereof

A technology for detecting gene chip and nosocomial infection, applied in the field of molecular biology technology and clinical detection, can solve the problems of inability to reduce medical expenses and increase costs, and achieve rapid and sensitive detection, high accuracy, specificity and repeatability. Effect

Active Publication Date: 2013-08-21
SHANDONG ACV BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This type of diagnosis does not reduce medical costs, but increases them

Method used

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  • Nosocomial infection multiple causative agent parallel detection gene chip and preparation method and application thereof
  • Nosocomial infection multiple causative agent parallel detection gene chip and preparation method and application thereof
  • Nosocomial infection multiple causative agent parallel detection gene chip and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Embodiment 1: the extraction of sample nucleic acid

[0073] RNA was extracted from clinical specimens using QIGEN Viral RNA mini Extraction Kit(CAT:52904). A total of 43 nucleic acid samples were extracted, of which 28 were positive samples with known subtypes, 10 were negative controls, and 5 were unrelated virus samples. Specimen information is as follows:

[0074]

[0075] (1) Add 560 μl of AVL buffer to a 1.5ml centrifuge tube;

[0076] (2) Add 140 μl clinical specimens to this centrifuge tube;

[0077] (3) Place at room temperature for 10 minutes, then centrifuge briefly;

[0078] (4) Add 560 μl absolute ethanol, mix thoroughly for at least 15 seconds, and centrifuge briefly;

[0079] (5) Carefully add about 630 μl of the mixed solution to the QIAamp column, put the QIAamp column on the collection tube, and then centrifuge at 8000rpm for 5s to allow the liquid to pass through the filter membrane;

[0080] (6) Discard the liquid in the collection tube and r...

Embodiment 2

[0087] Embodiment 2: PCR amplification and hybridization

[0088] Add the above-mentioned sample nucleic acid (one cartridge to detect one sample, and a blank control is also provided as needed. To test the repeatability of the reagent, each sample is repeated three times) from the sample addition hole into the cartridge, and the cartridge All reagents required for PCR amplification, hybridization detection, and cleaning are pre-installed, and the prepared nosocomial infection pathogen gene chip is also pre-placed in the hybridization detection tank at the bottom of the cartridge. Put the cartridge into the cartridge processor, write the reaction program from the system control software, run it, and the instrument will automatically perform PCR amplification and hybridization.

[0089] (1) Two rounds of PCR amplification were performed. The reagent used in the first round of PCR amplification was Qiagen one-step RT-PCR Kit, Cat No. 210212. The configuration of the reaction sys...

Embodiment 3

[0101] Example 3: Signal Detection

[0102] After the reaction is complete, take the cassette out of the processor and put it into the cassette reader. The cassette reader has detection devices such as a laser confocal scanning system and a photomultiplier tube. The read data is processed by the system analysis software and directly converted to The digital form is displayed, and the cartridge is thrown away directly after it is taken out of the reader. The closed system prevents pollution from occurring. The test results are as follows:

[0103]

[0104]

[0105]

[0106]

[0107]

[0108]

[0109]

[0110] Result analysis:

[0111] (1) The reagent can accurately type 28 positive bacterial nucleic acid samples with a specificity as high as 100%.

[0112] (2) The repeatability is good, 90% of the detection data intra-assay coefficient of variation (CV) <10%.

[0113] (3) 10 known negative samples were tested, and the results showed that there were no fa...

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Abstract

The invention belongs to the technical fields of molecular biological techniques and clinical tests, and particularly discloses a nosocomial infection multiple causative agent parallel detection gene chip and a preparation method and application thereof. The gene chip can be used for rapidly and sensitively detecting fourteen medical treatment infection typical causative agents such as acinetobacter baumannii, enterobacter cloacae, enterococcus faecalis and the like by using one reaction, and the detection data of coinfection of yeast and bacterium which cannot be provided by a traditional amplification method is offered. Furthermore, the gene chip also comprises a methicillin resistance target spot, thus realizing further detection of staphy lococcus infection, therefore, a clinician can realize which causative agent a patient is infected in one or two hours, and realize whether the patient has drug resistance, so as to decide whether the patient needs antibiotics or which antibiotics is needed, so that the gene chip has a greater clinical guiding significance.

Description

(1) Technical field [0001] The invention belongs to the field of molecular biology technology and clinical detection technology, and in particular relates to a gene chip for parallel detection of nosocomial infection multi-pathogens and its production method and application. (2) Background technology [0002] In recent years, with the development of medicine, the rate of nosocomial infection has been on the rise, which has attracted the attention of hospitals all over the world. Therefore, it has become a comprehensive research topic to put forward monitoring measures for nosocomial infection. Foreign advanced countries carried out research on nosocomial infection 30 years earlier than my country. The rate of nosocomial infection in my country is 9% to 20%. The health administrative departments at all levels in our country attach great importance to it. The Ministry of Health established 134 hospitals nationwide to participate in nosocomial infection monitoring in only 3 yea...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12Q1/04C12R1/01C12R1/445C12R1/19C12R1/12
Inventor 李艳艳张凯宁
Owner SHANDONG ACV BIOTECH CO LTD
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