SARS-CoV-2 Neo Assay Kit

Primers and probes for real-time RT-PCR detection of SARS-CoV-2 and its future variants for epidemiological research

Products

The SARS-CoV-2 Neo Assay Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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SARS-CoV-2 Neo Assay Kit (600)

Cat. No. / ID: 222115

Each tube contains a ready-to-use mixture of twelve primers and six probes. Probes coupled with different dyes for detection in FAM, ROX and Cy5 channels.
$650.00
SARS-CoV-2 Neo Assay Kit should be used in conjunction with QIAprep&amp Viral RNA UM Kit
The SARS-CoV-2 Neo Assay Kit (600) is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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SARS-CoV-2 Neo Assay Kit (2400)

Cat. No. / ID: 222117

Each tube contains a ready-to-use mixture of twelve primers and six probes. Probes coupled with different dyes for detection in FAM, ROX and Cy5 channels.
$2,200.00
SARS-CoV-2 Neo Assay Kit should be used in conjunction with QIAprep&amp Viral RNA UM Kit
The SARS-CoV-2 Neo Assay Kit (2400) is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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SARS-CoV-2 Neo Positive Control

Cat. No. / ID: 222710

Each tube contains a synthetic double-stranded DNA fragment covering six targets within the SARS-CoV-2 genome.
$101.00
SARS-CoV-2 Neo Positive Control should be used in conjunction with SARS-CoV-2 Neo Assay Kit and QIAprep&amp Viral RNA UM Kit
The SARS-CoV-2 Neo Positive Control is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

Features

  • Targets ORF1a, ORF1b, ORF3a, ORF7a, N1 and N2 within the SARS-CoV-2 genome
  • Compatible with nasopharyngeal and oropharyngeal swabs, lolly (lollipop swab), gargle and saliva
  • Detects six different targets in three color channels (FAM, ROX and Cy5)
  • Does not detect SARS-CoV-1 or other Sarbecovirus species
  • Ready-to-use assay mix (all-in-one tube)

Product Details

To ensure future-proof detection of SARS-CoV-2, we developed a novel assay kit covering six targets across four genomic regions (ORF1ab, ORF3a, ORF7a and N gene) of the virus. Using multiple targets in multiple regions increases the likelihood of successful detection.


Probes are coupled to different dyes and can be measured in the FAM, ROX and Cy5 channels on a cycler of your choice.


This kit can be used in combination with the QIAprep&amp Viral RNA UM Kit chemistry and SARS-CoV-2 Neo Positive Control to provide fast and easy detection. Sample types compatible for detecting SARS-CoV-2 include nasal, nasopharyngeal, oropharyngeal swabs, lolly (lollipop swab), saliva, and gargles.

Performance

The unique design of the SARS-CoV-2 Neo Assay enables reliable and highly specific SARS-CoV-2 detection. In silico analyses showed that all investigated SARS-CoV-2 viral sequences (>4.8 Mio) were successfully detected by this assay. The assay does not detect SARS-CoV-1, other Sarbecovirus species, and/or other organisms as demonstrated by wet-lab and in-silico analyses. The list of wet-lab-tested respiratory pathogens includes Influenza A (H3N2, H1N1), Influenza B, Parainfluenza (1-4), Coronavirus 229E, OC43, NL63, C. pneumoniae, Metapneumovirus, B. pertussis, Human Rhinovirus 1A, RSV A, Human Adenovirus 5, L. pneumophila and Bocavirus. RNA from different respiratory pathogen-positive controls (chemically inactivated viruses) was tested for cross-reactivity with the SARS-CoV-2 Neo Assay using the QIAprep&amp Viral RNA UM Kit. None of the pathogens examined showed cross-reactivity (data not shown).

 

The SARS-CoV-2 Neo Assay Kit can be used in combination with the QIAprep&amp Viral RNA UM Kit chemistry and is compatible with cyclers having FAM, ROX and Cy5 detection channels and samples collected in all non-fixation transport media. Pooling of up to 20 samples per reaction is possible.

 

The analytical sensitivity per channel was assessed using in-vitro transcribed nucleic acids and determined as follows: 1000 copies/ml in FAM, 1000 copies/ml in ROX and 1500 copies/ml in Cy5.

 

Principle

The SARS-CoV-2 Neo Assay is intended for use with the QIAprep&amp Viral RNA UM Kit, with the above-mentioned sample types, transport media and cyclers.

 

The SARS-CoV-2 Neo Assay consists of twelve primers and six probes. Primers and probes are pre-mixed in a ready-to-use 20x formulation in a single tube. The oligonucleotides amplify the following six targets in the viral RNA: ORF1a, ORF1b, ORF3a, ORF7a, N1 and N2. The unique target combination results in high mutational tolerance while ensuring highly specific SARS-CoV-2 detection.

The probes are coupled to different dyes which can be detected in the FAM, ROX and Cy5 channels.

Optionally, the SARS-CoV-2 Neo Positive Control containing a synthetic DNA fragment covering all six targets can be used in the experimental setup.

 

Procedure

The SARS-CoV-2 Neo Assay is optimized for use with the QIAprep&amp Viral RNA UM Kit. Follow a few simple steps as outlined in the Quick-Start Protocol and get your results in under one hour. The procedure does not require a separate RNA extraction.

