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Kit for detecting respiratory pathogens of community-acquired pneumonia

A technology for acquired pneumonia and respiratory tract, which is applied in the field of biomedical detection and can solve problems such as complex pathogen spectrum

Active Publication Date: 2019-07-09
广州市宝创生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0022] Due to the complex pathogenic spectrum and high incidence of CAP disease, there are very few nucleic acid detection products for multi-link detection. For the early diagnosis of CAP respiratory diseases, there are still many problems that need to be solved urgently: (1) The common respiratory pathogens of CAP are diverse. More than 20 common pathogens have been reported, and most cases are mixed infections, so the detection kit is required to realize the parallel detection of more than 20 pathogens without cross-interference between species, and has high sensitivity and specificity. There is no similar product or invention patent yet; (2) In addition to bacteria, common respiratory pathogens of CAP are mostly RNA viruses, DNA viruses, mycoplasma and chlamydia, etc., so the detection kit is required to have the ability to detect RNA and DNA nucleic acids at the same time, multi-indicator (3) Most CAPs are acutely onset and concentrated outbreaks, so the detection kits are required to have the ability of simple operation, rapid detection and high-throughput detection; (4) ) The variability of virus antigens is extremely strong, and the types of virus subtypes are increasing day by day, so the detection gene region, primers and probes of the detection kit are required to have good coverage and specificity

Method used

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  • Kit for detecting respiratory pathogens of community-acquired pneumonia
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  • Kit for detecting respiratory pathogens of community-acquired pneumonia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] This embodiment provides a kit for detecting respiratory pathogens of community-acquired pneumonia, which includes a reaction solution, a primer solution, an enzyme solution, a negative quality control, a positive quality control, a hybridization chromogenic solution, a hybridization membrane strip, and Desiccant; reaction solution includes PCR buffer solution, metal cations and purified water, enzyme solution includes reverse transcriptase, hot start Taq enzyme and UNG enzyme, negative quality control product is sterile normal saline, positive quality control product is for the purpose of detecting pathogens The cloning bacteria liquid or pseudovirus particles of the amplified sequence, the base sequence of the amplified sequence of the pathogen target is shown in SEQ ID NO.60-83; the concentration of metal cations is 1-3mM, and the concentration of Taq enzyme is 0.5-5U, The concentration of UNG enzyme is 0.05~0.5U, the concentration of cloning bacteria is 1×10 4 cfu / m...

Embodiment 2

[0097] Specifically, using the detection kit described in the above-mentioned embodiment 1, this embodiment shows the detection data of three groups of clinical samples. Clinical samples are nasopharyngeal swabs, alveolar lavage fluid or sputum samples from CAP patients with known pathogen infection (see Table 7: " / " in Table 7 indicates no clinically confirmed infection pathogen type, and these samples are negative ). The pretreatment of clinical samples is as follows: (1) nasopharyngeal swab sample: put the swab sample into 1.0ml of normal saline, wash it thoroughly, squeeze it dry, discard the swab, centrifuge at 12000rpm for 5min, discard the supernatant, and leave the precipitate Standby; (2) Alveolar lavage fluid sample: take 1.0ml sample, centrifuge at 12000rpm for 5min, discard the supernatant, then add 1.0ml of normal saline to wash once, centrifuge at 12000rpm for 5min, discard the supernatant, and save the precipitate for later use; (3) Sputum sample: a. Add 2 to 3...

Embodiment 3

[0115] In order to prove the rationality and feasibility of the present invention, the following performance evaluation research has been carried out to the detection kit:

[0116] (1) Minimum detection limit experiment

[0117] Preparation of reference products: purchase standard reference products from the Guangdong Microbial Culture Collection Center (see the table below), extract the nucleic acid of the reference product, measure the nucleic acid concentration, convert the copy number according to the relevant formula, and then use 0.2×TE to test the nucleic acid sample Serial dilution and detection.

[0118] Reference Detection concentration gradient (copies / ml) SA 10 4 、10 3 、500、100

PA 10 4 、10 3 、500、100

IFB 10 4 、10 3 、500、100

[0119] The standard reference substance of each concentration gradient is detected by the detection method of the detection kit in the above-mentioned embodiment, and the membrane strip detection resu...

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Abstract

The invention discloses a kit for detecting respiratory pathogens of community-acquired pneumonia, and belongs to the technical field of biomedical detection. The kit comprises a reaction solution, aprimer solution, an enzyme solution, a negative quality control substance, a positive quality control substance, a hybrid color developing solution, a hybrid membrane strip and a desiccant, wherein aprobe is attached to the hybrid membrane strip, the hybrid color developing solution comprises microspheres connected with a second coloring marker, and the color is developed after the microspheres are enriched. The detection kit involves 24 common respiratory pathogens of CAP, and has the advantages of high specificity, high sensitivity, large number of detection targets, simple and rapid operation and the like.

Description

technical field [0001] The invention belongs to the technical field of biomedical detection, in particular to a kit for detecting respiratory pathogens of community-acquired pneumonia. Background technique [0002] At present, the morbidity and mortality of infectious diseases have been high, among which respiratory tract infection ranks first among all kinds of infectious diseases. Community-acquired pneumonia (CAP) refers to the infectious lung parenchyma (including alveolar wall, that is, pulmonary interstitium in a broad sense) inflammation suffered outside the hospital, including pathogen infection with a clear incubation period. Pneumonia of internal onset. Globally, CAP is one of the most common respiratory infectious diseases that seriously threaten human health, with a wide range of transmission, high morbidity and high mortality. According to data from foreign CAP monitoring networks, the 30-day mortality rate of adult CAP patients is 8.6 %, the mortality rate of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/689C12Q1/04C12N15/11
CPCC12Q1/689C12Q1/701
Inventor 肖慧杨祥胜潘腾飞杨伊莹
Owner 广州市宝创生物技术有限公司
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