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Cliff Notes of 4 papers that Dissolve the Corman-Drosten RT-qPCR test.
https://twitter.com/Kevin_McKernan/status/1335357508618252289?s=20
https://twitter.com/Kevin_McKernan/status/1335357508618252289?s=20
We'll start with Muenchhoff et al. published in Eurosurveillance which concludes with Charite' RdRp assay is faulty and needs to be replaced. This manuscript demonstrates sensitivity issues. This would support False Negatives. But you must understand why it creates FNs...
The authors report reduced sensitivity. That is a direct result of the authors skipping simple primer design QC steps. Screening for Primer Dimers or Primer Hairpins is 20 year old free software on the web.
This is the RdRp probe. Hairpins and homodimers.
This is the RdRp probe. Hairpins and homodimers.
RdRp also has homology to the E primers which we report in Corman-Drosten report. While most labs run these tests in different wells (1-plex), its bad practice to have primer dimers between your 1-plexes knowing liquid handling of Millions of tests can create X-contam.
Lets see how these problems manifest and you'll get a better understand of whats going on.
Charite' RdRp assay craters. Author propose a fix. This fix is not at the WHO website.
Charite' RdRp assay craters. Author propose a fix. This fix is not at the WHO website.
Guess who is include on the author list which concludes these primers must be replaced.
Published June 18th.
But last time Drosten went to the WHO before he went to Eurosurveillance?
Why not this time?
Published June 18th.
But last time Drosten went to the WHO before he went to Eurosurveillance?
Why not this time?
"BUT REDUCED SENSITIVITY DOESN'T CREATE FPs".
I present to you Jung et al.
Promiscuous primers, not only fails to amplify your targets in some samples, they also amplify things they shouldn't in other samples. In this case they amplify Water (NTC).
https://pubs.acs.org/doi/10.1021/acsinfecdis.0c00464
I present to you Jung et al.
Promiscuous primers, not only fails to amplify your targets in some samples, they also amplify things they shouldn't in other samples. In this case they amplify Water (NTC).
https://pubs.acs.org/doi/10.1021/acsinfecdis.0c00464
Outlier you say.
I present to you Etievant et al.
Also find the E assay producing positive signals with water samples. This is other set of primers that are predicted to create dimers ONLY IF RdRp is around.
I present to you Etievant et al.
Also find the E assay producing positive signals with water samples. This is other set of primers that are predicted to create dimers ONLY IF RdRp is around.
Reports of such primer contamination arent just theoretical, they are reported in the peer reviewed literature (Etievant et al). Note, the study CT values are in question as the CD paper didn't disclose this important detail.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7355678/
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7355678/
2 papers is just Cherry Picking +1.
I present to you Gand et al.
https://www.mdpi.com/1422-0067/21/15/5585
Note the lack of specificity in this paper for Charite' primers.
I present to you Gand et al.
https://www.mdpi.com/1422-0067/21/15/5585
Note the lack of specificity in this paper for Charite' primers.
False positives mentioned in addition to reduced sensitivity.
There is more evidence of these artifacts on this thread.
In summary, the most popular qPCR test in Europe and on the WHO website is published as flawed in multiple peer reviewed articles and Drosten is on the author list making it really hard to deny. https://twitter.com/kevin_mckernan/status/1331564279254933507?s=21
In summary, the most popular qPCR test in Europe and on the WHO website is published as flawed in multiple peer reviewed articles and Drosten is on the author list making it really hard to deny. https://twitter.com/kevin_mckernan/status/1331564279254933507?s=21
Racing to the WHO to get them the primers your buddy sells or lab friends use, before you embark on a 24hr rubber stamp review had urgency.
Correcting the record at the WHO, when it became obvious (with Muenchhoff and others) that water samples were amplifying, Not so urgent.
Correcting the record at the WHO, when it became obvious (with Muenchhoff and others) that water samples were amplifying, Not so urgent.
There is more detailed version of this here: https://twitter.com/Kevin_McKernan/status/1333846936332464129?s=20
