Mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection kit and application thereof

A technology for Mycoplasma pneumoniae and Chlamydia pneumoniae, which is applied in the field of kits for the joint detection of Mycoplasma pneumoniae and Chlamydia pneumoniae nucleic acids based on double amplification technology, can solve problems such as easy pollution

Active Publication Date: 2020-03-24
武汉中帜生物科技股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These methods require special PCR amplification conditions, specialized laboratories and PCR machines, and are prone to contamination during the detection process

Method used

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  • Mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection kit and application thereof
  • Mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection kit and application thereof
  • Mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] [Example 1] Sensitivity Test

[0085] The known concentrations of Mycoplasma pneumoniae and Chlamydia pneumoniae were diluted in a ten-fold concentration gradient. The pathogens at each dilution concentration were lysed with the lysate in the kit (pathogen: lysate = 1:1). Double amplification detects pathogens at each diluted concentration and assays the results. The pathogen dilutions of each gradient were separately repeated 5 times during the detection. The pathogen titer value with a positive detection rate of 90%-95% is initially used as the minimum detection limit. After the initial value of the detection limit is determined, the stock solution of each pathogen subtype is diluted to the vicinity of the initial value titer, and the pathogens of different concentrations are detected with this kit, and each titer is detected 20 times to further determine the minimum detection limit. Accurate determination (choose positive rate above 95%). The specific results are...

Embodiment 2

[0100] [Example 2] Specificity Verification

[0101] Different microorganisms were lysed with cell lysate and then double-amplified for detection to verify the specificity of the probe design of the kit. The results are shown in the table below:

[0102] Table 3 Cross-reaction verification data between other pathogenic microorganisms

[0103]

[0104] The results showed that the pathogens detected by this kit were not involved in cross-reaction with other pathogens.

Embodiment 3

[0105] [Example 3] Validation of clinical samples

[0106] 1. Clinical sample information

[0107] A total of 359 samples were tested in the Maternal and Child Health Hospital of Hubei Province, including 196 male samples and 163 female samples, accounting for 54.60% and 45.40% respectively. Among the 452 samples, the oldest patient was 62 years old, the youngest was 1 month old, the mean age was 10.63 years old, the standard deviation was 11.2 years old, and the median was 6 years old. The diagnosis results of the enrolled patients were all related to respiratory tract infection, and the specific distribution was as follows: 196 cases of suspected pneumonia, 69 cases of (acute, asthmatic) bronchitis, 37 cases of (acute) upper respiratory tract infection, 20 cases of (infectious) fever, 19 cases of influenza and 18 others.

[0108] 2. Test results

[0109] 1) Mycoplasma pneumoniae test results

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Abstract

The invention discloses a mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection kit and an application thereof. A collected sample is split by a cell lysis solution to releasepathogen nucleic acid, and amplification of pathogen nucleic acid fragments is realized through reverse transcription and transcription processes. The amplified RNA product is added into a microporecoated with a coating probe, and a specific probe and an amplification probe are added at the same time, wherein the coating probe can be combined with one end of the specific probe CES to fix the amplified product RNA. One end of the specific probe LES is combined with the RNA product, and the other end of the specific probe LES is combined with the amplification probe to realize signal amplification. The amplification probe marked with multiple biotins is then combined with a streptavidin-HRP enzyme conjugate. Finally, an HRP enzyme chemiluminiscence substrate is added, and detection is carried out on a chemiluminiscence instrument. According to the invention, RNA extraction is not needed, and pollution is not likely to happen in the detection process; and the kit has the advantages of high sensitivity and high specificity, and can be widely applied to mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid combined detection.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a kit for combined detection of mycoplasma pneumoniae and chlamydia pneumoniae nucleic acid based on double amplification (RNA constant temperature amplification + multi-biotin signal amplification) technology and its application. Background technique [0002] Mycoplasma pneumoniae (Mycoplasmal pneumonia, MP) is the main pathogen of community-acquired pneumonia (CAP), and its infection has been on the rise in recent years. Mycoplasma pneumoniae is a cell-wall-free microorganism that can pass through sterile filters but cannot be cultured in vitro in standard bacterial culture media, intermediate between viruses and bacteria. MP can cause upper and lower respiratory tract infections, and is one of the important pathogens causing atypical pneumonia. Due to its small size, MP is usually transmitted through droplets. The prevalence of MP varies with seasons, genders, ag...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/682C12Q1/6834C12Q1/689C12Q1/04
CPCC12Q1/682C12Q1/6834C12Q1/689C12Q2600/166C12Q2531/119C12Q2547/101C12Q2545/101C12Q2545/113C12Q2563/103C12Q2521/107Y02A50/30
Inventor 李先强姜昕厉洁黄永伟陈巨龚丽君
Owner 武汉中帜生物科技股份有限公司
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