Thanks for that. Hence, what are the chances that there are labs performing this service on a large scale on behalf of Pro Tour Teams do you think? Or is recombinant EPO really the only viable option at a commercial level?
In closing however… do you think it’s theoretically possible to extract enough EPO from your own urine that you could reinject it to artificially raise your haemotocrit levels to the magic 50-52% mark? I’m told you’re allowed a 2% margin of error in the UCI regulations and that 50% is the cutoff mark before you’re ruled out of a race on medical grounds but not given a suspension.
You’re not going to get anywhere in this debate till you realise that there ain’t no such thing. The whole tour has been under a great deal of scrutiny for several years. Several riders have been caught or as good as caught by other means while testing absolutely positively certifiably clean. David Millar comes to mind.
It’s not. But it has nothing to do with this debate because no one’s saying he dopes because he wins. It’s a fundamental misunderstanding that happens every time we have this debate.
What will constitute proof?
Threemae, unless you were in France a few weeks ago racing around on your tricycle, you don’t have a rooster in this cockfight. So why the hostility?
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It doesn’t need to be insidious. Again, I guess you can doubt my expertise, and anyone who has ever run more than five gels in their lives can jump in and support or refut this, but sometimes bands just appear. Gels, be they Southern, Western (2D is just a more impressive Western), Northern or DNA are notorious for giving bands that aren’t supposed to be there.
About a month ago, I pulled up a band of exactly the size I was looking for on a DNA gel. It would have been a huge finding. Problem is that I sequenced the band and it turned out to be the gene for prostaglandin E. What the hell the gene for prostaglandin was doing in this sample, I have no idea, but had I gone just by the gel, I would have concluded that I had an interesting finding that I didn’t have. I avoided this by repeating the experiment several times and not being able to pull that band up again, leading me to question it and sequence it.
Lest you question my gel abilities, you’ll always see talks where people present their data, and just ignore bands that don’t make sense. There is even a term for them; they’re called “artifacts” and they pop up all the time.
Sometimes bands just appear. It’s called false positives. Which is why I say if they could repeat this with another LA sample, I would have been a lot more impressed. One gel doesn’t mean a thing.
Well, as far as facilities go, I supppose, though I don’t think it probable. I’ve heard of grad students setting up some positively state-of-the-art LSD or meth labs in a loft, so I guess one has to keep the “mother of invention” factor in mind.
As for harvesting human EPO, in the seminal paper on the subject, Miyake, et al. got about 10mg of EPO from 2550 liters of urine from people with aplastic anemia, a condition characterized by abnormally high levels of circulating erythropoietin (the EPO conc. in the urine collected was around 1-2 IU/ml - to put this in perscpective, dosing for anemics is around 20 IU/kg, 3 times per week). Getting EPO from one rider in useful amounts is going to be a chore. But no matter. Just get a boatload of urine from whoever; it’s all the same protein. It’s got to be handled reasonably carefully and processed quickly, because, as we’ve been discussing, urine isn’t the best stuff to keep EPO lying around in.
So sure, it’s possible one could be collecting urine from scores of people and passing big vats of it over columns and so forth, but this is not a small operation. The whole reason EPO was one of the first recombinant proteins brought to market is because this process, while not a total money loser, is a huge, messy, costly pain in the ass. Can it be done? Sure. Is it likely? I personally really don’t think so, but who knows.
“Or as good as caught” doesn’t fly with me. There’s either proof or there isn’t. In Armstrong’s case, there isn’t. The “evidence” presented is laughable. There are strict protocols re labeling, logging, handling of material, etc. that labs need to follow. Reporters are neither trained nor qualified to run drug tests. And they are biased. Therefore, their “conclusions” are worthless.
We’re having this debate because people are claiming LA dopes but have failed to offer proof. And even if one day someone offers up some empirical data to support these claims, it still does not make me a “bitch” for being healthily skeptical. Rational people should be skeptical, especially when the source is so clearly biased.
Yes, and twelve distinct bands of them just hapen to consistently be in the exact location where synthetic EPO shows up, in urine, which doesn’t contain a lot of protein anyway.
And remember, the lab doesn’t have to really do any extraction to run these samples. They aren’t putting Lance Armstrong in a blender, grinding him up, and using chromatography to get out the EPO. They’re running a straight 2-d gel from his urine. Again, what’s more likely here? The director of the French lab, the Canadian lab, and all of the other anti-dope labs agree that synthetic EPO bands don’t, “just show up,” in their samples.
