How Many list are show on the Purdue cytometry mail list for software sales...

[Unregistered]

Re: Last words from Mario (we hope) on data display



From: Howard Shapiro (h...@shapirolab.com)
Date: Sat Oct 04 1997 - 22:16:13 EST


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I will propose to Cytometry to write a perspective on the topic of FACS data display.


>I challenge any of those of you who


>champion dot plots (or even color dot plots) to join my effort and write a
>"counterpoint" analysis to provide a balancing viewpoint.


A forum with various viewpoints, well documented and illustrated,
would probably be very helpful to the readership.




>Calman writes about dot plots:

>> Furthermore it stresses the single cell nature of the
data- each dot is a cell.

No, no, no, no, no, no, no!


This is precisely the problem!

In dot plots, each
dot can be one cell, two cells, three cells, or a thousand cells. You can
never
know which.



I think what he probably meant was that a dot plot does allow you to
spot
numbers of occurrences below your lowest threshold value for
contouring,
including single occurrences; it is true that if there isn't a dot at
a
point on a dot plot there weren't any cells observed

with the corresponding data values.



I agree with Alice that the precise way of contouring can affect how the more frequent populations appear.

This is why the choice of contouring algorithms is so important!

One of the most robust (in that it is objective, not allowing
"user-defined" contouring levels, etc.) is probability contouring. This method
of contouring has been adopted by SAS Institute for use in their bivariate
displays--in addition, it is offered by several FACS data analysis packages.


>This method of contouring generates displays that are indepedent of the number of events collected --

something that no other display can do.


By "probability contouring" do you mean normalization, so the contour
lines
represent percentile values rather than absolute numbers of cells?


This is a very sensible method of displaying things, and facilitates
comparison of samples of unequal sizes.

In order to deal with rare events, however, you
still have to have dots or their equivalent added to the contour plot.


Thus, using Dot plots or color dot plots or user-defined thresholding,
I can make a variety of
>conclusions about the same sample depending solely on how many events I choose
>to collect (or display)!

>Jim Houston is 100% correct that the precise method of data display is critical.

>I urge reviewers and editors to demand that this information be included
in all
>FACS data displays.


Note, for example, that bivariate chromosome contour plots are
generally made with higher thresholds than plots of immunofluorescence...

and
it wouldn't be a bad idea to include a scale or to indicate which contour
lines represent which percentiles.


Once again, this brings us to the fundamental point of data display: to convey information accurately to the reader.

I highly recommend a book by Edward Tufte, "The Visual Display of Quantitative Information," about this topic (especially to programmers developing analysis packages).

This fabulous book

>shows how misleading different styles of graphs can be, and discusses some of
>the underlying principles of data display--principles largely ignored by
>developers of FACS data analysis programs.
>There was some discussion about art vs. science. Do not mistake artistry for
>disinformation! Of course there is art in science, and in the presentation of
>scientific data.


If not, we would only see tables of numbers that would be
>incomprehensible-- we are, after all, only human.



Tufte actually has three books out; each is a work of art as well as a
work of science.

In the Chapter on Data Analysis in the 3rd Edition of
Practical Flow Cytometry, I suggest that single parameter distributions be
represented using Tufte's minimalist version of the "Box and Whiskers" plot, which
shows the position of the median, 25th and 75th, and 5th and 95th percentile
values (or, alternatively, the full range of the data instead of 5th
and 95th %).

This could readly be extended to a two-dimensional version,
but
might be better represented by different colored (or differently
shaded)
areas than by contour lines.


-Howard

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RE: data display

From: David L. Haviland, Ph.D. (dhavi...@imm2.imm.uth.tmc.edu)
Date: Fri Oct 03 1997 - 15:31:37 EST

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Greetings all:

I have learned a great deal and have enjoyed the discussion concerning
data display, dot vs contour, etc.

To settle a curiosity that I had,

I privately asked M_Roederer this question.

