Expected use

The Gnomegen COVID-19 RT-qPCR Detection Kit is a real-time in vitro RT-PCR diagnostic test for the qualitative detection of SARS-CoV2 virus nucleic acid in nasopharyngeal and oropharyngeal swab samples collected from people suspected of COVID-19 by their healthcare provider. Tests are limited to laboratories certified under the Clinical Laboratory Improvement Amendments of 1988 (CLIA), 42 U.S.C. §263a, to perform highly complex tests.

The results are for the identification of SARS-CoV-2 RNA. SARS-CoV-2 RNA is generally detectable in nasopharyngeal and oropharyngeal swab samples during the acute phase of infection. Positive results are indicative of the presence of SARS-CoV-2 RNA. Clinical correlation with the patient’s history and other diagnostic information is necessary to determine the patient’s infection status. Positive results do not rule out bacterial infection or
coinfection with other viruses. The detected agent may not be the definitive cause of disease. Laboratories within the United States and its territories are required to report positive results to the corresponding public health authorities.

Negative results do not rule out SARS-CoV-2 infection and should not be used as a sole basis for patient management decisions. Negative results should be combined with clinical observations, patient history, and epidemiological information. The Gnomegen COVID-19 RT-qPCR Detection Kit is designed for you to use
specifically trained clinical laboratory personnel instructed and trained in the techniques of in vitro diagnostic procedures and real-time PCR. The Gnomegen COVID-19 RTqPCR Detection Kit should only be used under the
Emergency use authorization.

Summary and explanation

An outbreak of pneumonia caused by a new coronavirus (SARS-CoV-2) in Wuhan City, Hubei Province, China was identified and reported to WHO on December 31, 2019. The rapid spread of SARS-CoV-2 to numerous areas around the world requires preparedness and response in laboratory and healthcare facilities. Disponibility Specific and sensitive assays for the detection of the virus are essential for a case diagnosis, evaluation of the extent of the outbreak, follow-up of the intervention surveillance strategies and studies.

The Gnomegen COVID-19 RT-qPCR Detection Kit is an in vitro molecular diagnostic test which aids in the detection and diagnosis of SARS-CoV-2 and is based on nucleic acid amplification technology. The product contains oligonucleotide primers and double-labelled hydrolysis probes (TaqMan®) and control material used in RRT-PCR to Qualitative in vitro detection of SARS-CoV-2 RNA in respiratory samples.

Principles of the procedure

Oligonucleotide primers and probes were selected for the detection of SARS-CoV-2. CV03030005 R01 Gnomegen 4
of regions of the virus nucleocapsid (N) gene. The panel is designed for 2019-nCoV detection (two sets of primers/probes). An additional primer/probe configured to detect the human RNase P (RP) gene in control samples and clinical samples as well included in the panel.

RNA isolated and purified from upper and lower respiratory tract samples are reverse transcribed to cDNA and subsequently amplified on the Applied Biosystems 7500 Fast Dx real-time PCR instrument with SDS version 1.4 software. In the process, the probe hybridizes to a specific target sequence located between the forward and reverse primers.

During the extension phase of the PCR cycle, the 5 ‘nuclease activity of Taq. The polymerase degrades the probe, causing the indicator dye to separate from the extinguishing dye, which generates a fluorescent signal. With each cycle, additional reporter dye molecules are cleaved from their respective probes, increasing fluorescence.

Equipment and consumables required (but not supplied)

• Vortex mixer
• Microcentrifuge
• Micropipettes (2 or 10 μL, 200 μL and 1000 μL)
• Multichannel micropipettes (5-50 μL)
• Racks for 1.5 mL microcentrifuge tubes
• 2 cold blocks of 96 wells at -20 ° C
• Molecular grade water, nuclease free
• 10% bleach (1:10 dilution of commercial 5.25-6.0% hypochlorite bleach)
• DNAZapTM (Ambion, cat. # AM9890) or equivalent
• RNAse AwayTM (Fisher Scientific; cat. # 21-236-21) or equivalent
• Powder-free disposable gloves and surgical gowns
• Aerosol barrier pipette tips
• 1.5 mL microcentrifuge tubes (DNase / RNase free)
• 0.2 ml PCR Reaction Plates (Applied Biosystems; Catalog # 4346906 or # 4366932)
• MicroAmp 8-Plug Optical Strips (Applied Biosystems; Catalog # 4323032)

Reagent storage, handling and stability

• Store the assay and controls at -20 ° C.
• Store liquid HSC control materials at ≤ -20 ° C. CV03030005 R01 Gnomegen 8
• Protect fluorogenic probes from light.
• Primers, probes (including aliquots), and enzyme master mix should be thawed and kept on a cold block at all times during preparation and use.
• Controls and aliquots of controls must be thawed and kept on ice at all times during preparation and use.