It may or may not be laughable, but it is most certainly experimental, and simply should not hold weight when determining guilt or innocence. There is, IMO, considerable reasonable doubt about the integrity of the entire procedure, from storage, to handling, to interpretation of data, to the manner in which the data was revealed. No one has any business declairing guilt about anyone based on this data, even if it could be proven to be 100% correct, in no small part because it can’t be, by the lab’s own admission. What’s laughable is the lack of anything resembling integrity and due process. We know results we should never have known about a test that is not officially sanctioned and cannot be further verified about an individual who just happens to be the best cyclist in history.
Unacceptable. Dubious techniques, dubious motivation, dubious ethics, insufficient regulatory oversight, probably non-actionable, and most certainly worth maintaining the highest standards of privacy over. The fact we know any information about individuals whose samples were being used for validation of an assay that has not been fully vetted by the appropriate regulatory bodies is no better than illegal search-and-siezure, IMO. And when people show so much enthusiasm for catching an individual that they sacrifice integrity in the process, serious questions about evidence tampering are most certainly NOT out of line.
It wasn’t “consistent”. It happened once with one sample. I’m making one simple statement that an n of 1 is nice, but no conclusions should be drawn. Any number of things can happen to make a false positive once. If it comes up positive from three to five independent samples, then I’ll conclude that he is doping.
Whether the science is bad or good, reliable or unreliable is not nearly as important to me as the incredible breach of ethics that has gone on here that allowed L’Equipe to even make this an issue.
If he is guilty, then he sure is a good actor - he does the righteous indignation thing really well. He has shown up on all the big TV interviews sans lawyers, plus he is speaking to print reporters freely.
It seems to me he is either setting himself up for a big fall, or else he is, you know, innocent.
Just to be sure that this “expertise” of yours was acquired naturally, would you mind peeing in this cup?
What’s the difference between the natural stuff and the recombinant stuff?
The amino acid sequences are the same, but post-translational modifications would be different. I believe that they are glycosylated differently, meaning that sugars are attached to the protein in different conformations.
Post-translational modification. Human EPO and recombinant EPO have the same amino acid sequence, but chinese hamster ovary (CHO) and baby hamster kidney (BHK) cells, which have been used to produce it in large quantities in vitro, make slightly different modifications to that amino acid chain once it’s been polymerized and mostly folded than the human kidney cells that naturally produce EPO in adults. Specifically, human cells normally modify EPO so that specific sorts of sugar chains are added to four particular amino acids (three different asparagines, which provide the N-links and one serine, which provides the O-link), though there is a natural heterogeneity in the number of sugar subunits (specifically sialic acid) that get tacked on. CHO and BHK cells do a pretty good job, but after they process EPO, it’s not quite the same as the human variety. You can find characteristic differences in sialation, in particular, which are due in part to the basic biochemistry of the cells, and in part to the batch process and the contiditinos the protein is exposed to in culture. These differences can be seen with a sufficiently sensitive assay.
It’s actually somewhat surprising to me that a 2D protein gel electrophoresis method would be favored over a chromatography or mass spectrometry method, but maybe it’s a convenience thing, I don’t know. Anyway, depending on the numbers of these sugars, and where they are, there will be mass and charge differences between EPO glycoforms, and those glycoforms can be resolved on a 2D gel, which first separates proteins by charge, and then by apparent size.
Yes. There may be slight differences in immunogenicity, and so forth, but for the most part recombinant EPO and native human EPO are remarkably similar.
Well, what exactly do you mean by guilt or innocence? If this were a criminal trial over doping, there are certainly, “shadows of a reasonable doubt” and I wouldn’t hesitate to acquit Mr. Armstrong if I were on a jury. But this isn’t a criminal trial. As observers of the sport, we’re well within out rights to decide if Mr. Armstrong is a doper or not. Now the evidence is pretty overwhelming that he is. 95 to 5, I’d venture. And what will result from all of this? Nothing. There’s not a snowball’s chance in hell that he’ll be sanctioned for this because the A sample was used up back in 1999, and he shouldn’t be because athletes have a right to due process and A and B sample testing with the same positive results. I’ve said this earlier in the thread if you had bothered to read it. You don’t disagree that it’s now clear that it’s more likely than not that Armstrong doped, do you?