"We know each plot (contour/dot/density) has its benefits and
pitfalls.
However, does anyone know of an instance where a conclusion or part
thereof
has been in serious doubt because a conclusion was drawn off a dot
plot,
when a contour plot would have suggested a different conclusion?"

Frankly, I expected "no" as the respone but was suprised when Mario
stated
there had been a few instances. So my question to the group is

does anyone have such a reference(s) of a flow cytometric "opps"? Were manuscript retractions involved?

Many thanks in advance,
David

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RE: data display

From: Donnenberg, Albert (donnenber...@MSX.UPMC.EDU)
Date: Tue Sep 30 1997 - 15:41:23 EST
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Dot plot or probability plots, what do you do when compensation forces
a significant number of events into the first channel? Dot plots don't
get any blacker than black, some contour algorithms truncate the first
channel of data!

In the 4-color world it is sometimes impossible to keep events out of
the gutter when PMTs are balanced and compensation is optimized.
Albert D. Donnenberg, Ph.D.



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RE: data display

From: Gib Otten (G...@cellgenesys.com)
Date: Tue Sep 30 1997 - 19:32:44 EST

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Somtheing that's been overlooked in this discussion.

While a dot plot by itself may be misleading because of the dot density problem, most
investigators identify regions of interest and indicate the percent cells in those regions. The same should be done, and probably is being done, with contour plots.


Gib Otten, Ph.D.
Senior Scientist, Gene Therapy Applications
Cell Genesys, Inc.
342 Lakeside Drive
Foster City, CA 94404
Tel: (415) 425-4515
email: g...@mail.cellgenesys.com
> ----------
> From: Alice.L.Gi...@dartmouth.edu
> Sent: Monday, September 29, 1997 9:48 AM
> To: Cytometry Mailing List
> Subject: re: data display

> I know that dot plots can be misleading for all the reasons that

Mario Roederer describes ---

BUT I also know that, by choice of the contouring
algorithm, you can make a contour plot look any way you want: shoulders on peaks

can be emphasized or can be made to disappear, double peaks can be made to look like single peaks, etc etc etc.. These problems are not solved by showing the dots that are below the contouring threshold, as they relate to the levels of coutours above the threshold.

Dot plots can be misleading, but contour plots are a can of worms.

OK Mario (and anyone else) -- looking forward to your response!

> Alice

> Alice L. Givan
> Englert Cell Analysis Laboratory
> Dartmouth Medical School
> Lebanon, New Hampshire
> NH 03756 USA
> tel 603-650-7661
> fax 603-650-6130
> e-mail gi...@dartmouth.edu


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[Unregistered]

From: David.C.McFarl...@gsk.com

Sent: Wednesday, July 19, 2006 9:06 AM
To: Cytometry Mailing List

Subject: Re: FloJo and DiVa file IDs

Ray,

I had this discussion with BD a while ago. This is something that
blindsided me.

We actually have an SOP that we follow for tube IDs
such that they are all unique. Basically, we use accession numbers
of
the format FCM00001 for experiment names
and then the tube IDs would be FCM00001_001, etc.

In the older versions of Diva, this was the "FCS" file name that showed up in the folder when experiments were exported.


However, those weren't "real" FCS files.

Evidently,

people were trying to import these bogus FCS files

back into Diva,

or

elsewhere,

and it was creating errors.

In their infinite wisdom,

BD's fix

was

to change the names of these files to something unrecognizable to the users so that they wouldn't be tempted to do that.

They have a system of generating their own pseudo-unique IDs in this manner.

I say pseudo-unique, because it isn't clear to me where/how the
numbering starts if, for instance, you have several instruments
running Diva in the same lab, or if upgrade to a new version by doing
a clean install.

Or perhaps the machine crashes.

Can the numbering be set to pick up where if left off?

I think what they really mean is unique to that particular incarnation of the database on that local machine which isn't unique enough in my mind.

This really screwed up my system.

