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Freeze-dried Mycoplasma Pneumoniae Nucleic Acid Detection Kit(Enzymatic Probe Isothermal Amplification)

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Freeze-dried Mycoplasma Pneumoniae Nucleic Acid Detection Kit(Enzymatic Probe Isothermal Amplification)

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Introduction

[Product Name]Freeze-dried Mycoplasma Pneumoniae Nucleic Acid Detection Kit (Enzymatic Probe Isothermal Amplification)
[Packaging Size] 48 tests/kit
[Intended Use]

This kit is intended for in vitro qualitative detection of Mycoplasma pneumoniae (MP) nucleic acid in human throat swabs.
Mycoplasma pneumoniae (MP) is the smallest prokaryotic microorganism with a cell structure and no cell wall between bacteria and viruses[1]. MP mainly causes respiratory tract infections in humans, especially in children and young people[2]. MP can cause Mycoplasma hominis pneumonia, respiratory tract infections in children and atypical pneumonia. The clinical symptoms are diverse, mostly severe cough, fever, chills, headache, sore throat, upper respiratory tract infection and bronchopneumonia are the most common. Some patients may develop severe pneumonia from upper respiratory tract infection, and severe respiratory distress or even death may occur[3,4]. MP is one of the common and important pathogens in community-acquired pneumonia (CAP)[5], accounting for 10%-30% of CAP, and the proportion can increase 3-5 times when MP is prevalent[6]. In recent years, the proportion of MP in CAP pathogens has gradually increased. The incidence of Mycoplasma pneumoniae infection has increased, and because of its nonspecific clinical manifestations, it is easy to be confused with bacterial and viral colds. Therefore, early laboratory detection is of great significance for clinical diagnosis and treatment[7].

This kit is used to detect MP nucleic acid in human throat swab samples. The results are for clinical reference only, and can not be used as the only basis of the clinical diagnosis. The clinical diagnosis must be comprehensively analyzed in combination with the patient' s clinical manifestations and other laboratory tests.

[Test Principle]

The kit was to detect nucleic acid of MP using Enzymatic Probe Isothermal Amplification (EPIA). Primers and Probe with RNA bases (rProbe) were developed tailored to the P1 gene of MP and added Bst enzyme and RNaseH. Two ends of RNA bases in rProbe were labeled with fluorophores as reporter and quencher. During amplification, the target sequence was amplified in isothermal condition using DNA polymerase and strand replacement of Bst enzyme, RNaseH cleaves RNA bases on 

target gene-probe hybrid strands, enabling to separate the reporter dye from the quencher dye and to generate the fluorescent signal. The remaining segment of rProbe RNA can continue the amplification process to generate fluorescent signals further. The fluorescent signals can be monitored by detection system, thereby realizing the detection of the target nucleic acid.

Recombinant pseudo-virus of Pine chloroplast gene was developed as internal control in the kit to monitor the quality in laboratory processing including validity of reagent and laboratory operation to avoid false negative.

[Storage Condition and Shelf-life]
1. Unopened kit: It should be stored at no higher than 30°C and protected from light, and the shelf-life is 12 months.
2.Test should be performed immediately after reconstitution of the freeze-dried reaction buffer, internal control and positive control, the remaining internal control and positive control should be stored below -15°C and protected from light. The validity period is 1 week, and repeated freezing and thawing should be avoided ( limited to 2 cycles).

3.Transportation conditions: It can be transported stably for 1 month when the temperature is not higher than 37°C.
4. See the packaging label for the production date, production batch number and expiration date.

[Applicable Instruments]

Applied Biosystems 7500 Real-Time PCR Systems, SLAN ®-96P Real-Time PCR Systems, LightCycler® 480 Real-Time PCR system, Easy Amp Real-time Fluorescence Isothermal Detection System(HWTS1600)
[Requirements for Samples]

1. Sample collection
Throat swabs collection: wipe the posterior pharyngeal wall and bilateral tonsils at the back of the uvula (palatine or uvula) with moderate force with a polyester fiber head swab, avoid touching the tongue, take out the posterior sampling tube, and break the plastic handle at the contact part of the hand, soak the swab into the sampling buffer, and tighten the cap. Throat swab samples can be preserved with a preservative solution such as normal saline or Hank's solution.
2. Sample storage
Samples can be stored for no more than 7 days at 2-8°C and can be stored for no more than 4 months below -18°C. If the sample needs to be stored for a long time, it should be stored below -70°C. Repeated freezing and thawing of samples should be avoided.

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