Respiratory panel real time PCR detection (500 tests)

Respiratory panel real time PCR detection (500 tests)

Cat. No.: BMG-QP2179-02
Size: 500 tests


Description

A broad range of respiratory viruses cause acute local and systemic infections that may be severe in children, the elderly and immunocompromised patients. The detection and identification of specific viral type in patients who have symptoms of respiratory tract infection will aid physicians in the timely diagnosis and administering the appropriate treatment for these patients, as well as prevent spreading of the viruses to at risk individuals. Upper and lower respiratory tract infections are caused by a broad range of microbes, including RNA and DNA viruses, bacteria, and even fungi, and yet are often symptomatically similar. This panel is designed for specific and qualitative detection of respiratory viruses in specimens such as oropharyngeal swabs, nasopharyngeal swabs or sputum. The product is intended for use by qualified laboratory personnel well trained in nucleic acid amplification techniques and in vitro diagnostic procedures. Positive results do not rule out bacterial infection or co-infection with other pathogens. Negative results do not preclude microbial infection and should not be used as the sole basis for patient management decisions. Test results must be combined with clinical observations, patient history, and epidemiological information.

Summary and explanation of the test

The respiratory Panel Real Time PCR Detection Kit is designed for simultaneous detection of SARS-CoV-2, Influenza A, Influenza B, Respiratory Syncytial Virus, Adenovirus, Human Metapneumovirus, Rhinovirus, ParaInfluenza 1, ParaInfluenza 2, ParaInfluenza 3 and ParaInfluenza 4. RNA/DNA is extracted from respiratory specimens, amplified using RT-PCR and detected using fluorescent reporter dye probes.

Principle

The respiratory viral panel assay is a real-time reverse transcription polymerase chain reaction (rRT-PCR) test. The multiples assay contains 11 primer/probe sets that target the RNA/DNA of influenza A, influenza B, SARS-CoV-2, Respiratory Syncytial Virus, Adenovirus, Human Metapneumovirus, Rhinovirus, ParaInfluenza1, ParaInfluenza2, ParaInfluenza3 and ParaInfluenza4. The assay also contains a primer/probe set to detect the human RNase P gene (RP) in clinical specimens or control samples. The kit also contains a positive control (Plasmid DNA) and a negative control for testing process monitoring.


Reagents and materials provided
 

Item	Component	Quantity		Volume /T
		100 Tests / Box	500 Tests / Box
1	PCR Mix I PCR Mix II PCR Mix III	1.5 mL/ tube × 1	1.5 mL/ tube × 5	14 µL
2	RT-PCR Enzyme Mix	330 μL/ tube × 1	1550 μL/ tube × 1	1 µL
3	Positive Control 1	50 μL/ tube × 1	250 μL/ tube × 1	5 μL
4	Positive Control 2&3	50 μL/ tube × 1	250 μL/ tube × 1	5 μL
5	Negative Control	50 μL/ tube × 1	250 μL/ tube × 1	5 μL

Storage instructions

Store the kit frozen at -20±5°C and avoid repeated freeze-thaw cycles (less than 5 frozen-thaw times allowed). The product is valid for 12 months, and the expiry date of the product is shown on the label.

Specimen type and preservation

Specimen type: oropharyngeal swabs, nasopharyngeal swabs and sputum.
Specimen preservation: process specimens for viral RNA extraction and nucleic acid detection as soon as possible. Specimens can be stored at 4°C for 24 hours, or frozen at -80°C for longer storage, if not processed immediately. Avoid repeated freeze-thaw cycles during transport and storage of the specimens

Assay PCR reaction setup (at reagent preparation area)

A. Reagent Preparation
To calculate the number (n) of PCR reaction tubes (n= number of specimens to be tested + number of positive controls + number of negative controls), add n x14 μL of SARS-CoV-2 PCR Mix and n x 1 μL of RT-PCR Enzyme Mix to a clean tube, vortex to mix well, centrifuge and aliquot 15 µL of the mix into each PCR reaction tube.

B. Addition of Viral RNA
Add 5 μL of viral RNA extracts of the specimen or positive/negative control to each PCR tube according to the reaction preparation table below for testing. Then immediately cap the tube tightly, briefly spin the mixture and move it to the amplification area for RT-PCR detection

• PCR Reaction Preparation

C. RT-PCR amplification (at Amplification Area, use Applied Biosystems 7500 Real-Time PCR System as an example)

1. 1. Place the complete PCR reaction tubes into the fluorescent quantitative PCR analyzer and label positive control, negative control and testing specimen on the Thermocycler software.
2. 2. Enter the PCR reaction volume: 20 μL.
3. 3. Select fluorescence detection channels: JOE, FAM and Cy5 channels. Cy5 is the E gene indicator channel, FAM is the ORF1ab indicator channel, and JOE is the RNase P gene indicator channel (internal control).
4. 4. Select PCR cycle parameter setting following the instructions below, save the file and start the PCR run.
5. 5. Analyze the results according to your thermocycler manufacturer’s instructions. If the positive and negative control samples do not meet their respective requirements, the PCR run must be repeated.

Determination of results

1. The results can only be determined when the results of both controls meet their requirements.
2. Criterion for a positive specimen: a specimen can be reported as positive when FAM, Cy5 and JOE channels are all positive.
3. Criterion for a negative specimen: a specimen can be reported as negative when the JOE channel is positive and the ROX, FAM and Cy5 channels are negative.

Compatibility of real-time PCR thermocyclers
Product performance

Product analysis performance evaluation results:

1. 1. The analytical sensitivity of this kit is 200 copies/mL.
2. 2. Cross-reaction: no cross-reaction with other pathogens such as, enterovirus types A, B, C and D, EB virus, measles virus, human cytomegalovirus, rotavirus, norovirus, mumps virus, varicella zoster virus, mycoplasma pneumoniae, chlamydia pneumoniae, legionella, bordetella pertussis, haemophilus influenzae, staphylococcus aureus, streptococcus pneumoniae, streptococcus pyogenes, klebsiella pneumoniae, mycobacterium tuberculosis, aspergillus fumigatus, candida albicans, candida glabrata, cryptococcus neoformans, coronavirus (HKU1, OC43, NL63, 229E), SARS-CoV-1, and MERS coronavirus.

REFERENCE

1. National Health Commission & State Administration of Traditional Chinese Medicine. Diagnosis and Treatment Protocol for Novel Coronavirus Pneumonia (Trial Version 7). [R]. March 3, 2020