So asked the same question you just did, "Is there a way
to retain my Tube IDs

and/or

can I configure Diva to use the same unique filenaming convention that I do".

Of course, the answer was no.

Even worse,

even if you export as FCS files, although recognizable, the files aren't exactly the same as the entered ID.

The specimen name

and

an underscore are added before the tube ID

and

".fcs" is added to the end.

This is more aggravation for me to deal with in regard to data management in a GLP setting.

It's too bad. We like exporting entire experiments for archiving purposes,

but

it is easier to reconcile file name/tube ID discrepancies when exported as
FCS, so we will probably have to make a change.

At present we've just been adding a file note to the archive documents and explaining that the original naming scheme will reappear when imported back into
Diva.

This isn't entirely true when FCS files are imported into Diva
since you have to import them into an open experiment, which could be
named differently than the exported FCS files.



Again this is an issue for me in a GLP validated instrument setting and would warrant more workarounds or further explanation as to why reimported raw data
doesn't look "exactly" as it did when collected.

Let me know if you here anything different from what I've told you as
the situation seems to change frequently and this may no be the most
current info.

Regards,

Dave

David McFarland
Principal Scientist
GlaxoSmithKline


"Ray Hester" <rhes...@jaguar1.usouthal.edu>
17-Jul-2006 16:16

To "Cytometry Mailing List"
<cytome...@flowcyt.cyto.purdue.edu>
cc

Subject FloJo and DiVa file IDs

Can someone tell us how to export DiVa files as 'Experiments', not as
'FCS files', so that the individual tubes within the 'Experiment'
retain
their specific identities, e.g., 'T4/8', rather than some numerical
value, e.g., '2437'. When we export DiVa data as 'FCS files', the
specific tube ID is retained.

We have a FloJo 7.1 USB dongle

and it's being used primarily on PCs.

Thanks.

Ray Hester
Univ. of South Alabama


I must say that it came as a complete surprise to me that

BD

has not notified all Aria-containing laboratories about this bug

(I was visiting some other laboratories last week, and this news came as a
complete surprise to them!

The laboratories were relieved to find out that the recent sorting failures
were not their fault).

Normally, just in the last two weeks, well after the problem was identified.

Henc BD
has been responsive to problems,

but

in this case they have completely dropped the ball.





Diva 5.0 Software on Aria

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From: Mario Roederer roede...@drmr.com

Date: Tue Oct 10 2006 - 08:37:47 EDT

Dear Aria Users:

We would like to alert you to a serious bug in the Diva 5.0 software
that applies only to sorting on the Aria.

There is a significant

bug

that causes misapplication of gates, such that your sorting is completely incorrect.

We notified BD about this

bug

on on September15,

and

they were able to reproduce it before the end of September.

The problem occurs on any sorting that requires new gates; sorting
using old templates seems to work in at least some cases.

We have taken the solution to go back to the previous version of

Diva software (4.1.2) until BD can fix the issue.

Note that we have seen no issues with Diva 5.0 software on our LSR II analyzers,

and

continue to use 5.0 on the LSR II.

I must say that it came as a complete surprise to me that BD has not
notified all Aria-containing laboratories about this bug

(I was visiting some other laboratories last week, and this news came as a
complete surprise to them! The laboratories were relieved to find
out that the recent sorting failures were not their fault).

Normally, just in the last two weeks, well after the problem was identified. HencBD
has been responsive to problems,

but

in this case they have completely dropped the ball.

It cost us two important experiments to discover the bug;

in visiting other laboratories, I discovered those laboratories lost several important experiments e,

I felt it necessary to let everyone know before more valuable
experiments are lost.

mr

Received on Tue Oct 10 10:58:00 2006
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[Unregistered]

From: Novo, David [email protected]

Date: Fri Aug 17 2007 - 18:52:53 EDT


Hello Kate,


The FC 500 data files are indeed a complicated beast that give you a
lot of information.


There are indeed two separate data sets inside the one
FC500 file.

