Source of Richardson test slide

[JOEL B. SHEFFIELD]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Tina,

Well put.  Those rules do make a lot of sense.  However, as
you also mention, acquisition software now plays a significant
role in image generation.  The simple software that we have
now will compensate for white balance and subtract a
background image automatically.  There are also sharpening
kernels and binning effects to consider.  

On the other hand, I remember from my em days that lots of
people developed plates "by eye", and used cropping and
dodging techniques to emphasize certain areas.  

How do we deal with this?

Joel
---- Original message ----
>Date: Sun, 22 Jun 2008 14:22:55 -1000
>From: Tina Carvalho <[hidden email]>  
>Subject: Re: An alarming amount of image manipulation  
>To: [hidden email]
>
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>So this might be a good time to remind everyone that the
Microscopy
>Society of America's Education Committee has a subcommittee
on the Ethics
>of Digital Imaging. Several years ago we tackled this problem
>(intellectually) and came up with amazingly straightforward
guidelines:
>
>The only thing you can do to a digital image without
reporting it as
>manipulation is changing contrast and brightness, and
levels/gamma/curves
>over the entire image. *Anything* else needs to be reported
as an image
>manipulation in the Materials and Methods. In addition, the
original
>(unmanipulated) image must be stored as an uncompressed TIFF
on archival
>media. It should be available to anyone who requests
inspecting the
>original.
>
>We hoped that this would alert (un-educated) researchers to
the fact
>that Photoshoping an image means changing the data, and they
would
>educate themselves about image manipulation programs and how
they work and
>what they are actually dong to an image.
>
>Now, of course there are all kinds of things you can do before
>snapping/saving an image, especially with confocal and
fluorescence
>acquisition software, and we have to rely on people's
internal morality
>meter... But even there researchers should be able to
adequately report
>and describe their software and parameters. If they can't,
it's probably
>more a matter of not understanding what they are doing, a
problem that
>seems to be rampant these days. Of course, there is also the
question of
>deliberate fraud, but I suspect that's truly less that is
insinuated in
>the article and more a question of un-education.
>
>Aloha, Tina
>
>> I was a bit surprised at the statistics cited in this article:
>>
>> http://chronicle.com/free/2008/05/3028n.htm
>>
>> Does this mean that all journals will start hiring image
manipulation  

>> detectives someday? Could be an interesting career.
>>
>>
>> John Oreopoulos, BSc,
>> PhD Candidate
>> University of Toronto
>> Institute For Biomaterials and Biomedical Engineering
>> Centre For Studies in Molecular Imaging
>>
>> Tel: W:416-946-5022
>>
>
>****************************************************************************
>* Tina (Weatherby) Carvalho               *

[hidden email]           *
>* Biological Electron Microscope Facility * [phone]  
              *
>* University of Hawaii at Manoa           *
http://www.pbrc.hawaii.edu/bemf* 
>****************************************************************************

Joel B. Sheffield
Professor, Department of Biology
Temple University
Philadelphia, PA 19122
[hidden email]
http://astro.temple.edu/~jbs

[Russell Spear]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi
Some of this is the same as Kodabromide F1 - F5 for contrast
manipulation.  How many of you didn't keep undiluted D19 to rub on an
area of a print, that wasn't coming up fast enough in the developer?  Or
done unsharp-masking using photo-plates.  The same goes with software,
you can pretty much do what you want if you are good at it.  My policy
in our imaging group; is we watermark all unapproved images, when we
release them to investigators, and will not release images for
publication, until we see the written the materials and methods
description of image manipulation.


Russell N. Spear
Sr. Research Specialist
Dept. of Plant Pathology
Univ. of Wisconsin
1630 Linden Dr.
Madison WI 53706

voice 608.263.2093
fax     [phone]

[Dr. Mark A. DeCoster]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

I have also brought up these ethical concerns in our Doctoral Seminar course
for Biomedical Engineering here.  I remember one of the examples I used of a
"scientist's nightmare" which included false results generated by shared
software ("homemade" and in this case, for crystallography analysis)-- but
this brings up the point also, that with all of the different image analysis
software being used, it should also be helpful for more journals (and
reviewers) to insist that the version of the image software used be stated:
"Photoshop version 6, Image J ver.1h,  Image ProPlus ver. 6.1",  etc.  

Since these are the tools being used, we need to know.

Best,

Mark  

Dr. Mark A. DeCoster
Associate Professor
Biomedical Engineering
Louisiana Tech University

http://www2.latech.edu/~decoster/


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On
Behalf Of Shalin Mehta
Sent: Sunday, June 22, 2008 10:12 AM
To: [hidden email]
Subject: Re: An alarming amount of image manipulation

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

We realized it is a big concern these days when we took a course on
ethics in research (mandatory for anyone doing bioengineering or life
sciences here). We came across quite interesting cases of high profile
fraud.

