[Cytometry] Teaching Flow Cytometry

Nebe-Von-Caron, G g.nebe-von-caron at alere.com
Fri May 6 14:20:52 EDT 2011

Hi Till

First of all you should let  your users appreciate the difference
between a flow cytometer and a fax machine, particularly as you use a
particle analyser that can do other things than only fluorescent
activated cell sorting .  Reading TFM, The Fantastic Manual, is a good
start but usually not as entertaining as reading Howards book to learn
how to obtain low noise valves for building excellent amplifiers.
However there are a couple of small "Introduction to Flow Cytometry"
booklets available for free from vendors who want your address details
in return, even Howards big book. Once potential users have read through
one of the smaller booklets you have a good discussion basis, so let
them read one of those before they come to see the person who is
responsible for the cytometer. And learning works much better if you
have some basic information about the things you want to learn more

You can teach students all sorts of tricks on how to do things and the
basic principles of flow cytometry, but the thing that helps to be a
really successful cytometrist is to have "the Knack"
http://www.youtube.com/watch?v=CmYDgncMhXw  The most important element
in teaching is to live by example and to transfer the enthusiasm and
passion you have for your scicence.  To prime the enthusiasm of others
it helps to explain why this technology is such a central research tool
in science. It comes from its ability to differentiate cells / particles
and generate population statistics as it can measure the relevant
numbers and features of cells particles, identify
subpopulations/subcommunities with similar properties, find (rare)
events and separate events of interest by gating or regions of interest
and sometimes even physically separating them. The key is in the
appreciation of learning that all systems are made up from individual
units with differences from each other as those differences create
complexity and stability. To understand the the "big picture" you have
to appreciate the differences at the elementary level, or as said by
''to find you in infinity of space and time one must first divide and
then combine", the basic theorem of scientific studies such as systems

There are a lot of cytometry courses around and usually if you have a
manager above you he/she will be much happier to send staff on a course
elsewhere than having you set up an in house course. First of all it is
difficult to budget, but more important, for a manager a trainer /
consultants outside the managers company is always better / more
intelligent than in house staff as they are not working under the
managers supervision, e.g. some managers tend to have difficulties
admitting that someone below them can train people in something that
they cannot train in themselves, thus admitting that they are not on top
of everything. If your boss is a leader and not a manager that's
different. Institutes and Universities are a bit more open in this as
they are more used to the idea of teaching but as you have a couple of
enthusiasts up / down the road and considering that you are not the only
cytometrist in Darmstadt (in fact if I am not mistaken you have even one
of the original cytometry specialists living quite near) I would contact
some of the local flower gurus for example in Heidelberg as it is always
easier to organise training course together with some other
cytometrists, particularly if you are not an expert yourself. They tend
to be very helpful bunch, each of them having their own area of
expertise which they can be very enthusiastic about. Also some of them
already run courses, most of them on a break even budget / cost basis
and you might as well want to point your new starters to such courses
because of the networking opportunity. Other considerations should be
whether you can or cannot get an in house budget for the time you spent
in training people. If in house training is seen to be "for free" it is
usually not treated with the same respect. In the end you still need to
conduct some in house training including other steps to consider such as
lab internal processes, data handling and particularly health and safety
issues. Howard correctly said that unfortunately a lot of students come
up with the idea of it all being hush hush wizardry of which they do not
need to understand the minor details as they use a cytometer with an
attached operator. Ideally they want to be served with a 12 colour
antibody panel and someone to prepare things for them and clear up after
them, push a button and become a manager writing in their CV that they
done 12 colour cytometry. You don't do them a favour by not letting them
work out things for themselves as scientific thinking is not achieved by
giving them answers but by learning how to phrase the question and what
to use as control.