This is allowed by the FCS standard.


The first data set is a FCS 2.0 file.

quite a typical FCS 2.0 file, stored in 10 bit with the data already saved compensated and This is converted to log (if necessary).

This is quite clever to put it there because any analysis software that can read FCS 2.0 files should be able to read this file, and not even know that there was a second data set hiding afterwards.

I am not sure why the BD CBA software

would not be able to handle this,

maybe there is a hidden setting or something that allows this.


The second data set is a high resolution data set that is in FCS 3.0
format. This contains all the high resolution linear data, that is stored uncompensated, and is ideal for performing software compensation.


I guess our web site is either too high, or too low,

and

fell out of the range of your previous search :-)

FCS Express can read both the FCS

2.0 and FCS 3.0 portion of the FCS 500 data files. You can even export
them as individual files if you like.

When loading the files you can either default to load the FCS 2.0 or FCS 3.0 portion, or be prompted which to choose.



I also know that WinList can read both portions of the FC500 file.

I am not familiar withFlowJo's capabilities in this regard.

If you contact them,

I am sure they will be glad to answer this point.


-Dave


-----------------------------------


David Novo


President


De Novo Software


[email protected]

[Unregistered]

Advertising

From: Mario Roederer ([email protected])

Date: Fri Nov 01 2002 - 16:46:47 EST


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________________________________________

I agree that there have been far too many commercial-oriented emails
on the list.


I appreciate the efforts of most manufacturers to
withhold from advertising on this list

(plus, of course, the efforts of Steve & Paul to filter those out).

The list, which is an outstanding forum for exchange of information,
has been occasionally used to identify substantially new products

which can significantly impact on how we do experiments.

I feel that the Molecular Probes email of 10/30

clearly does not fall into this category;

the new product advertised was no more than a slight
modification of the existing one.

Such an email should be directed solely to the person requesting information;

if that person then collates responses and puts it back on the list then so be it.

But

for manufacturers to directly respond in this way is

counter-productive to the goals of this list.


I would like to propose a 6-month moratorium on all emails that are
no more than advertisements.

Note that I write "would like to"--because I'm not sure that this is possible.

I don't want to put any additional on us on Steve or Paul to filter out the borderline emails.


While these may be easy to identify when they come from
manufacturers,
it could just as well be considered blatant advertising when they come from a user.


Therefore, perhaps we can see if the commercial participants on this
list could exercise self-restraint rather than requesting a formal
censorship of advertising emails.


Thus, if you are a manufacturer, and you are responding to somebody's
request for information, do so privately to that person ONLY.

It is up to the person requesting information to decide whether or not the
information received in response to the query warrants a summary on
the list.


If you are not a manufacturer, and are responding to somebody's
request for specific information, please consider whether your
response
(that identifies a specific product or manufacturer)

is of general enough interest to warrant the list.

If it does not, then simply send it privately to the person who requested information, and let them decide whether to post the summary of responses.


In general,

I urge people to err on the side of caution and send
their information only to the person who requested it.

Realize that
if several people want the same information, they can always request
it from the original poster!

I have posted queries to the list;

people have sent me emails asking me to forward to them the
responses, whichI did.


This process can significantly cut down on emails that might be
viewed as too commercial.


mr
________________________________________
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[Unregistered]

From: Ray Hicks (rh...@cus.cam.ac.uk)

> Date: Fri Jul 04 1997 - 04:15:27 EST

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> ---------------------------------------------------------------------------­­-----
> Hi Vincent,

> I'm developing an analysis program, based on FCS Assistant, called > FCSPress.

So far it's quite rudimentary and only allows you to plot
histograms.

It doesn't do gating or analysis yet.

And

there is little support for the edit menu functions (cut, copy, paste, undo).

Unlike most cytometry programs available on the mac,

it allows you to open a data file and optionally manipulate its contents,

graphs are plotted from the active data window into graph windows,
which are wysiwyg for printing and saving as PICT )(sort of like cricket graph).