The SARS-CoV-2 Neo Assay uses the human sampling control of the QIAprep&amp Viral RNA UM Kit to monitor successful sampling and PCR amplification of the sample of your choice. You can use any qPCR instrument capable of detecting FAM, ROX, Cy5 and HEX (for the human sampling control).

The simple workflow is automatable on your platform of choice. If using a pool of 10 samples per reaction on a high-capacity robotic platform, a throughput of 250,000 samples per week can be achieved when working 24/7. 

Applications

The SARS-CoV-2 Neo Assay Kit is intended for molecular biology applications such as epidemiological research using real-time RT-PCR.

Supporting data and figures

Resources

Application/Protocol Documents (1)
Kit Handbooks (1)

FAQ

Can the SARS-CoV-2 Neo Assay Kit detect all variants?
Yes. We have assessed 4.8 mio sequences of a 6-month period within Gisaid -- none of the sequences showed a drop-out (at least 1 had to be positive). Not a single sequence showed a potential drop-out in more than 1 color channel. Additionally, we have wet-lab checked real samples for Alpha, Delta, Omicron, and artificial positive control samples for Alpha, Kappa, Beta, and Gamma.
FAQ ID - 3891
Can I use ROX as passive reference dye with the SARS-CoV-2 Neo assay?
No. Two of the viral targets are detected in the ROX channel. Passive reference dye is not necessary to evaluate results with the SARS-CoV-2 Neo Assay Kit.
FAQ ID - 3894
What color channels are used?
SARS-CoV-2 Neo Assay detects SARS-CoV-2 in FAM (N1+N2), ROX (ORF1a+ORF1b), and Cy5-spectrum (ORF3a+ORF7a). When combined with QIAprep&amp Viral RNA UM Kit, the human sampling control is detected in HEX.
FAQ ID - 3885
Is it possible to scale up sample volume and reaction volume?
Although this is theoretically possible, the protocol is optimized for 20 µl reaction volume and all steps, volumes and reagents concentrations have been optimized to deliver optimal results in these conditions.
FAQ ID - 3884
Anything important when starting the cycler run?
Yes. Please make sure that the data collection is switched on for FAM, HEX/JOE/VIC, ROX/TexasRed, Cy5/Quasar to collect viral amplification signals as well as the human sampling control amplification signal.
FAQ ID - 3888
Is it possible to use ABI instruments without ROX as a passive reference?
Yes. Select None as a passive reference in the ABI instruments, allowing to use the ROX as a reporter dye.
FAQ ID - 3889
What is the shelf-life of the Neo Assay Kit?
Up to 36 months
FAQ ID - 3890
Can I use the IC RNA of the QIAprep& Viral RNA UM Kit?
No. The IC RNA Assay is detected in Cy5. The SARS-CoV-2 Neo Assay detects two viral targets in the Cy5 channel. When using the IC RNA with the SARS-CoV-2 Neo Assay, the signals cannot be discriminated.
FAQ ID - 3895
Which SARS-CoV-2 targets can the SARS-CoV-2 Neo Kit detect?
The kit detects N1 and N2 (CDC assays) as well as ORF1a, ORF1b, ORF3a, and ORF7a proprietary designs.
FAQ ID - 3881
Do we have specificity and sensitivity data for this kit?
Isolated RNA of the following viruses:
  • Influenza: Inf A H3N2, Inf A H1N1, Inf B
  • Parainfluenza: PIV1, PIV2, PIV3, PIV4
FAQ ID - 3883
What is the minimum and maximum concentration of effective RNA for the test?
We have successfully tested positive samples with the lowest available Ct around 16 and very late Ct close before Ct 40, indicating a very high dynamic range that is compatible with the real life samples.
FAQ ID - 3887
Which positive control should I use?
You can purchase the SARS-CoV-2 Neo Positive Control. It contains all the targets which will be detected by the assay.
FAQ ID - 3896
What is the percentage of false negative results that QIAGEN obtained during validations?
We don't have such performance data for the Neo kit. In general, the kit is very sensitive with and LOD of around 100-1500 copies/ml or 8-12 copies per reaction. Sampling control increases negative predictive value.
FAQ ID - 3886
Why did we not choose commonly used genes such as the S gene, E gene, or RdRP?
The S gene is subjected to high selection pressure and has a very high mutation rate. Therefore, an assay in the S gene is prone to fail sooner or later. Our goal was to develop a long-term solution.

The E gene is very conserved among different Sarbecoviruses. As specificity is very challenging with the E gene, we did not choose it. As an example, the Charité E gene assay was designed to be inclusive for many Coronaviruses and is not specific for SARS-CoV-2.

Original RdRP assay shows suboptimal sensitivity with all assays we have tested – e.g. the Charité RdRP assay can be up to 10 Cts later than N1+N2, and the redesigned RdRP assay still showed a Ct shift of 4-6 Cts.
FAQ ID - 3892
Why did you choose ORF1a+ORF1b and ORF3a+ORF7a?
These regions were known to be well-expressed and have a favorable balance between conservation among related viruses and mutation prevalence/mutation hot spots. ORF3a and ORF7a have not been used earlier which gives us an advantage.
FAQ ID - 3893
Do we have specificity and sensitivity data for this kit?
Yes. The SARS-CoV-2 Neo Kit has been wet-lab checked against several respiratory pathogens, as well as bioinformatically assessed versus the NCBI database. The primers & probesb are specific for SARS-CoV-2. 
FAQ ID - 3882