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Kit for detecting mycoplasma pneumonia

A technology of Mycoplasma pneumoniae and a kit, which is applied to measurement devices, instruments, disease diagnosis and other directions, can solve the problems of prone to cross-reaction, high requirements on culture conditions, and complicated operations, and achieves the advantages of spectrophotometric detection, shortened reaction time, The effect of increasing sensitivity

Inactive Publication Date: 2019-02-15
廖朝晖
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The isolation and culture method is the gold standard for diagnosing MP infection, but it requires high culture conditions and a long culture period (10-14 days). It is generally used for scientific research and clinical retrospective analysis, and has little significance for the early diagnosis of MP. It is rarely used now. Clinical diagnosis; molecular biology including: nested PCR, real-time quantitative PCR, loop-mediated isothermal amplification reaction (LAMP) technology, nucleic acid sequence-dependent amplification technology (NASBA), real-time fluorescent nucleic acid constant temperature amplification detection technology (SAT) Composite PCR and other technologies, molecular biology techniques have high sensitivity and specificity, and are suitable for early clinical diagnosis. However, in addition to the shortcomings of molecular biology itself, such as high false positive rate and easy contamination of specimens, studies have found that MP in healthy people The carrier rate is high (0.1%-13.5%), and the DNA can survive in the respiratory tract for a long time after infection (7 weeks to 7 months). Combined with clinical judgment
In addition, it is also affected by factors such as different specimen types, sampling sites, sampling personnel, and experimental methods used in the laboratory. At present, there is no unified standard, which brings certain difficulties to the evaluation of laboratory diagnostic methods; serological methods are commonly used in the diagnosis of MP infection in my country. , the experimental methods include complement fixation test (CFT), gelatin particle agglutination test (PA) and ELISA and other methods
The overall sensitivity of CFT is poor, the specificity of detecting antibodies is not high, and it is prone to cross-reaction, resulting in false positives, so it is rarely used clinically
The sensitivity and specificity of PA test and ELISA are significantly better than CFT, and are widely used clinically in some countries, and have good consistency with molecular biology results. However, commercial serological methods for detecting MP infection are all manual method, the operation is cumbersome and time-consuming, and its sensitivity is also affected by laboratory conditions and the skills of experimenters.

Method used

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  • Kit for detecting mycoplasma pneumonia

Examples

Experimental program
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Effect test

Embodiment 1

[0043] Preparation and detection of Mycoplasma pneumoniae P1 latex-enhanced immunoturbidimetric assay kit

[0044] (1) Preparation of reagent R1:

[0045] Weigh 23.83g of Hepes, 9g of NaCl, 15g of PEG8000, 0.5g of sodium azide, and 5g of bovine serum albumin, adjust the pH to 7.4, and dissolve in 1L to obtain reagent R1.

[0046] (2) Preparation of reagent R2:

[0047] The first step: with the MES buffer solution of 0.02M / L, the Mycoplasma pneumoniae cell membrane component (Mycoplasma pneumoniae is donated by the Chinese Academy of Medical Sciences, and the cell membrane component is obtained by processing) is diluted to a concentration of 2 mg / ml of the Mycoplasma pneumoniae cell membrane component dilution; Styrene latex microspheres (136nm) were washed 3 times with distilled water, and the supernatant was removed.

[0048] Step 2: Dilute the washed polystyrene latex microspheres to a mass concentration of 1% with MES buffer solution having a pH of 5.3 and a concentration...

Embodiment 2

[0053] Preparation and detection of Mycoplasma pneumoniae P116 latex-enhanced immunoturbidimetric assay kit

[0054] (1) Preparation of reagent R1:

[0055] Weigh 23.83g of Hepes, 9g of NaCl, 15g of PEG8000, 0.5g of sodium azide, and 5g of bovine serum albumin, adjust the pH to 7.4, and dissolve in 1L to obtain reagent R1.

[0056] (2) Preparation of reagent R2:

[0057] Step 1: Dilute the human mycoplasma pneumoniae antigen P116 with 0.02M / L MES buffer solution to a concentration of 2 mg / ml of mycoplasma pneumoniae antigen P116 dilution; the polystyrene latex microspheres are washed with distilled water.

[0058] Step 2: Dilute the washed polystyrene latex microspheres to a mass concentration of 1% with MES buffer solution having a pH of 6.0 and a concentration of 0.02M / L. Add 0.01g of EDC to 1L of the above-mentioned latex dilution, stir and react at room temperature for 30min, centrifuge at a speed of 15000rpm for 15min after the reaction to remove the supernatant, resusp...

Embodiment 3

[0063] Preparation and detection of Mycoplasma pneumoniae P30 latex-enhanced immunoturbidimetric assay kit

[0064] (1) Preparation of reagent R1:

[0065] Weigh 23.83g of Hepes, 9g of NaCl, 15g of PEG8000, 0.5g of sodium azide, and 5g of bovine serum albumin, adjust the pH to 7.4, and dissolve in 1L to obtain reagent R1.

[0066] (2) Preparation of reagent R2:

[0067] Step 1: Dilute the human mycoplasma pneumoniae antigen P30 with 0.02M / L MES buffer to a concentration of 2mg / ml mycoplasma pneumoniae antigen P30 dilution; polystyrene latex microspheres (136nm) are washed with distilled water.

[0068] Step 2: Dilute the washed polystyrene latex microspheres to a mass concentration of 1% with MES buffer solution having a pH of 5.0 and a concentration of 0.02M / L. Add 0.01g of EDC to 1L of the above-mentioned latex dilution, stir and react at room temperature for 30min, centrifuge at a speed of 15000rpm for 15min after the reaction to remove the supernatant, resuspend with 0.0...

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Abstract

The invention relates to a kit for detecting mycoplasma pneumonia. The kit comprises an R1 reagent and an R2 reagent. The R1 reagent comprises a first buffer, a first stabilizer, a first protectant, afirst preservative, an opacifier and water. The R2 reagent comprises mycoplasma pneumonia antigens, polystyrene latex microspheres, a second buffer, a second stabilizer, a second protectant, a secondpreservative and water. The kit of the invention has the advantages of high sensitivity, high detection speed, stable result, wide linear range, good repetition, easy to standardize and specimen collection, and small in specimen usage amount and the like. Also, the kit of the invention has the advantages that it provides a novel method for early, rapid, high sensitivity and specificity for the diagnosis of clinical mycoplasma pneumonia infection.

Description

technical field [0001] The invention relates to the field of in vitro diagnostic reagents, in particular to a kit for detecting mycoplasma pneumoniae. Background technique [0002] Mycoplasma pneumoniae (Mycoplasma pneumoniae, MP) is one of the important pathogens of children's community-acquired pneumonia. MP infection accounts for 10%-30% of children's community-acquired pneumonia infection, and it mostly occurs in families, communities, schools and camping areas. crowded public places. MP infection is seasonal, mostly occurs in autumn, and is mainly transmitted by droplets. Most MP infections are limited. Whether clinical infection symptoms appear depends on whether MP adhered to the surface of the host’s respiratory tract can be cleared, but there are still 3%- 13% cause clinical symptoms, the main symptom is dry cough, body temperature at 37.7±1.0°C, and last for 1-3 weeks, may be accompanied by sore throat and muscle pain, lack of specific clinical features, and appro...

Claims

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Application Information

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IPC IPC(8): G01N33/569
CPCG01N33/56933G01N2800/12
Inventor 廖朝晖
Owner 廖朝晖
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