You can put as many graphs from as many files as you like into a graph window, and you can constrain their plotting to an optional grid of variable size.

You can annotate the graphs manually in variable fonts and styles, as well as semi-automatically from text held in the source file.

I've got quite a way to go before the program is finished, but it seems to be quite stable so far.

If you'd like a copy let me know and I'll e- mail it to you (or anyone else who'd like a copy).

Ray

At 10:29 am +0000 3/7/97,

Vincent Falco wrote:

I have BD Vantage generated list files copied to a PC via FACSNET.

I have an investigator who wishes to

generate multiple histograms in gallery format.

Which software product can do this the easiest(least steep learning curve)?

Thanks.

Ray Hicks


__________________________________________________ ______________________
> |University of Cambridge |Tel 01223
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> |Department of Medicine |Fax 01223
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> |Level 5, Addenbrookes Hospital |e-mail
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[Unregistered]

FCSPress Training Sessions?

From: Ray Hicks ([email protected])

Date: Thu Oct 11 2001 - 19:42:36 EST
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--------------------------------------------------------------------------------
Most of you will findFCSPress so easy to use that you won't need
training sessions.

You can download a copy to try it out for yourselves at

http://www.fcspress.com/html/FCSPArea.html

(Who's computer is this?)

where you'll also find documentation and details of a special offer.
If you find you DO need some assistance using FCSPress,
I'm planning on staying in Cambridge for quite some time, and I'd be happy to answerany ofyour queries via e-mail at

[email protected]
Cheers,
Ray
--
e-mail mailto:[email protected]
Web http://www.FCSPress.com
(Who's computer is this?)
Tel +44 797 453 8647
+44 1223 871081
Fax +44 870 7408595
--------------------------------------------------------------------------------
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FlowJo Training Sessions (Midwest)

From: Jennifer Wilshire ([email protected])

Date: Thu Oct 11 2001 - 06:20:11 EST
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Next in thread: Ray Hicks: "FCSPress Training Sessions?"

Rand locations are listed below. Thanks, Jennifer Wilshire FLOWJO
SEMINARS ______________________________________ Monday, October 15th
University Hospitals Clinical Sciences Center 600 Highland Ave,

Madison WI Room K4/418 10:00 AM reply: Ray Hicks: "FCSPress Training Sessions?"

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I will be in Wisconsin, Minnesota, and Iowa offering FlowJo seminars
and training sessions next week. All of the hosting sites have

graciously opened the sessions to members of the Cytometry list.


Times

______________________________________
Tuesday, October 16th University of Minnesota Cancer Center 425 E.
River Road Room 450 9:00 AM ¬ Introduction to FlowJo 10:30 AM ¬
Advanced Topics _______________________________________ Tuesday,
October 16th University of Minnesota, St. Paul Building AS/VM Room
295M 2:00 PM _______________________________________ Friday, October
19th University of Iowa South East 310 General Hospital Bean
Conference Room 10:00 AM _______________________________________ If
you are interested in hosting a FlowJo seminar, please let me know.
Upcoming trips are in the planning stages for: -NJ, PA, DC -Florida.
-
Boston (home) Jennifer Wilshire, Ph.D. Application Scientist
[email protected] Tree Star, Inc. FlowJo Home
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[Unregistered]

From: J. Paul Robinson <j..._at_flowcyt.cyto.purdue.edu>

Date: Fri Dec 28 2007 - 13:43:46 EST


> Beware, the end is nigh!

No, not an apocalyptic prediction - but 2007 is definitely coming to an end.

Not before time I would say - it s been a busy year.

But

I have some strong words to end the year

and

I am going to say them!!

Of course you don t have to read them!


Cytometry is now 40 years old and it s been sort of decaying a bit.


What do I mean?

I am amazed at how conservative

and

frankly boring the field has become.

Why?

It s time to move to the 21st century folks.

I'm getting older and frankly,

its time to kick some butt

as my younger colleagues often say.