We had to do a presentation and we presented something on image
manipulation. While thinking about it I stumbled upon an interesting
way of quickly checking fake data. It is to 'scan the histogram' with
'narrow look up table', i.e. set the contrast to maximum and view the
image as you change the brightness.

The idea being that when someone has merged different images to appear
one or copied e.g. bands - scanning the histogram will show clear
signs. In the case of merging,one can see an edge where merge has been
done and in the case of copy one can see that two regions of the image
change identically as we scan the histogram. With that, I found that I
could actually detect discrepancies in example fake data given in
Rossner's editorial (What is in a picture? by Rossner and Yamada).

I tend to think that these problems will subside as ideas like open
source, reproducible research
(http://sepwww.stanford.edu/research/redoc/), and giving credit based
on 'first appearance anywhere and not just journal' will permeate
gradually - they have already done so in computer science where it is
very easy to allow someone to reproduce your work simply by uploading
files.

cheers
shalin



On Sun, Jun 22, 2008 at 10:43 PM, Guy Cox <[hidden email]> wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Interesting - especially the statement that images on film
> cannot be doctored. I guess it's irrelevant nowadays, but that
> is so far from the truth.
>
> One funny story - which I've told before but some time ago.
> Many years ago I (with colleagues) published a paper in a
> well-known journal which included a photo of some gels -
> taken by the departmental photographer.  One of the gels had
> cracked when it was taken out of the tube, so there was a
> dark hairline across it in the photo.  In the published paper
> that line had disappeared.  You can hardly accuse me, or my
> two co-authors, of fraud since this was done by the journal's
> art department without any reference to us!
>
>                                                       Guy
>
>
> Optical Imaging Techniques in Cell Biology
> by Guy Cox    CRC Press / Taylor & Francis
>    http://www.guycox.com/optical.htm
> ______________________________________________
> Associate Professor Guy Cox, MA, DPhil(Oxon)
> Electron Microscope Unit, Madsen Building F09,
> University of Sydney, NSW 2006
> ______________________________________________
> Phone [phone]     Fax [phone]
> Mobile [phone]
> ______________________________________________
>     http://www.guycox.net
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On

Behalf Of John Oreopoulos

> Sent: Sunday, 22 June 2008 10:57 PM
> To: [hidden email]
> Subject: An alarming amount of image manipulation
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> I was a bit surprised at the statistics cited in this article:
>
> http://chronicle.com/free/2008/05/3028n.htm
>
> Does this mean that all journals will start hiring image manipulation

detectives someday? Could be an interesting career.
>
>
> John Oreopoulos, BSc,
> PhD Candidate
> University of Toronto
> Institute For Biomaterials and Biomedical Engineering Centre For Studies
in Molecular Imaging
>
> Tel: W:416-946-5022
>
> No virus found in this incoming message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008
9:27 AM
>
>
> No virus found in this outgoing message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008
9:27 AM
>
>



--
~~~~~~~~~~~~~~~~~~~~~~~~~
Shalin Mehta
mobile: [phone]
blog: shalin.wordpress.com
~~~~~~~~~~~~~~~~~~~~~~~~~~
Bioimaging Lab, Block-E3A, #7-10
Div of Bioengineering, NUS Singapore 117574
website: http://www.bioeng.nus.edu.sg/optbioimaging/colin/index.html

Liver Cancer Functional Genomics Lab, #6-05
National Cancer Centre, Singapore 169610
http://www.nccs.com.sg/researcher/02_04d.htm

[Guy Cox]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Two quite separate comments on this thread.

1. Often, in response to the referees, I have to change figures.
Maybe group them differently (which means changing letters) or
alter the labelling.  I suppose I should go back to the original
images and re-create the entire plate, but if the changes are
minor it's much easier to (for example) black out 'a' and paste
in 'b', or take out an arrow and put in a letter, etc.  All these
things would presumably show up as 'fraud' ....

2.  A point nobody has yet raised is deconvolution imaging
systems.  Clinicians love them, but they have no training in physics
(and, to be honest, not much in science).  None of them has the
first idea what the software is doing, just that it's giving them
the pictures they want.  How does anyone handle that?

                                                   Guy



Optical Imaging Techniques in Cell Biology
by Guy Cox    CRC Press / Taylor & Francis
    http://www.guycox.com/optical.htm
______________________________________________
Associate Professor Guy Cox, MA, DPhil(Oxon)
Electron Microscope Unit, Madsen Building F09,
University of Sydney, NSW 2006
______________________________________________
Phone [phone]     Fax [phone]
Mobile [phone]
______________________________________________
     http://www.guycox.net
-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Michael Cammer
Sent: Monday, 23 June 2008 2:30 AM
To: [hidden email]
Subject: Re: An alarming amount of image manipulation

