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https://biobot.io/biobots-wastewater-monitoring-the-omicron-variant/ https://biobot.io/biobots-wastewater-monitoring-the-omicron-... indicates they use PCR. How

by epakai 5y ago

https://biobot.io/biobots-wastewater-monitoring-the-omicron-variant/ https://biobot.io/biobots-wastewater-monitoring-the-omicron-... indicates they use PCR. How does PCR testing return a quantitative result? I thought the purpose was to amplify (reproduce) proteins to enable detection, and this precludes finding original concentrations.

jhj 5y ago
The cycle threshold (Ct) is the quantitative value from PCR. This is mapped to counts per volume based on results in https://pubmed.ncbi.nlm.nih.gov/32607521/ https://pubmed.ncbi.nlm.nih.gov/32607521/ I believe.
- dsizzle 5y ago
  Yes. I think the device used is a bit more sophisticated than simply obtaining the cycle number where it crosses the threshold. There's a variation of PCR called quantitative PCR (qPCR) that measures a fluorescence signal more or less continuously and the growth rate allows one to work backwards to the initial count. Here's a basic description from the maker of the device they used (Taqman) https://www.thermofisher.com/blog/ask-a-scientist/what-is-qpcr/ https://www.thermofisher.com/blog/ask-a-scientist/what-is-qp...
  - mlyle 5y ago
    There's a lot of ways you can get fancier and more precise quantitative results, sure. The basics is the cycle threshold. The next step further is just to try and come up with a fractional cycle number through interpolation-- e.g. if cycle 20 was not-quite-positive-but-very-close and cycle 21 was strongly positive, that implies a value close to 20, versus if cycle 20 was very far from positive and 21 was just barely over the threshold, that implies the real value is just under 21. And there's further, but diminishing gains from fitting the fluorescence curves.
jonas21 5y ago
https://en.wikipedia.org/wiki/Real-time_polymerase_chain_reaction https://en.wikipedia.org/wiki/Real-time_polymerase_chain_rea...
ImaCake 5y ago
You can measure how efficient a PCR is by how long it takes to get a positive result. PCR requires a loop of different temperatures to cause different steps in the reaction to occur. This loop is usually repeated 30-40 times. You measure if its positive by looking for a (continuous) light signal. The light signal increases each time you loop through the temperatures. When the light signal passes a pre-determined threshold, that cycle is said to be the one where you got the positive result. The more virus DNA/RNA in the sample, the less loops you need for a positive result.

by epakai 5y ago

jhj 5y ago

The cycle threshold (Ct) is the quantitative value from PCR. This is mapped to counts per volume based on results in https://pubmed.ncbi.nlm.nih.gov/32607521/ https://pubmed.ncbi.nlm.nih.gov/32607521/ I believe.

dsizzle 5y ago
Yes. I think the device used is a bit more sophisticated than simply obtaining the cycle number where it crosses the threshold. There's a variation of PCR called quantitative PCR (qPCR) that measures a fluorescence signal more or less continuously and the growth rate allows one to work backwards to the initial count. Here's a basic description from the maker of the device they used (Taqman) https://www.thermofisher.com/blog/ask-a-scientist/what-is-qpcr/ https://www.thermofisher.com/blog/ask-a-scientist/what-is-qp...
- mlyle 5y ago
  There's a lot of ways you can get fancier and more precise quantitative results, sure. The basics is the cycle threshold. The next step further is just to try and come up with a fractional cycle number through interpolation-- e.g. if cycle 20 was not-quite-positive-but-very-close and cycle 21 was strongly positive, that implies a value close to 20, versus if cycle 20 was very far from positive and 21 was just barely over the threshold, that implies the real value is just under 21. And there's further, but diminishing gains from fitting the fluorescence curves.

jonas21 5y ago

https://en.wikipedia.org/wiki/Real-time_polymerase_chain_reaction https://en.wikipedia.org/wiki/Real-time_polymerase_chain_rea...

ImaCake 5y ago

You can measure how efficient a PCR is by how long it takes to get a positive result. PCR requires a loop of different temperatures to cause different steps in the reaction to occur. This loop is usually repeated 30-40 times. You measure if its positive by looking for a (continuous) light signal. The light signal increases each time you loop through the temperatures. When the light signal passes a pre-determined threshold, that cycle is said to be the one where you got the positive result. The more virus DNA/RNA in the sample, the less loops you need for a positive result.