Addenda to the Science paper

[V99]

Don't they also say that no mouse or xmrv infected cells had been in the lab?

 

[madmax]

Switzer's Assay

I may have overlooked something but I think they only used Switzer's assay to rule out contamination. All of the cell lines and 101 patient materials tested negative for mouse contamination.

We used Switzer's assay because he was very vocal that we were not detecting XMRV, but mouse DNA. His assay is a real time PCR that has not one but two fluorescent probes that is specific to mouse mitochondrial DNA.

 

[VillageLife]

We used Switzer's assay because he was very vocal that we were not detecting XMRV, but mouse DNA. His assay is a real time PCR that has not one but two fluorescent probes that is specific to mouse mitochondrial DNA.

Is that Max Pfost - wpi researcher, *waves* good to see you!

 

[SOC]

We used Switzer's assay because he was very vocal that we were not detecting XMRV, but mouse DNA. His assay is a real time PCR that has not one but two fluorescent probes that is specific to mouse mitochondrial DNA.

How delightful, then, that you found no mouse DNA contamination using the assay so "kindly" provided by Mr Switzer. I'm sure he was very pleased with the successful application of his extremely sensitive assay.

 

[Otis]

How delightful, then, that you found no mouse DNA contamination using the assay so "kindly" provided by Mr Switzer. I'm sure he was very pleased with the successful application of his extremely sensitive assay.

Yes, perhaps he began to doubt his own test.

@madmax, feel free to pop in anytime, your insights are surely appreciated!

 

[V99]

We used Switzer's assay because he was very vocal that we were not detecting XMRV, but mouse DNA. His assay is a real time PCR that has not one but two fluorescent probes that is specific to mouse mitochondrial DNA.

Amazing how confident these numpties are before they have even tested anything. Me thinks Mr Switzer is trying to imitate Reeves?

 

[anciendaze]

Amazing how confident these numpties are before they have even tested anything. Me thinks Mr Switzer is trying to imitate Reeves?

Psst! I have it on good authority this trait runs in their family.

 

[taniaaust1]

So nice of the CDC to help **grin**

 

[SOC]

We used Switzer's assay because he was very vocal that we were not detecting XMRV, but mouse DNA. His assay is a real time PCR that has not one but two fluorescent probes that is specific to mouse mitochondrial DNA.

BTW, did the use of his "extremely sensitive assay" specific to mouse mitochondrial DNA finally shut him up? About contamination, at least?

 

[madmax]

I would say they were satisfied that we conducted their assay, and presented them with the data.

 

[George]

I would say they were satisfied that we conducted their assay, and presented them with the data.

I would say that it was a good political and scientific move on the part of the researchers. (grins)

 

[V99]

Madmax, are there 8 patients missing from the addendum?

 

[Forbin]

Does anybody see anything or know of anything about "Highly Viremic" aspect of this?

I believe what they are saying is that Single Round PCR on DNA from PBMC's was a test of such low sensitivity for XMRV that, in order to get any results, they had to employ it on a subset of patients who were "highly viremic," which I assume means that those patients showed a high level of co-infection with other viruses, such as EBV, cytomegalovirus, HHV-6, etc...

I would also assume that if you were "highly viremic," you would have been automatically excluded from the CDC study.

 

[George]

I don't know Forbin, I don't see the correlation between co-infection viriemia and XMRV viriemia.

 

[V99]

The way I read it, was that they needed to show it was unlikely to be contamination, so they used normal PCR to test for xmrv. To make sure this would show positive samples, they guessed that those who display signs of high viremia, would be more likely to be identified as having xmrv using this basic method.

 

[V99]

That's not what you were asking was it. Do they mean severe symptoms that suggest a virus?

 

[George]

Actually V, I think that's exactly it. I think they had to sort of guess who would be the most likely to come up positive in the PCR. Everybody's got to start somewhere. (grins) Thanks V.

 

[V99]

So they created a subgroup for the basic PCR test. He he he.

 

[madmax]

Madmax, are there 8 patients missing from the addendum?

Yeah I noticed that, also noticed that they put the wrong version up. Should be changed soon.

 

[Forbin]

The addendum actually says:

In our October 2009 publication, we established XMRV infection in the blood products of our patient population by five different methods. Of these methods, single-round PCR on DNA from peripheral blood mononuclear cells (PBMCs), the least sensitive method, required us to use samples from a subset of chronically ill patients we had observed to have persistent viremia. In Figure 1A of our Science paper, we showed that DNA of 7 of 11 patients exhibited the expected gag and env PCR amplification products from single-round PCR with XMRV primers. We included this figure to demonstrate that nested PCR, which inevitably raises questions of contamination, is not essential to detect XMRV in highly viremic ME/CFS patients.

Although "highly viremic" might be interpreted as "showing symptoms of viral infection," the phrase "persistent viremia" indicates (to me, at least) the detection of a known virus or viruses.