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

In my experience, when somebody shows up with micrographs that have cells pasted in or background "distractions" stamp tooled out and I suggest maybe taking more pictures or doing the experiment again, they complain that this would cost more and delay them.  "The data are what they are and I'm just showing people and if I don't make it look good, then they won't publish it."  Sometimes junior lab members will go back to their PIs and say that I wasn't helpful; I become the problem.  I will not apologize for refusing to use the lasso tool around a band on a gel and S curve adjust it in Photoshop Curves or for refusing to paint out "an anomalous bad cell off in the corner of the image".  Yes, both these requests have come through our facility along with a bunch of other egregious ones.  Some scientists really feel justified because the competition does it.  "Hey, everybody does it," they say.  Although, more often (and a search will show that we've discussed this before), people simply don;t understand what they are doing.  They simply misuse the tools for making figures.  As with example of film, before you can begin to alter images in an intentional manner or make even reasonably good pictures of anything, you need to be at least knowledgeable of darkroom technique.  But with the new digital tools, any dummy can be a photographer and can cut and paste and manipulate in complicated ways.
-Michael

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Interesting - especially the statement that images on film cannot be
> doctored. I guess it's irrelevant nowadays, but that is so far from
> the truth.
>
> One funny story - which I've told before but some time ago.
> Many years ago I (with colleagues) published a paper in a well-known
> journal which included a photo of some gels - taken by the
> departmental photographer.  One of the gels had cracked when it was
> taken out of the tube, so there was a dark hairline across it in the
> photo.  In the published paper that line had disappeared.  You can
> hardly accuse me, or my two co-authors, of fraud since this was done
> by the journal's art department without any reference to us!
>
>                                                        Guy
>
>
> Optical Imaging Techniques in Cell Biology
> by Guy Cox    CRC Press / Taylor & Francis
>     http://www.guycox.com/optical.htm
> ______________________________________________
> Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope Unit,
> Madsen Building F09, University of Sydney, NSW 2006
> ______________________________________________
> Phone [phone]     Fax [phone]
> Mobile [phone]
> ______________________________________________
>      http://www.guycox.net
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]]
> On Behalf Of John Oreopoulos
> Sent: Sunday, 22 June 2008 10:57 PM
> To: [hidden email]
> Subject: An alarming amount of image manipulation
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> I was a bit surprised at the statistics cited in this article:
>
> http://chronicle.com/free/2008/05/3028n.htm
>
> Does this mean that all journals will start hiring image manipulation
> detectives someday? Could be an interesting career.
>
>
> John Oreopoulos, BSc,
> PhD Candidate
> University of Toronto
> Institute For Biomaterials and Biomedical Engineering Centre For
> Studies in Molecular Imaging
>
> Tel: W:416-946-5022
>
> No virus found in this incoming message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> 21/06/2008
> 9:27 AM
>
>
> No virus found in this outgoing message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> 21/06/2008
> 9:27 AM
>
>

_________________________________________
Michael Cammer   http://www.aecom.yu.edu/aif/

No virus found in this incoming message.
Checked by AVG.
Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008 9:27 AM
 

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Checked by AVG.
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[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> Well put.  Those rules do make a lot of sense.  However, as
> you also mention, acquisition software now plays a significant
> role in image generation.  The simple software that we have
> now will compensate for white balance and subtract a
> background image automatically.  There are also sharpening
> kernels and binning effects to consider.  

Right. It seems to be up to each individual to figure out how their
software works, what each algorithm is doing to their image/data before
acquisition, and what any post-processing steps are doing as well. This is
clearly too much self-education for most people, and I guess I would hope
that each software supplier would include a nice, big, fat booklet on how
this all works. How likely is that? Anyway, it was easy for us to come up
with guidelines and hope that it would make people think, but where do we
go from there? I had hoped to come up with guidelines for journal editors
and distribute them, but we (the two of us currently on the committee)
haven't had time to move on this.

Ideas and comments, and any participation welcome.

> On the other hand, I remember from my em days that lots of
> people developed plates "by eye", and used cropping and
> dodging techniques to emphasize certain areas.  

Back in the Really Old Days the development of negatives and the
production of photographic prints were detailed in the Materials and
Methods of a paper, right down to the concentration of developer, minutes,
etc., plus any dodging and burning. This eventually got shortened to just
negative and paper type and grade, and shortened more, until not much was
reported. Made for easier reading. I don't miss those days. BUT if we now
go back to more reporting of the details of image acquisition, it will
serve to promote reproducibility and, one hopes, make people think about
what they're doing so that they do not unintentially manipulate their
data.

The guideliens are pretty straightforward for black and white, but get
squirrely for color. For example, I think that if you can adjust Levels in
Photoshop, you can then go into levels and adjust each color level
independently. I love this, and it only makes me a little uneasy when I do
it... bring up the red, depress the green a bit... Hmmm, where to
stop? Presumably this is similar to changing the voltage on the PMTs, but
not really, but...

Again, comments appreciated. Do we need guidelines? How do we set
them? How do we get the info out to both editors and researchers? How do
we educate everyone, including the grad student who is just grabbing a few
images for a thesis and not making a living of it?

