[Cytometry] accreditation anyone?

Nebe-Von-Caron, G g.nebe-von-caron at alere.com
Fri May 13 17:56:41 EDT 2011

Sorry but this needed a long answer to be clarified so here it goes:

Anyone knowing me should actually know that I am classified as "mostly
harmless" and those who don't will hopefully take my word for it without
demanding a certificate. Those who refer to me as "nicht ganz dicht" are
actually correct as well. Sorry if I have come across picking upon a
particular paper in order to upset some people who tend to measure
output in dots per inch but most of you I hope would know if they read
my posts that this is not my style. I know that many believe that
Germans have no sense of humour, let me assure you they do, just the
question if it can be translated with those long sentences we make. Most
people will have forgotten the beginning of the sentence by the time we
come to the end, so in that sense:

In fact I actually did not want to diminish that paper in Any way as
there seem to be few about the subject anyhow. I actually liked their
approach to look at the diminishing fluorescence looking for significant
change and hoped that people would spot some of the parameters that are
important in achieving staining saturation and perhaps some people to
comment on the effects that could possibly cause the titration to be
different for the high intensity CD14 cells. I had speculated on three
different causes and hoped to get entries for possible controls. Looks
like some people seem to be too embarrassed to take on what was meant to
be a fun challenge and this mail might have probably taken the courage
out of many, so I might as well speculate in public so everyone can chip
in, accredited or not.
My theories were that it could be a higher numbers of cells, the
existence of soluble CD14 in the blood sample or the response of the
cells to put more CD14 on the surface in response to the antibody coming
onto the surface, an effect you can sometimes see with adhesion
molecules. In fact I once had seen a similar effect with CD14-PE that
with increased incubation time I got brighter and brighter CD 14 so I
was intrigued by the graph.

I hoped to compliment people for suggesting controls for those effects
such as washing the blood to remove free CD14 to test different cell
numbers, incubation times or temperatures or other ways to inhibit cell
activity and suggestions to look for patching and capping.

As unfortunately the staining conditions were not described in detail
but only referred to as "according to the manufacturers instructions"
which is perfectly correct statement - in fact you would be in trouble
if you did not use their protocol working in an accredited lab as you
are accredited not for your attempt to think but to follow a procedure
that is reproducible, not failsafe. As I could not get hold of those
(probably because BC restructures their site) I hoped people would come
forward with suggestions of what can change / influence the antibody
staining other than the concentration which would have obviously been
time and temperature. Now the authors were not meant to play around with
that for their paper so this is in no way a critique of them either.

The question on the blood sample was only as they did not mention the
anticoagulant which for example is one of the things that can affect
your adhesion molecule expression or the ability to do eat antibody....
- probably specified in the manufactures instructions as well. In that
context you could also consider other factors that are important when
taking blood samples like if the person was resting or just climbed up
flights of stairs...... 

Now if you feel they should have included all that stuff in their paper
you lost the titel. Some of the info would have been nice but was not
essential for what they wanted to show and they would probably still be
collecting data now. In fact you could of course use the method to look
after how long an incubation time you get no more benefit from it etc.

The second mail came about when I tried to copy the math started to
wonder if I should use the CV between the mean values of the different
blood samples or the CV for the CD14 distribution when I noticed
something odd with the values. I assumed the high CD14 to be 3% more
fluorescent than the normal ones with the mean peak going from 7000 to
7200 so I thought this was not so dramatic after all. Only then I
actually noticed something odd with the numbers which is why I asked the
bonus question. Now I could of course make a fool of myself here as I
know Joe's software to be quite good with getting the means tight when I
use it but it appeared to me that the data axis appeared linear whilst
the data look like they are log data assuming that the filled curve
represents the control (I don't bother about isotype or otherwise) and
the width of the positive population would make me think the same. I do
not know how that impacts the way of testing and I am happy to be told
how it should be done by someone with a certificate for statistics which
I have not. 

So whilst I think the approach is in principle correct following the fix
of the scale I would think Mario is probably the one to correct me it is
just to show once more that nothing is absolutely failsafe, not even a
review process - let alone an accreditation process, and of course I can
be completely wrong as well - and that would not be the first time

The only believers in the accreditation and certificates are those who
lack the belief so they need a certificate instead. Don't get me wrong,
we need process control to ensure that steps in a process are followed
which increases safety magins as humans tend to have a habbit of
forgetting things - or finding convenient shortcuts - but if we turn
paerwork it into a self fulfilling cult we only try to please the gods
of the paper pushers, the lawyers,  so that in case of a dispute you
have a certificate to show that someone has done a certain test or
trainig. Remember that paperwork is there to be kept to a minimum. The
rest of the time should be spent having ideas. To say it in Einsteins
"The intuitive mind is a sacred gift and the rational mind is a faithful
servant. We have created a society that honors the servant and has
forgotten the gift."

So sorry about the misunderstanding Mr Grivel, I am not into fighting. I
smile about the belief in the printed word and the hierachies built upon
it. It's not worth the figth as the logos will prevail in the long run
and why should I stop someone being happy about a form  - as long as
they do not waste my money for it but they charge the ones who want
them. Now if any of the listmembers ever assume - that is to make an ass
out of u and me- at least assume something positive so it's fun. In my
case assume that I do not take hostages (Honi soit qui mal y pense). If
I would they would probably be subjected to ribtickling until they start
laughing as I am very much into laughter. If I come across an enchanting
laugh I enjoy it for a long time, if from some friends I only call /
hear very occasionally or, as for example in a recent phone
conversation, with someone I did not know - but I think I do from the
laughter ;). So for those who are lucky enough to understand German
another link to youtube:


Enjoy and remember that people will keep harsh words and raised voices
in their head because they are coupled with emotions - as you should
know from the last link - but a laughter will stay in their hearts. So
thanks for the laughter and the smiles some people send me from time to
time and I hopefully raise with some of you. I feel sorry for those who
can't laugh so let's help them. Life is too short to spend it in misery!

Gerhard Nebe-von-Caron  | Research Scientist and Biomedical Engineer 

-----Original Message-----
From: cytometry-bounces at lists.purdue.edu
[mailto:cytometry-bounces at lists.purdue.edu] On Behalf Of Grivel, Jean
Charles (NIH/NICHD) [E]
Sent: 13 May 2011 03:14
To: cytometry at lists.purdue.edu
Subject: [Cytometry] accreditation anyone?

Dear Gerhard Nebe-von-Caron,

I have always enjoyed reading your posts and found your answers
extremely useful and filled with  invaluable knowledge. I must say that
the turn taken by your last posts stupefies me.  I understand and
respect your position regarding accreditation. I do not agree with the
way of have chosen to make your point.   I doubt that your trivia brings
anything to the debate, it singles out a few persons who may have
benefited from more constructive intervention on your side.
My point is that you could peak the fight with accreditation grating
institution, ISAC, teaching institutions, Peer review at Cytometry,
etc.. The authors of this paper have tried to improve upon a widely used
method, while imperfect, their effort should be recognized and improved
upon by more knowledgeable scientist like yourself. These author should
not singled out as examples of what is wrong with accreditation , and
they certainly should not be kept hostages of this fight.
Jean-charles Grivel

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