Instruction Manual
Table Of Contents
- Intended Use
- Summary and Explanation
- Principles of the Procedure
- Materials Required (Provided)
- DTT solution 500mM
- SARS-CoV-2 PCR Mix
- Equipment and Supplies
- Warnings and Precautions
- Specimen Collection, Handling, and Storage
- Procedure
- For viscous samples (sputum and bronchial wash): Performed inside BSC 2
- NOTE: In the event that the test system becomes inoperable, notify supervision or designee for further direction. Patient specimens must be stored in a manner that maintains the integrity of the specimen.
- Quality Control
- Limitations
- Conditions of Authorization for the Laboratory
- Performance Characteristics
- References
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The Quest Diagnostics SARS-CoV-2 RNA, Qualitative Real-Time RT-PCR aids in the detection of SARS-CoV-2 RNA and
diagnosis COVID-19 and is a real-time reverse transcription polymerase chain reaction test. The test’s primer and probe
sets were designed to detect RNA from individuals suspected of COVID-19 by their healthcare provider. Testing is limited
to
Quest Diagnostics laboratories in San Juan Capistrano CA, Chantilly VA, and Marlboro MA, or other laboratories
designated by Quest Laboratories that are also certified under the Clinical Laboratory Improvement Amendments of 1988
(CLIA), 42 U.S.C. §263a, to perform high complexity tests.
In sample pooling, specimens are identified from populations based on positivity rate (for example, by county, zip code or
by client), and up to four patient specimens are combined into a pool, and the pool is tested as described in this package
insert. If the pool is positive or inconclusive or invalid, then each of the constituent samples is re-tested as a separate
individual specimen. If the pool is negative, then each constituent sample is reported as negative.
Principles of the Procedure
The test is a real-time RT-PCR test intended for the qualitative detection of nucleic acid from the SARS-CoV-2 in upper
respiratory specimens (for example, nasopharyngeal swabs, oropharyngeal swabs, sputum, BAL, and tracheal aspirates).
The assay is composed of two principal steps: (1) extraction of RNA from patient specimens, (2) one-step reverse
transcription and PCR amplification with SARS-CoV-2 specific primers and real-time detection with 2019-nCoV specific
probes. The assay targets regions of the virus nucleocapsid gene (N1 & N3) and is designed for the detection of SARS-
CoV-2. Amplification and detection are accomplished using TaqMan chemistry on the ABI 7500. To ensure the absence of
non-specific PCR inhibition of a sample, an internal positive amplification control (IPC) is included with each specimen. A
sample can be interpreted as negative only if the analysis of the IPC indicates that amplification has occurred in the reaction
tube but no signal from target reporter dye has been detected. Detection of viral RNA not only aids in the diagnosis of illness
but also provides epidemiological and surveillance information.
Materials Required (Provided)
• MagnaPure Extraction
• MagNA Pure 96 DNA and Viral NA – Small Volume Kit Roche Diagnostics #06 543 588 001 (3 x 192
isolations)
• MagNA Pure 96 External Lysis Buffer or other comparable lysis buffer that will be validated
• Omega Extraction
• Mag-Bind Viral RNA Xpress Kit (Omega Bio-Tek, Cat. M6219-2304)
• 4X 1-Step RT-qPCR Master Mix, CG
• Exogenous NA Primer Pair
•
Exogenous NA
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• TE Buffer, pH 8.0
• Quest V-C-M transport medium, Quest PBS Specimen Transport Tubes or other comparable transport medium
that will be validated
• Poly (A)
• DEPC-water
• PBS, 1X
• DTT
Reagents:
nCoV RT-PCR Mix Primers and Probes
2019-nCoV_N1 Forward Primer
2019-nCoV_N1 Reverse Primer