Aloha, Tina

>
> How do we deal with this?
>
> Joel
> ---- Original message ----
> >Date: Sun, 22 Jun 2008 14:22:55 -1000
> >From: Tina Carvalho <[hidden email]>  
> >Subject: Re: An alarming amount of image manipulation  
> >To: [hidden email]
> >
> >Search the CONFOCAL archive at
> >http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> >So this might be a good time to remind everyone that the
> Microscopy
> >Society of America's Education Committee has a subcommittee
> on the Ethics
> >of Digital Imaging. Several years ago we tackled this problem
> >(intellectually) and came up with amazingly straightforward
> guidelines:
> >
> >The only thing you can do to a digital image without
> reporting it as
> >manipulation is changing contrast and brightness, and
> levels/gamma/curves
> >over the entire image. *Anything* else needs to be reported
> as an image
> >manipulation in the Materials and Methods. In addition, the
> original
> >(unmanipulated) image must be stored as an uncompressed TIFF
> on archival
> >media. It should be available to anyone who requests
> inspecting the
> >original.
> >
> >We hoped that this would alert (un-educated) researchers to
> the fact
> >that Photoshoping an image means changing the data, and they
> would
> >educate themselves about image manipulation programs and how
> they work and
> >what they are actually dong to an image.
> >
> >Now, of course there are all kinds of things you can do before
> >snapping/saving an image, especially with confocal and
> fluorescence
> >acquisition software, and we have to rely on people's
> internal morality
> >meter... But even there researchers should be able to
> adequately report
> >and describe their software and parameters. If they can't,
> it's probably
> >more a matter of not understanding what they are doing, a
> problem that
> >seems to be rampant these days. Of course, there is also the
> question of
> >deliberate fraud, but I suspect that's truly less that is
> insinuated in
> >the article and more a question of un-education.
> >
> >Aloha, Tina
> >
> >> I was a bit surprised at the statistics cited in this article:
> >>
> >> http://chronicle.com/free/2008/05/3028n.htm
> >>
> >> Does this mean that all journals will start hiring image
> manipulation  
> >> detectives someday? Could be an interesting career.
> >>
> >>
> >> John Oreopoulos, BSc,
> >> PhD Candidate
> >> University of Toronto
> >> Institute For Biomaterials and Biomedical Engineering
> >> Centre For Studies in Molecular Imaging
> >>
> >> Tel: W:416-946-5022
> >>
> >
> >****************************************************************************
> >* Tina (Weatherby) Carvalho               *
> [hidden email]           *
> >* Biological Electron Microscope Facility * [phone]  
>               *
> >* University of Hawaii at Manoa           *
> http://www.pbrc.hawaii.edu/bemf* 
> >****************************************************************************
>
> Joel B. Sheffield
> Professor, Department of Biology
> Temple University
> Philadelphia, PA 19122
> [hidden email]
> http://astro.temple.edu/~jbs
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> 1. Often, in response to the referees, I have to change figures.
> Maybe group them differently (which means changing letters) or
> alter the labelling.  I suppose I should go back to the original
> images and re-create the entire plate, but if the changes are
> minor it's much easier to (for example) black out 'a' and paste
> in 'b', or take out an arrow and put in a letter, etc.  All these
> things would presumably show up as 'fraud' ....

Yup. Sigh. This is the education part, and I guide people through this a
lot. The trick is to learn to use Layers in Photoshop, and save every
iteration (disk space is cheap) so that you can go back and move the
images, letters, arrows, etc. on a figure plate. Once you do this one
time, you realize how valuable it is!

> 2.  A point nobody has yet raised is deconvolution imaging
> systems.  Clinicians love them, but they have no training in physics
> (and, to be honest, not much in science).  None of them has the
> first idea what the software is doing, just that it's giving them
> the pictures they want.  How does anyone handle that?

Education. Give them a cheat sheet that lays out how it works. Give them a
test. Whatever. I know it can be a pain, but researchers should (in an
ideal world) understand what it is that they are doing. I can go on a rant
about labeling "kits" where you follow directions, but never have any idea
what is actually happening. I can go on lots of rants...