We talk so much like it is the same old cytometry
it has always been.


Wake up people - times are changing

- look at all these new small companies


trying to stick their noses in "our" field!


True we need to do the core work and do it well,

but lets not forget that fundamental tools of cell analysis are changing

and

if we don't keep ourselves up-to-date and educated on what's happening....

before we know it,

a new field will emerge and we will be like the old electron microscopists who are still wondering what happened ......


I know most of us work in the field

and

like what we do,

but

I think its time to open up a little

and

try to do some serious integration of our field.

It s not happening very effectively on the most part

I would say.


Cytometry is about integration of the tools of the field into the vast reaches of biological problems that we can contribute to solving.

Cytometry is about advancement of the field,
that means always looking ahead.

ISAC will soon be the International Society for Advancement

of Cytometry a 21st century Society not a 20th century Society.

Cytometry is not flow cytometry!!

Let s not kid ourselves about this folks.

Cytometry is about measuring cells -

however you do it -

and

flow cytometry is just one component of many.

I understand that it may be the only tool some of you use -

I don t want to take away from that

or

de- emphasize its value or importance.

But,

we so often hear people talk about our field

only in context of just flow cytometry.

Recently, when we polled the ISAC community on

changing

our name from "analytical Cytology"

to Advancement of Cytometry"

we received comments like

"hey I don t do flow cytometry,

so

why are you reducing the breadth of the field?"

Ouch - they think "cytometry" means "flow cytometry".

We have a long way to go before


we convince the community that we cover all

aspects of cytometry.


And

let s also remember the growing membership

in

India and China

(that s half the worlds population right there)

it s high time we paid much more attention to these countries

as a field Awtar Krishan can t be the only person to drive cytometry training

and

education for 1.2 billion Indians can he?

Well he has been up to now.

Who is taking on the mantle of training and education of cytometry in
China?

So here's the scoop.


That's one of the reasons

why the Purdue Web portal is going to change.

We tried to make the change this past year,

but

there were too many other things happening here to achieve it.

But

come middle of 2008,

I am resolved that you will see a huge difference in the Purdue site.

It s been the default cytometry communication portal nowfor many, many years.

We have focused on good clean fun with cytometry

quality, timely, simple - no spam.

Many people like that actually.
> The
> portal is almost overwhelming for us 22,000 daily page requests with
> over 2 Gigs daily download. In 2007 alone, downloads of 208,000
> powerpoint files, 233,000 PDF files, 8800 movies, 38,000 word
> document
> files. The education pages and the Cytometry Discussion Archive are
> the
> most hit for sure. Over 125,000 distinct files from our portal were
> accessed in 2007.

> But all good things must come to an end.

Come July 2008,

the usual Purdue web portal may well be no more.

It will be replaced with something entirely new.


Hopefully most will find it more useful

and


relevant - some will not like it.

Maybe we will be able to make everyone happy

....ha!..C'est la vie.


Some of you will be beta testers and advisers I hope.


So my best wishes to all in the cytometry field for 2008.

Regarding the past year on the discussion list, its been lively, with an average of 7 messages per day with 754 different individuals submitting at least one message. 139 messages had at least 6 responses. There were 1205 unique subject lines. Subscribers came from 64 top level domains. The usual bunch of suspects answered lots of messages and Marty Bigos seems to have too much time as he answered the most (thanks Marty!!).

Tragically, the second most prolific responder was Randy Fisher who passed away on December 5. Randy's responses were always short, to the point and accurate. It hurts to lose one of our own, particularly when it's one
of our most active members.

But that s the point isn't it. For many
years to come, we have the value of Randy's hundreds of suggestions over the years archived for the many new people who enter our field. Many of you probably never actually met Randy - but I bet most of you feel you knew him. One of the mysteries of the web I suppose. Our condolences to Randy's family - perhaps they didn't know how many people knew Randy "electronically" - but we all did.
You know we are a small field when
it comes to the big world of science so when we lose one person, the entire field morns.

