Kinloch NN, Ritchie G, Dong W, Cobarrubias KD, Sudderuddin H, Lawson T,Matic N, Montaner JSG, Leung V, Romney MG, Lowe CF, Brumme CJ, Brumme ZL. SARS-CoV-2 RNA quantification using droplet digital RT-PCR. The Journal of Molecular Diagnostics. 2021 May 29. doi: 10.1016/j.jmoldx.2021.04.014.
The results and/or conclusions contained in the research do not necessarily reflect the views of all CITF members.
A group of researchers at Simon Fraser University, the BC Centre for Excellence in HIV/AIDS and Providence Health care led by CITF-funded researcher Dr. Zabrina Brumme have developed a method to quantify SARS-CoV-2 virus levels in human biological specimens using a technique called droplet digital RT-PCR (RT-ddPCR).
Key points:
- This method can be used to determine the exact number of copies of SARS-CoV-2 in a biological sample without the need to calibrate to a standard curve
- The method can also normalize the number of viral copies by the total amount of biological material collected, which can be important for samples collected on swabs, like SARS-CoV-2
- The team also developed a mathematical equation to determine the viral load of a sample from existing diagnostic test results without re-testing the samples
This recent work published in The Journal of Molecular Diagnostics is funded by the CITF, Genome BC and the Canadian Institutes for Health Research. The team felt that this assay was important to develop because the molecular diagnostic tests currently used for COVID-19 yield only semi-quantitative results: positive, negative or indeterminate. While these classic tests do produce a numeric readout (called the Ct value), these values are not standardized across diagnostic test platforms.
While semi-quantitative tests are sufficient for diagnostics, the ability to accurately quantify viral copy numbers can be very useful in some contexts, for example, when monitoring the effect of an intervention on reducing virus levels in the body.
The technique described in this manuscript offers an attractive platform: the test requires a very small amount of sample and does not require calibration to controls.
The team also developed a way to normalize the number of viral copies by the total amount of biological material collected. This is important because SARS-CoV-2 samples are collected on a swab, which is then put into liquid preservative. Because variable amounts of material can be collected on the swab, and different swabs come with different liquid preservative volumes, it can be useful to normalize viral levels to the total amount of biological material collected, so that viral levels can be meaningfully compared across samples.
Importantly, the team also developed a mathematical formula to infer the viral load of a sample from the original diagnostic test results. This means that existing diagnostic test results can now be converted to actual numbers of SARS-CoV-2 viruses without re-testing samples.