Aloha, Tina

>
> Optical Imaging Techniques in Cell Biology
> by Guy Cox    CRC Press / Taylor & Francis
>     http://www.guycox.com/optical.htm
> ______________________________________________
> Associate Professor Guy Cox, MA, DPhil(Oxon)
> Electron Microscope Unit, Madsen Building F09,
> University of Sydney, NSW 2006
> ______________________________________________
> Phone [phone]     Fax [phone]
> Mobile [phone]
> ______________________________________________
>      http://www.guycox.net
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Michael Cammer
> Sent: Monday, 23 June 2008 2:30 AM
> To: [hidden email]
> Subject: Re: An alarming amount of image manipulation
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> In my experience, when somebody shows up with micrographs that have cells pasted in or background "distractions" stamp tooled out and I suggest maybe taking more pictures or doing the experiment again, they complain that this would cost more and delay them.  "The data are what they are and I'm just showing people and if I don't make it look good, then they won't publish it."  Sometimes junior lab members will go back to their PIs and say that I wasn't helpful; I become the problem.  I will not apologize for refusing to use the lasso tool around a band on a gel and S curve adjust it in Photoshop Curves or for refusing to paint out "an anomalous bad cell off in the corner of the image".  Yes, both these requests have come through our facility along with a bunch of other egregious ones.  Some scientists really feel justified because the competition does it.  "Hey, everybody does it," they say.  Although, more often (and a search will show that we've discussed this before), people simply don;t understand what they are doing.  They simply misuse the tools for making figures.  As with example of film, before you can begin to alter images in an intentional manner or make even reasonably good pictures of anything, you need to be at least knowledgeable of darkroom technique.  But with the new digital tools, any dummy can be a photographer and can cut and paste and manipulate in complicated ways.
> -Michael
>
> > Search the CONFOCAL archive at
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> > Interesting - especially the statement that images on film cannot be
> > doctored. I guess it's irrelevant nowadays, but that is so far from
> > the truth.
> >
> > One funny story - which I've told before but some time ago.
> > Many years ago I (with colleagues) published a paper in a well-known
> > journal which included a photo of some gels - taken by the
> > departmental photographer.  One of the gels had cracked when it was
> > taken out of the tube, so there was a dark hairline across it in the
> > photo.  In the published paper that line had disappeared.  You can
> > hardly accuse me, or my two co-authors, of fraud since this was done
> > by the journal's art department without any reference to us!
> >
> >                                                        Guy
> >
> >
> > Optical Imaging Techniques in Cell Biology
> > by Guy Cox    CRC Press / Taylor & Francis
> >     http://www.guycox.com/optical.htm
> > ______________________________________________
> > Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope Unit,
> > Madsen Building F09, University of Sydney, NSW 2006
> > ______________________________________________
> > Phone [phone]     Fax [phone]
> > Mobile [phone]
> > ______________________________________________
> >      http://www.guycox.net
> > -----Original Message-----
> > From: Confocal Microscopy List [mailto:[hidden email]]
> > On Behalf Of John Oreopoulos
> > Sent: Sunday, 22 June 2008 10:57 PM
> > To: [hidden email]
> > Subject: An alarming amount of image manipulation
> >
> > Search the CONFOCAL archive at
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> > I was a bit surprised at the statistics cited in this article:
> >
> > http://chronicle.com/free/2008/05/3028n.htm
> >
> > Does this mean that all journals will start hiring image manipulation
> > detectives someday? Could be an interesting career.
> >
> >
> > John Oreopoulos, BSc,
> > PhD Candidate
> > University of Toronto
> > Institute For Biomaterials and Biomedical Engineering Centre For
> > Studies in Molecular Imaging
> >
> > Tel: W:416-946-5022
> >
> > No virus found in this incoming message.
> > Checked by AVG.
> > Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> > 21/06/2008
> > 9:27 AM
> >
> >
> > No virus found in this outgoing message.
> > Checked by AVG.
> > Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> > 21/06/2008
> > 9:27 AM
> >
> >
>
>
> _________________________________________
> Michael Cammer   http://www.aecom.yu.edu/aif/
>
> No virus found in this incoming message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008 9:27 AM
>  
>
> No virus found in this outgoing message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1513 - Release Date: 22/06/2008 7:52 AM
>  
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Dale Callaham]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Wow, I didn't know I was keeping company with such a manipulative bunch!

I'm just replying to this last message as a general continuation of the
thread.

Where is the line? On a camera for color imaging we all do a white
balance; and maybe there is a flatfield or background subtract applied.
And the CCD probably has the "fix bad pixels" box checked. These have a
well-understood reason for use, but they are systematically manipulating
pixel values. Flatfields and "fix-bad-pixels" are not array processes in
that they alter specific pixels in specific ways to make the image look
better: they are altering values to nominally fix errors that would make
the captured image not appear as the observed image. Will these
manipulations be detected and flagged by the sleuths checking images?