To end 2007, let me make a big plug for a program we began at the 2006 ISAC congress.

Gary Durack from iCyt and myself started a small not- for-profit charity called

"Cytometry for Life" in response to Stephen
Lewis' compelling plea for some low cost CD4 devices. Our field
has done a lot of talking about this,

but

only a few people have really tried to do anything practical.

Well, folks we have all been doing cytometry for a very long time - it's time to do something.
"Cytometry for life"

(http://www.cytometryforlife.org)

is working hard. We have made tremendous progress in just one year. It would be great if you all decided to jump on board and play a small part.
You can give money, advice, moral support, talk to your politicians, community healthcare, charities,

whatever.

but get involved as be recognized as the cytometry community to solve this problem of bringing low cost, portable devices to the 65% or more of African's who don t live in the cities and towns where current CD4 technologies are available. Our goal is to work in areas not being served by current technologies.

We have heard these calls before, but folks we have to deal with this problem - it's your problem

if you call yourself a "cytometry" person.

Email me if you can help -

consider donating to the program, let's make it work. By the end of 2008,

I want to be telling you that the program is getting to
people who need this desperately. Help us achieve that for 2008.

I hope many of you got hold of a copy of our new double DVD set Cytometry 60 years of Innovation
if not ask your local rep from virtually any company in our field.

It might give you a good sense of
how strong the foundation in our field really is.
I will see many of you at the 2008 congress in Budapest. I know some of you think its going to be expensive so I took several hours myself and created a webpage for
the cheap ones out there so you have no excuses not to go...

(Cheap european Flights).

It's been a privilege to serve for the past 19 months as President of ISAC.

I will gladly pass that hat to Bob Murphy in May. ISAC is alive and well - membership is growing daily.

I would not be surprised to see us top 2000 by the end of the Congress in May.

I know that about 60% of the members of this list are NOT ISAC members.

Perhaps you should consider joining

the

Society that keeps many of you in business?

Welcome to the ISAC! - Mambo

> My best wishes for you all in 2008 from Purdue Paul
> --
> J. Paul Robinson
> SVM Professor of Cytomics
> Professor of Immunopharmacology & Biomedical Engineering
> Director, Purdue University Cytometry Laboratories
> President, International Society for Analytical Cytology

[Unregistered]

From: Ray Hicks
Sent: Wednesday, October 17, 2001 8:42 PM
To: Cytometry Mailing List

Subject: Re: Bad Flow Data & reviewing -- What can we do?


Many good points Mario,
but

I'm going to take you back a few years to our
discussion on dot plot versus contour, and how misleading contours are.
I'd
reverse your logic in " remember that contour plots are also
histograms (2D histograms),

and

they have no numbers on the "Z" axis
corresponding to event frequency. Why should univariate histograms have
them?", and suggest that contour plots need even more annotation.

I'm sorely tempted to attach a few figures to this e-mail,

but

I've restrained myself, and made them available at:http://www.fcspress.com/seeWhatIMean.gif (Who's computer is this?)

and

http://www.fcspress.com/512AlongTheAxis.gif (Who's computer is this?)

The first <http://www.fcspress.com/seeWhatIMean.gif (Who's computer is this?) > shows how strikingly different contour plots of the same data can be

(the data is from the FlowJo tutorial set, the figures are made in FlowJo 3.2 and FCSPress 1.3).

The top left dotplot is from FlowJo,

and

shows the crowding you object to, the upper central plot is FlowJo's default contour plot of SSCvFSC with ten thousand
cells, the upper right plot is a plot of 1600 cells gated from the same
file
- doesn't look like fewer cells does it?
The lower left plot is a log 50% contour plot of the data in the top left
and top centre plots, what is one to make of those contours based on four
cells that jump out in the lower left?

