3CR Bioscience have developed ProbeSure™ COVID-19 One Step RT-PCR Kits to help fight the battle against Coronavirus SARS-CoV-2. The key challenges are:
- Increasing throughput without significant infrastructure investment
- Low copy number detection to address the issue of false negatives
- High reagent costs
Click on the links below to see how the ProbeSure™ COVID-19 testing RT-PCR kits developed at 3CR Bioscience Ltd addresses these challenges.
▾ Operational Efficiency
▾ Endpoint Detection COVID-19 testing is generally performed by real-time PCR (qPCR). Increasing the throughput can only be achieved by increasing the number of qPCR machines, along with an associated increase in FTE and infrastructure. The ProbeSure™ COVID-19 One Step RT-PCR kits can be analysed in real-time, but they have been optimised for endpoint analysis also. Endpoint analysis allows a far greater rate of data generation and a similarly increased rate of data analysis compared to real-time analysis. Using the endpoint PCR approach, bulk thermal cycling can be carried out on any Peltier or water bath based system and a single plate reader can then be used to quickly analyse many more PCR plates per unit time than would be the case for real-time PCR analysis.
Real-time data compared to endpoint data
▾ Technical Enhancements
▾ Low Copy Number Detection The ProbeSure™ COVID-19 One Step RT-PCR Kit has been developed to allow very high sensitivity, which has been tested on patient samples as well as FDA approved synthetic RNA from Twist Bioscience. The synthetic RNA samples were tested using ProbeSure™ COVID-19 One Step RT-PCR kits containing assays manufactured by Integrated DNA Technologies™ (IDT). The three markers assayed were N1, N2 and RNase P. A serial dilution was performed on the control sample with the resulting sensitivity shown to be as low as 1 copy per μL in a 5μl reaction. Using 5μL volumes enables testing to be performed in 384-well PCR plates, reducing both time and expense.
RNA control dilution series test results
▾ Optimised Endpoint Clustering
ProbeSure™ COVID-19 One Step RT-PCR kits were used to analyse suspected Covid samples in both real-time and endpoint. A plate of samples was thermally cycled on an ABI7900HT real-time instrument with samples shown to have Ct values ranging from 12 to 37. The plates were subsequently analysed in endpoint on a Tecan Spark plate reader and showed 100% concordance with the real-time data. The link below shows how this wide range of Ct values can be viewed as a tight cluster in endpoint analysis. Distinct endpoint clusters are seen containing all samples, despite the real-time Ct range being as broad as 12 to 37.
Wide range of Ct values produce tight cluster plot with ProbeSure™ COVID-19 One Step RT-PCR kits
▾ Multiplex Capabilities
The ProbeSure™ COVID-19 One Step RT-PCR Kit is available with a choice of singleplex or multiplex assays. Running a multiplex reaction further increases throughput and analysis rate, whilst reducing the cost per reaction. The multiplex assay, which combines N1, N2 and RNase P assays, produces data for all three markers in one PCR plate well per RNA sample, enabling a three-fold 3x increase in capacity.
▾ Cost Benefits
▾ Low Reaction Volume
The ProbeSure™ COVID-19 One Step RT-PCR Kit has been formulated such that testing can be performed in very low reaction volumes in higher density plates without affecting the performance, reducing cost per test very significantly. The results show above are all from 5μL reactions containing 2μL of sample.
▾ Kit Pricing
The ProbeSure™ COVID-19 One Step RT-PCR Kit has been priced to allow more data to be derived for a set budget. See our pricing table for costs.