Dale

Tina Carvalho wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>> 1. Often, in response to the referees, I have to change figures.
>> Maybe group them differently (which means changing letters) or
>> alter the labelling.  I suppose I should go back to the original
>> images and re-create the entire plate, but if the changes are
>> minor it's much easier to (for example) black out 'a' and paste
>> in 'b', or take out an arrow and put in a letter, etc.  All these
>> things would presumably show up as 'fraud' ....
>
> Yup. Sigh. This is the education part, and I guide people through this a
> lot. The trick is to learn to use Layers in Photoshop, and save every
> iteration (disk space is cheap) so that you can go back and move the
> images, letters, arrows, etc. on a figure plate. Once you do this one
> time, you realize how valuable it is!
>
>> 2.  A point nobody has yet raised is deconvolution imaging
>> systems.  Clinicians love them, but they have no training in physics
>> (and, to be honest, not much in science).  None of them has the
>> first idea what the software is doing, just that it's giving them
>> the pictures they want.  How does anyone handle that?
>
> Education. Give them a cheat sheet that lays out how it works. Give them a
> test. Whatever. I know it can be a pain, but researchers should (in an
> ideal world) understand what it is that they are doing. I can go on a rant
> about labeling "kits" where you follow directions, but never have any idea
> what is actually happening. I can go on lots of rants...
>
> Aloha, Tina
>
>> Optical Imaging Techniques in Cell Biology
>> by Guy Cox    CRC Press / Taylor & Francis
>>     http://www.guycox.com/optical.htm
>> ______________________________________________
>> Associate Professor Guy Cox, MA, DPhil(Oxon)
>> Electron Microscope Unit, Madsen Building F09,
>> University of Sydney, NSW 2006
>> ______________________________________________
>> Phone [phone]     Fax [phone]
>> Mobile [phone]
>> ______________________________________________
>>      http://www.guycox.net
>> -----Original Message-----
>> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Michael Cammer
>> Sent: Monday, 23 June 2008 2:30 AM
>> To: [hidden email]
>> Subject: Re: An alarming amount of image manipulation
>>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> In my experience, when somebody shows up with micrographs that have cells pasted in or background "distractions" stamp tooled out and I suggest maybe taking more pictures or doing the experiment again, they complain that this would cost more and delay them.  "The data are what they are and I'm just showing people and if I don't make it look good, then they won't publish it."  Sometimes junior lab members will go back to their PIs and say that I wasn't helpful; I become the problem.  I will not apologize for refusing to use the lasso tool around a band on a gel and S curve adjust it in Photoshop Curves or for refusing to paint out "an anomalous bad cell off in the corner of the image".  Yes, both these requests have come through our facility along with a bunch of other egregious ones.  Some scientists really feel justified because the competition does it.  "Hey, everybody does it," they say.  Although, more often (and a search will show that we've discussed this before), p

eople simply don;t understand what they are doing.  They simply misuse the tools for making figures.  As with example of film, before you can begin to alter images in an intentional manner or make even reasonably good pictures of anything, you need to be at least knowledgeable of darkroom technique.  But with the new digital tools, any dummy can be a photographer and can cut and paste and manipulate in complicated ways.

>> -Michael
>>
>>> Search the CONFOCAL archive at
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>> Interesting - especially the statement that images on film cannot be
>>> doctored. I guess it's irrelevant nowadays, but that is so far from
>>> the truth.
>>>
>>> One funny story - which I've told before but some time ago.
>>> Many years ago I (with colleagues) published a paper in a well-known
>>> journal which included a photo of some gels - taken by the
>>> departmental photographer.  One of the gels had cracked when it was
>>> taken out of the tube, so there was a dark hairline across it in the
>>> photo.  In the published paper that line had disappeared.  You can
>>> hardly accuse me, or my two co-authors, of fraud since this was done
>>> by the journal's art department without any reference to us!
>>>
>>>                                                        Guy
>>>
>>>
>>> Optical Imaging Techniques in Cell Biology
>>> by Guy Cox    CRC Press / Taylor & Francis
>>>     http://www.guycox.com/optical.htm
>>> ______________________________________________
>>> Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope Unit,
>>> Madsen Building F09, University of Sydney, NSW 2006
>>> ______________________________________________
>>> Phone [phone]     Fax [phone]
>>> Mobile [phone]
>>> ______________________________________________
>>>      http://www.guycox.net
>>> -----Original Message-----
>>> From: Confocal Microscopy List [mailto:[hidden email]]
>>> On Behalf Of John Oreopoulos
>>> Sent: Sunday, 22 June 2008 10:57 PM
>>> To: [hidden email]
>>> Subject: An alarming amount of image manipulation
>>>
>>> Search the CONFOCAL archive at
>>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>>
>>> I was a bit surprised at the statistics cited in this article:
>>>
>>> http://chronicle.com/free/2008/05/3028n.htm
>>>
>>> Does this mean that all journals will start hiring image manipulation
>>> detectives someday? Could be an interesting career.
>>>
>>>
>>> John Oreopoulos, BSc,
>>> PhD Candidate
>>> University of Toronto
>>> Institute For Biomaterials and Biomedical Engineering Centre For
>>> Studies in Molecular Imaging
>>>
>>> Tel: W:416-946-5022
>>>
>>> No virus found in this incoming message.
>>> Checked by AVG.
>>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
>>> 21/06/2008
>>> 9:27 AM
>>>
>>>
>>> No virus found in this outgoing message.
>>> Checked by AVG.
>>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
>>> 21/06/2008
>>> 9:27 AM
>>>
>>>
>>
>> _________________________________________
>> Michael Cammer   http://www.aecom.yu.edu/aif/
>>
>> No virus found in this incoming message.
>> Checked by AVG.
>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008 9:27 AM
>>  
>>
>> No virus found in this outgoing message.
>> Checked by AVG.
>> Version: 7.5.524 / Virus Database: 270.4.1/1513 - Release Date: 22/06/2008 7:52 AM
>>  
>>
>
> ****************************************************************************
> * Tina (Weatherby) Carvalho               * [hidden email]           *
> * Biological Electron Microscope Facility * [phone]                 *
> * University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
> ****************************************************************************

