REF 89-03 Form F
The SARS-CoV-2 Triplex PCR kit is one-step reverse transcription real-time PCR multiplex test for detection of SARS-CoV-2 nucleic acids from nasopharyngeal swabs, oropharyngeal swabs and saliva samples from individuals suspected of COVID-19.
SARS-CoV-2 Triplex PCR kit simultaneously detects three targets, in ORF1ab, ORF8, and N protein coding regions, each on separate detection channel. The detection of 3 regions increases the specificity of the test and makes it stable against potential mutations of the virus in the regions where primers or probes bind.
As internal control of quality of RNA extraction and reverse transcription reaction, the test uses specific set of primers and fluorescent probes for transcript of one of the human “housekeeping genes”.
Positive control reaction uses mix of in vitro transcribed RNAs corresponding to the target SARS-CoV-2 genome regions.
| Assay Principle
|| one-step reactions of reverse transcription and real-time PCR with fluorescent probe detection.
| Test results
|| nasopharyngeal swabs, oropharyngeal swabs and saliva samples in transport medium or NaCl solution
||10 copies per reaction
|| PCR-Mix SC2 in a 2 mL tube,
Taq DNA-polymerase with reverse transcriptase
| Controls included in the kit
|| • endogenous internal control
• positive control
• negative control
| Validated extraction methods
Magnetic DNA/RNA Extraction kit (Astra Biotech GmbH)
Prime DNA/RNA Extraction kit (Astra Biotech GmbH)
| Validated Thermocycler
Qiagen Rotor-Gene Q
Corbett Research Rotor-Gene 3000/6000
Bio-Rad CFX96/CFX96 Touch
Roche Light Cycler 96
Thermo Fisher Scientific/Applied Biosystems - QuantStudio-5
| Detection channels required for the PCR cycler:
|| ROX, HEX, Cy5, FAM
||-18...-30 °C, Storage at ≤+25 °C is allowed for no more than 5 days
||for in-vitro Diagnostics
Confirmed detection capability for the following virus variants:
VOC Alpha 202012/01 GRY (B.1.1.7), VOC Beta GH/501Y.V2 (B.1.351), VOC Gamma GR/501Y.V3 (P.1), VOC Delta G/478K.V1 (B.1.617.2), VOI Eta GR/484K.V3 (B.1.525), B.1.1.523, ŔŇ.1, VOC Alpha 202012/01 GRY (AY.4). VOI Epsilon GH/452R.V1 (B.1.429+B.1.427/CAL.20C), VOI Zeta GR/484K.V2 (P.2), VOI Theta GR/1092K.V1 (P.3), VOI Iota GH/253G.V1 (B.1.526), VOI Kappa G/452R.V3 (B.1.617.1), VOI Lambda GR/452Q.V1 (C.37), VOI Mu GH (B.1.621+B.1.621.1), VUM GH/490R (B.1.640). VOC Omicron GR/484A (B.1.1.529+BA.1+BA.2+BA.3+BA.4+BA.5).
Advanced test design
• three SARS-CoV-2 specific targets chosen from least variable regions in ORF1ab, ORF8, and N genes
• multiplicity of targets increases the robustness of the test in case of possible arising mutations in the regions where primers or probes bind
Control of the entire diagnostic process
• endogenous internal control (transcript of one of human «housekeeping genes») allows to monitor the quality of sampling, RNA extraction procedure, reverse transcription and PCR amplification, as well as detection reactions
• positive control (in vitro transcribed RNA of target regions) allows to monitor the quality of reverse transcription and PCR amplification and detection reactions
• negative control allows to monitor the cases of possible contamination
Single tube design
• simultaneous detection of three targets + internal control in one tube
• internal control is included in master mix - no additional pipetting required
Magnetic DNA/RNA Extraction kit
Prime DNA/RNA Extraction kit