The lower central plot is a dot plot from FCSPress, plotting data at 512 points along the axis

(the data has a range of 512 "channels"),

FCSPress has dithered the plot to represent how it would

(and does) print on a printer which isn't limited to screen resolution

(using the "clarify option),

you'll notice that using higher resolution avoids much of the coalescing to a black blob that you object to in dot plots

(the second figure, <http://www.fcspress.com/512AlongTheAxis.gif (Who's computer is this?) >,
.
shows this graph at full size with no dithering)
The lower right plot shows a density plot from FlowJo,
the smoothing belies the sparsity of the data.

What's an expert to do when presented with this kind of thing?

Would labelling the upper left and lower left plots as having the same number of cells be enough to make you see them as representing the exact same data
set?

The dot plot of 1600 cells (not shown for brevity) clearly has fewer
cells than that of 10000, and does a better job of warning the viewer,
expert or not, of how confident they should be in making conclusions based
on the plot than numbering the events on the two contour plots (upper left
and upper right).
Oh, alright then, I've put a further figure up with two dot plots and two
contour plots with paired numbers of events at:

http://www.fcspress.com/nowDoYouSee.gif (Who's computer is this?)

The other issue I take is;

how is the collective going to select the
experts?Surely the people who are publishing this stuff ARE people

"with a modicum of experience in flow".

Putting the responsibility on editorial
boards is probably going to end up in a status quo.

How about pressuring your lab-fellows

to sling the FACS aspect of papers, that

they're reviewing anyway,

in your direction?


Ray

ps

as an aside,

there's something freaky happening on the axes of these graphs -

they're 512 channel data,

but

the linear FSC axis runs out
just past 200,

and one of the events exceeds the maximum for side scatter
(ie the one that juumps above the red line in the left hand plots -

has this beenfixed in later versions of

FlowJo?

Would this be

something an expert could

criticise/reject

a paper for?

[Unregistered]

From: Randy T. Fischer ([email protected])
Date: Thu Jul 17 1997 - 03:25:15 EST
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[ attachment ]
________________________________________
I agree with both

Marty and Gunter

in the very important issue

ofstandardizing data formatting.

I would point out that

lobbying ISAC

is only, however, part of the answer.

Regardless of what ISAC may choose to recommend,
it is still up to the

manufacturers to implement what they
want to do,

and

if they do not agree
with ISAC,

then too bad for ISAC

and

the flow community.

A potentially more powerful

force for change

might be the

FDA,

which regulates machines

used in CLINICAL settings.

If the

FDA

could be persuaded


to

requireall

CLINICAL data be universally

both
accessible and readable,
then the manufacurers

would be forced


to upgrade machines

and

software or lose


the
LUCRATIVE CLINICAL market.

This would make anlyzing data from different sources easier, and could facilitate the exchange

ofcrucial clinical results

from various trials where multiple sites

and

machines are in use.

So how does this get done?

Gunter

(and Paul's agreeing response)

are correct this needs to be revisited at Asilomar,

with perhaps an additional idea.

Any concrete standardization protocol,

FCS3.0

or

whatever

it ends up being designated,

should be then presented to any

and

all regulatory agencies

by

ISAC

to ensure

no

individual manufacturer decides

FCS3.0 in their format is acceptable,
even

if it is not universally readable.

Randy T. Fischer
NIA/NIH
GRC
Baltimore, MD 21224
[email protected]




NOW

PAUL AND THE CREW RUN ISAC

THE DEVELOPERS FORM THE MAIL HAVE TAKEN CONTROL!

They are only

THE

FDA

AWAY FROM TOTAL CONTROL OF THE MARKET!

look at all these small companies
trying to stick there nose in our field!

THE COALITION FOR CHANGE!

[Unregistered]

GOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOOGLE
PURDUE CYTOMETRY MAIL LIST


READ HOW THEY GREW AND TOOK CONTROL OF ISAC CONGRESS!

FILTERING OUT THE LIST!

DEVELOPERS OF SOFTWARE

MACHINES

CODE

THERE MUST BE CHANGE