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> Where is the line? On a camera for color imaging we all do a white
> balance; and maybe there is a flatfield or background subtract applied.
> And the CCD probably has the "fix bad pixels" box checked. These have a
> well-understood reason for use, but they are systematically manipulating
> pixel values. Flatfields and "fix-bad-pixels" are not array processes in
> that they alter specific pixels in specific ways to make the image look
> better: they are altering values to nominally fix errors that would make
> the captured image not appear as the observed image. Will these
> manipulations be detected and flagged by the sleuths checking images?

As far as I understand it (and I may be totally wrong) white balance uses
the same algorithms as, say, Photoshop Levels, a channel at a time, so
it's "legitimate" (unreportable) by our rules. Flatfield correction would
sort of be Levels on a gradient, doesn't fit the criteria, so how about
reporting it? Bad pixels, well, a bad pixel obscurs data, so an arbitrary
fix based on neighboring pixels is about the same, except that now you
expect that pixel to be "true". How do you report that for an individual
pixel?

This is useful, though. Let's hash this out and come up with a consensus,
if possible.

Aloha, Tina

>
> Dale
>
> Tina Carvalho wrote:
> > Search the CONFOCAL archive at
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> >> 1. Often, in response to the referees, I have to change figures.
> >> Maybe group them differently (which means changing letters) or
> >> alter the labelling.  I suppose I should go back to the original
> >> images and re-create the entire plate, but if the changes are
> >> minor it's much easier to (for example) black out 'a' and paste
> >> in 'b', or take out an arrow and put in a letter, etc.  All these
> >> things would presumably show up as 'fraud' ....
> >
> > Yup. Sigh. This is the education part, and I guide people through this a
> > lot. The trick is to learn to use Layers in Photoshop, and save every
> > iteration (disk space is cheap) so that you can go back and move the
> > images, letters, arrows, etc. on a figure plate. Once you do this one
> > time, you realize how valuable it is!
> >
> >> 2.  A point nobody has yet raised is deconvolution imaging
> >> systems.  Clinicians love them, but they have no training in physics
> >> (and, to be honest, not much in science).  None of them has the
> >> first idea what the software is doing, just that it's giving them
> >> the pictures they want.  How does anyone handle that?
> >
> > Education. Give them a cheat sheet that lays out how it works. Give them a
> > test. Whatever. I know it can be a pain, but researchers should (in an
> > ideal world) understand what it is that they are doing. I can go on a rant
> > about labeling "kits" where you follow directions, but never have any idea
> > what is actually happening. I can go on lots of rants...
> >
> > Aloha, Tina
> >
> >> Optical Imaging Techniques in Cell Biology
> >> by Guy Cox    CRC Press / Taylor & Francis
> >>     http://www.guycox.com/optical.htm
> >> ______________________________________________
> >> Associate Professor Guy Cox, MA, DPhil(Oxon)
> >> Electron Microscope Unit, Madsen Building F09,
> >> University of Sydney, NSW 2006
> >> ______________________________________________
> >> Phone [phone]     Fax [phone]
> >> Mobile [phone]
> >> ______________________________________________
> >>      http://www.guycox.net
> >> -----Original Message-----
> >> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Michael Cammer
> >> Sent: Monday, 23 June 2008 2:30 AM
> >> To: [hidden email]
> >> Subject: Re: An alarming amount of image manipulation
> >>
> >> Search the CONFOCAL archive at
> >> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>
> >> In my experience, when somebody shows up with micrographs that have cells pasted in or background "distractions" stamp tooled out and I suggest maybe taking more pictures or doing the experiment again, they complain that this would cost more and delay them.  "The data are what they are and I'm just showing people and if I don't make it look good, then they won't publish it."  Sometimes junior lab members will go back to their PIs and say that I wasn't helpful; I become the problem.  I will not apologize for refusing to use the lasso tool around a band on a gel and S curve adjust it in Photoshop Curves or for refusing to paint out "an anomalous bad cell off in the corner of the image".  Yes, both these requests have come through our facility along with a bunch of other egregious ones.  Some scientists really feel justified because the competition does it.  "Hey, everybody does it," they say.  Although, more often (and a search will show that we've discussed this before),!

