7 replies to this topic

[LivingWellWithCML]

Hey all,

So, I'm still wondering a bit about PCR sensitivity.  I got my report today (peripheral blood PCR test) and this is what it says:

• Reliably detects 10 positive K562 cells in a background of 10(to the 6th) negative cells.  The ratio they are calculating is BCR-ABL1/G6PDH.

For Emory Winship in Atlanta, the baseline ratio is still a mystery to me (and I've given up asking them about it), but is "ten positive in a background of one million" considered good PCR sensitivity?  And is G6PDH a pretty standard housekeeping gene used for BCR-ABL PCR?

Thanks for your help...

[valiantchong]

10 / 1 000,000 meaaning you are almost -6 log, that will bring you to almost -6 log PCRU....Most of the lab only detect -4.5 log.. I am surprised that the lab you test is that sensitive. Wondering if this is tie to IS scale ? If it is tie to a IS scale it will be then multiply by another factor....

There are record that some more sensitive lab could detect -7 or -8 log...That is the present limit I think...

Yes I think BRC-ABL1 is a standard reference gene used most of the time these days...

[LivingWellWithCML]

Emory isn't on IS.  They have their own mystery baseline ratio that is based on an average PCR ratio for CML Chronic Phase patients who have not received therapy yet...presumably Emory-only data.  When I look at their baseline, I think this sensitivity level is more like -4.0 log as the limit of detection, but I don't have a way to compare this to IS...

[Trey]

Really, unless a lab has ancient equipment and uses outdated primers, the sensitivity is about the same everywhere.  It is advertised generally as 1 in 100K.  From what I can tell, Aussies are using old stills from the woods of Tennessee for PCRs, and Moonshine primers aged in Kentucky Bourbon barrels.  But other than that, the difference is mainly lawyers arguing about how many logs they allow to be advertised as their accuracy.  I have previously stated that PCR equipment manufacturers say that their equipment is not authorized as medical diagnostic tools.  In fact, the #1 PCR maker in the world has this warning on their  machines: "For life science research only.  Not for use in diagnostic procedures."   Ha ha ha, that is a good one, since that is what is being done on all of us. 

So your PCR is certainly good enough.  However, the G6PDH housekeeping gene has been abandoned by most labs for BCR-ABL PCRs.  It is generally accepted that ABL or b2m are the better choices.  But as long as you always use the same lab, the trend is the most important indicator of response.

IS and sensitivity are two separate issues.

Georgia is close to Tennessee, right?

[valiantchong]

Hi,

Normally the detection log detection limit is stated in the report. My report max limit detection varies from 1 to 10K - 50K normal cells reference back ground meaning ~ 4 log to -4.5 log. And this ratio is then multiply with a baseline factor to tie to IS scale.

Well, could ask what is the confirm limit of the detection in your lab... Anyway I guess you are doing great near to PCRU,,, congratulation or did you ask your doc if you could be considered PCRU ?

There are reports that on present PCR dectection limit on some normal healthy person could be - 7 or - 8 log meaning 1 cell in 10, 000, 000 to 100, 000, 000 normal cells. About 70% of normal people could be detected. But I do not know if this is due to our present technology limit of due to the sensitivity error is inherent in the detection method.

I know that DNA-PCR method is more senstive to RT-PCR or Nested PCR technic, nevertheless you could check out the method used in your lab and how confident if the detection method used with your doctor. The DNA-PCR method was used to detect some successfull STIM trial patients to check if for MRD, however only 1 or 2 still found to be still undetectable - PCRU. But most of them are detectable with DNA-PCR.

[Pin]

Booooo! Don't make me worry about the standard of our testing equipment! I hope this is just one of those jokes about how backward we all are down here, kangaroos to work etc.

[LivingWellWithCML]

Yup, this must be a Georgia / Tennessee thing - I'd like to get ahold of that primer ... I hear it packs quite the punch.

Why are ABL and b2m better choices than G6PDH - is there a difference in accuracy?  For no good reason whatsoever, I'm now wondering why a leading research hospital in a major metropolitan area would use an outdated housekeeping gene when they could get a better test (?) with a more mainstream choice.  And I just checked all of my PCR reports post-dx and they are running RQ-PCR.  The report only states that "-3.0 log reduction or greater from Emory's baseline" is an MMR, but it doesn't state the maximum log reduction that the test can reliably detect.  I've calculated their baseline ratio from past reports using my actual log reduction levels (Baseline Ratio ~ 0.07900), and it looks to be ~ -4.0 log reduction max sensitivity.  Unless I choose to fly to Houston or Portland every 3 months vs. driving 25 minutes into downtown Atlanta, then I will need to settle into this level of sensitivity.

Thanks for your input -

[valiantchong]

Hi,

Not to be too worry on the testing, most important is that you know you are in MMR, and doin very well... unless you are planing to be in the STIM trial. Then you will need to have a more sensitive measurement method.

Well I guess most important you know is that you have the present baseline and need to monitor your trend base on your present baseline as long as it is from the same lab and is trending downwards or not goin up trend...you are assured you are doing well, that is what more important...