Have fun


-----Original Message-----
From: cytometry-bounces at lists.purdue.edu
[mailto:cytometry-bounces at lists.purdue.edu] On Behalf Of Howard Shapiro
Sent: 06 May 2011 14:53
To: Dettmering, Till
Cc: hsfc at elist.tufts.edu; flow list Purdue flow list Purdue
(cytometry at lists.purdue.edu)
Subject: Re: [Cytometry] Teaching Flow Cytometry

Till Dettmering wrote:

> I'm a PhD student for a year now and I'm working with flow cytometry
for 2.5 years. Since there is no real dedicated FACS-'guru' in our group
who I could have learned from, I had to teach practically everything
about flow cytometry to myself, including maintenance of the device (a
Partec PAS III). I find myself more often in the situation in which I
have to teach new students who have never seen a flow cytometer how to
work with it and how to interpret the data. Although it works out quite
well, I'm very interested on your experiences on how to best teach new
people flow cytometry. What do you start with? Do you have something
like curriculum you follow when teaching it? What did you start with
when you learned it? What should one pay special attention to when
> I'm very interested in hearing your experiences!

Problem number one these days seems to be motivating the new people to
think; too many of them want to mix samples with magic juice, put them
in the magic box, press the magic buttons on the box and do the
necessary magic mouse clicks, keystrokes, etc. on the computer, and save
the PowerPoint presentation and the manuscript that will be submitted
online to one prestigious journal or another. Then it's back to social

Those of us who write flow books and vent on this Mailing List have
tried to spell out the basics. Cytometry, as I have often said, begins
with the cell, even at the level of the word itself. You have to know
what information you want from the cells in your samples, and what
reagents you need to use and parameters you need to measure to get that
information, and it helps to consider alternatives when they are there. 

The cell identification game is like "Where's Waldo (his name is Wally
in some places)?" - The game wouldn't be any fun if Waldo were the only
person in the crowd wearing magenta and chartreuse; the intellectual
challenge comes in picking out physical features and the patterns of the
colors on his clothing and distinguishing them from those on everybody
else in the crowd among which he is hidden. Generally speaking, one can
always identify a cell from a given species by detecting a specific
nucleic acid sequence, or a combination of antigens. In unlysed blood,
CD45 is good for discriminating white blood cells from red cells but, in
cases where you know you won't encounter nucleated red cells, it's much
simpler to use a permeant DNA dye. Simpler is always better.

One can learn these kinds of tricks from books and material available
online. It's harder to learn how to set up an instrument, verify that it
is working properly, and deal with it if it isn't. The theoretical part
of that is accessible in books, but which knobs you turn and/or how you
set up the software depend entirely on the specific cytometer you're
using. Manufacturers offer training courses, and provide manuals, but,
as is the case with computers, you can sometimes RTFM and find that it
doesn't really tell you all you'd need to set up a flow lab in a
postapocalyptic venue. Oral tradition only goes so far; even prehistoric
hunters seem to have needed seminars with presentations painted on cave

When TFM leaves you in the dark, you can get computer questions answered
online, sometimes for nothing, or you can buy something like the books
in David Pogue's "the missing manual(R)" series, published by O'Reilly.

What we need in cytometry are equivalents. Practical Flow Cytometry had
its genesis in my "Building and Using Flow Cytometers: The Cytomutt
Breeder's and Trainer's Manual," and the material from that on building
and using instruments, which appeared in the first couple of editions of
PFC, has apparently been sufficient to enable dozens of people to put
together instruments and keep them running for decades without access to
service from outside. 

It does not strike me as unreasonable that manufacturers should support
their user groups in generating similar documentation, but the users
could get organized to do this without the manufacturers' help, which
would minimize the likelihood of problems being swept under the rug. The
material, which should cover software from both manufacturers and third
parties as well as apparatus, should be kept online in a presumably
neutral site such as this one. 

I've seen people with little or no lab experience get up to speed, at
least with relatively simple flow cytometers, in weeks, and people with
decades of experience fail to do so in months to years, and I can't
always explain either.  There hasn't been a Mozart in cytometry yet, and
even he started small. If you can't hear a sonata playing in your head,
don't try for a symphony, and, if you can't do two-color fluorescence
and two-angle scatter, don't try running a 4-way sorter with 7 beams and
24 colors. I spent several years training in surgery, and would have had
to spend several more before anybody let me do any significant part of a
heart operation; it is, admittedly, harder to kill people by running a
flow cytometer incompetently than it is to do so when you are waving a
scalpel around vital organs, but the malpractice of adding junk to the
medical and scientific literature has its own adverse consequences.

So let's get organized. 

I'll be at my 50th College Reunion during ISAC, which declined my offer
to give a plenary talk by remote video, but catch up to me online if
you're willing to help.


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