In has been over a year since the outbreak of COVID-19 started in Wuhan City, China. Since then, it has spread across the world. While the development of vaccines plays an important role in containing the disease, monitoring the spread remains vital in this battle against COVID-19. So far, samples have been mostly collected through nasopharyngeal swabs. However, as this method is rather unpleasant, reliable workflows based on saliva are becoming more common.
Canadian manufacturer Norgen Biotek has developed a complete SARS-CoV-2 workflow which can also be combined with other brands’ products.
Want to check if Norgen’s products fit into your workflow?
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Step 1 Sample collection & preservation
Saliva RNA Collection and Preservation Device
CE-IVD marked kits are available. These are in accordance with EU Directive 98/79/EC.
- Non-invasive, user-friendly sample collection. Self-collection suitable (see training video)
- Inactivates viruses including SARS-CoV-2 upon mixing (App. Note 94)
- Preserved RNA is stable for up to 2 months at room temperature
- Compatible with many downstream applications, incl. PCR and qPCR
- Compatible with most commercial isolation methods
Read more on Norgen’s product page
Alternatively, Norgen also offers Total Nucleic Acid Preservation Tubes Dx (for swabs). These tubes are authorized for diagnostic use by Health Canada and are CE-IVD marked in accordance with EU Directive 98/79/EC.
Saliva samples were stored at ambient temperature in Norgen's Saliva RNA Preservative and monitored over a 2 month period. At regular intervals, RNA was isolated from 250 µL of preserved samples and RT-PCR was performed on three housekeeping genes. All of the genes were preserved and stable over a 2 month period.
When samples are taken by non-professionals, there will be additional variation. What happens when the amount of saliva collected varies? Norgen tested saliva samples collected in their Saliva RNA Preservative outside of the ideal range of 1:1 and stored them at room temperature for 3 days. One-step RT-PCR results indicated that the variation of the saliva input volumes did not significantly influence the RNA gene detection.
Step 2 Viral RNA isolation
Saliva/Swab RNA Purification Kits
This kit is very similar to Norgen’s topselling RNA purification kit, except that it is optimized for the extraction of RNA from saliva. CE-IVD marked kits, which are in accordance with EU Directive 98/79/EC, are also available.
- Quick and easy RNA isolation from saliva and swab samples
- Isolate total RNA, from large rRNA down to microRNA without phenol or chloroform
- Very sensitive & linear down to a few cells/viral copies without the need for carrier RNA
- Suitable for a variety of downstream applications, incl. Small RNA Sequencing
Why are Norgen’s silicon carbide columns so much better for RNA purification?
Read our blog article on it!
Isolation from swabs as well as saliva
Duplicate nasopharyngeal swabs, oropharyngeal swabs and saliva samples were collected from different donors into Norgen’s Total Nucleic Acid Preservative Tubes (69200) and Saliva RNA Collection and Preservation Devices (RU53800), respectively. Then the samples were spiked with three different concentrations from Norgen’s E/RdRP/RP Positive Control (included in Norgen’s Covid-19 RT-PCR kit TM67200) and processed for RNA isolation using Norgen’s Saliva/Swab RNA Purification Kit (69100). The linear recovery of the different spike-in amounts indicated high sensitivity of the isolation procedure as well as the high quality of the RNA purified.
Integration into a SARS-CoV-2 assay
Curative, Inc. developed an RT-PCR test for SARS-CoV-2 that makes use of Norgen’s Total RNA Purification 96-well Kit (24350/24380). They employ the use of this kit both manually or automated on the Tecan Resolvex A20. They have recently received Emergency Use Authorization from the FDA for in vitro diagnostic use of this kit at KorvaLabs LLC.
Performance against the competition
One of the most important factors affecting the sensitivity of any RT-qPCR viral assays is the RNA extraction method, particularly in asymptomatic individuals with extremely low viral loads. Read these application notes comparing Norgen’s Saliva/Swab RNA Purification Kit against leading competitors!
Step 3 PCR based detection of SARS-CoV-2
COVID-19 TaqMan RT-PCR Kit (E/RdRP genes) Dx
Norgen’s COVID-19 TaqMan RT-PCR Kit (E/RdRP genes) Dx is an in vitro diagnostic test for the qualitative detection of SARS-CoV-2 specific RNA using a multiplexed TaqMan® fluorescence detection assay (FAM and HEX/VIC) based on the Charité/Berlin protocol.
- Convenient ready-to-use 2X One-Step RT-PCR Master Mix
- Primer & Probe Mixes contain all E gene/RP and RdRP gene assays in individual tubes
- Positive Control contains the three SARS-related target genes: E gene, RdRP gene as well as the human RP (internal control)
- All assays are premixed to the working concentrations recommended by the Charité/Berlin protocol.
- Manufactured under ISO 13485:2016 (RUO)
Sensitive
The analytical sensitivity was determined by analyzing six serial dilutions (200 000 copies to 2 copies) of Norgen's positive control from the COVID-19 TaqMan RT-PCR Kit (E/RdRP gene)(Cat. TM67200). The arrow indicates a copy number of 10.
Specific
The E/RdRP/RP Positive Control was used to test the kits specificity against RNA purified from other related pathogens. RdRP reactions were only prepared for samples required for confirmation and discriminatory detection based on the first gene detection result. Amplification was measured by real time RT-PCR.
Detects SARS-CoV-2 variants
Variant analysis showed that the E gene assays has alignment scores of 100% for the multiple SARS-CoV-2 variants (24.03.2021).
Norgen Covid-19 workflow products
| Cat-No. | Item | Size | Price (CHF) |
|---|---|---|---|
| 53800 | Saliva RNA Collection and Preservation Devices Dx | 50 units | 787.00 |
| RU53800 | Saliva RNA Collection and Preservation Devices | 50 units | 750.00 |
| Dx69200 | Total Nucleic Acid Preservation Tubes Dx | 50 tubes | 448.00 |
| 69200 | Total Nucleic Acid Preservation Tubes | 50 tubes | 431.00 |
| Dx69100 | Saliva/Swab RNA Purification Kit Dx | 50 preparations | 411.00 |
| 69100 | Saliva/Swab RNA Purification Kit | 50 preparations | 387.00 |
| Dx69300 | Saliva/Swab RNA Purification 96-Well Kit Dx | 2 plates | 1'096.00 |
| 69300 | Saliva/Swab RNA Purification 96-Well Kit | 2 plates | 1'065.00 |
| Dx17200 | Total RNA Purification Kit Dx | 50 preparations | 375.00 |
| DxTM67200 | COVID-19 TaqMan RT-PCR Kit (E/RdRP genes) Dx | 500 reactions | 3'775.00 |
| TM67200 | COVID-19 TaqMan RT-PCR Kit (E/RdRP genes) | 500 reactions | 3'533.00 |
| DXTM67100 | 2019-nCoV TaqMan RT-PCR Kit Dx | 50 reactions | 424.00 |