  p

> eople simply don;t understand what they are doing.  They simply misuse the tools for making figures.  As with example of film, before you can begin to alter images in an intentional manner or make even reasonably good pictures of anything, you need to be at least knowledgeable of darkroom technique.  But with the new digital tools, any dummy can be a photographer and can cut and paste and manipulate in complicated ways.
> >> -Michael
> >>
> >>> Search the CONFOCAL archive at
> >>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>>
> >>> Interesting - especially the statement that images on film cannot be
> >>> doctored. I guess it's irrelevant nowadays, but that is so far from
> >>> the truth.
> >>>
> >>> One funny story - which I've told before but some time ago.
> >>> Many years ago I (with colleagues) published a paper in a well-known
> >>> journal which included a photo of some gels - taken by the
> >>> departmental photographer.  One of the gels had cracked when it was
> >>> taken out of the tube, so there was a dark hairline across it in the
> >>> photo.  In the published paper that line had disappeared.  You can
> >>> hardly accuse me, or my two co-authors, of fraud since this was done
> >>> by the journal's art department without any reference to us!
> >>>
> >>>                                                        Guy
> >>>
> >>>
> >>> Optical Imaging Techniques in Cell Biology
> >>> by Guy Cox    CRC Press / Taylor & Francis
> >>>     http://www.guycox.com/optical.htm
> >>> ______________________________________________
> >>> Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope Unit,
> >>> Madsen Building F09, University of Sydney, NSW 2006
> >>> ______________________________________________
> >>> Phone [phone]     Fax [phone]
> >>> Mobile [phone]
> >>> ______________________________________________
> >>>      http://www.guycox.net
> >>> -----Original Message-----
> >>> From: Confocal Microscopy List [mailto:[hidden email]]
> >>> On Behalf Of John Oreopoulos
> >>> Sent: Sunday, 22 June 2008 10:57 PM
> >>> To: [hidden email]
> >>> Subject: An alarming amount of image manipulation
> >>>
> >>> Search the CONFOCAL archive at
> >>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>>
> >>> I was a bit surprised at the statistics cited in this article:
> >>>
> >>> http://chronicle.com/free/2008/05/3028n.htm
> >>>
> >>> Does this mean that all journals will start hiring image manipulation
> >>> detectives someday? Could be an interesting career.
> >>>
> >>>
> >>> John Oreopoulos, BSc,
> >>> PhD Candidate
> >>> University of Toronto
> >>> Institute For Biomaterials and Biomedical Engineering Centre For
> >>> Studies in Molecular Imaging
> >>>
> >>> Tel: W:416-946-5022
> >>>
> >>> No virus found in this incoming message.
> >>> Checked by AVG.
> >>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> >>> 21/06/2008
> >>> 9:27 AM
> >>>
> >>>
> >>> No virus found in this outgoing message.
> >>> Checked by AVG.
> >>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> >>> 21/06/2008
> >>> 9:27 AM
> >>>
> >>>
> >>
> >> _________________________________________
> >> Michael Cammer   http://www.aecom.yu.edu/aif/
> >>
> >> No virus found in this incoming message.
> >> Checked by AVG.
> >> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date: 21/06/2008 9:27 AM
> >>  
> >>
> >> No virus found in this outgoing message.
> >> Checked by AVG.
> >> Version: 7.5.524 / Virus Database: 270.4.1/1513 - Release Date: 22/06/2008 7:52 AM
> >>  
> >>
> >
> > ****************************************************************************
> > * Tina (Weatherby) Carvalho               * [hidden email]           *
> > * Biological Electron Microscope Facility * [phone]                 *
> > * University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
> > ****************************************************************************
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Csúcs Gábor]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Dear Tina,

Do you think that your guidelines are sufficient? If someone is allowed
to change the brightness/contrast and gamma settings for the different
fluorescent channels separately and save those "new", edited images
(especially if I do repeated cycles of this procedure) - it is pretty
easy to create completely new images - showing e.g. more or less
co-localization. The problem is that also the above mentioned procedures
are able to manipulate the information content of an image... (OK they
are always just reducing it - but still, one could use them to "cut out"
the unwanted stuff). Don't you think so?

Greetings   Gabor

--
Gabor Csucs
Light Microscopy Centre, ETH Zurich
Schafmattstrasse 18, HPM F16
CH-8093, Zurich, Switzerland

Web: www.lmc.ethz.ch
Phone: [phone]
Fax: [phone]
e-mail: [hidden email]

[Ignatius, Mike]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

What happened to JPEG 2000?  I recall a Congress of HCS imagers, where
this was the key watermark required in regulated situations. Any changes
to the original image were apparent by this capture mode?  

Also anyone interested in how images are abused, err manipulated in the
world of commerce, esp. fashion, would love this article in the New
Yorker, May 12th.
http://www.newyorker.com/reporting/2008/05/12/080512fa_fact_collins.
For example: "In the March issue of Vogue Dangin tweaked a hundred and
forty-four images..."  

Mike Ignatius

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On
Behalf Of Tina Carvalho
Sent: Monday, June 23, 2008 2:07 PM
To: [hidden email]
Subject: Re: An alarming amount of image manipulation

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> Where is the line? On a camera for color imaging we all do a white
> balance; and maybe there is a flatfield or background subtract
applied.
> And the CCD probably has the "fix bad pixels" box checked. These have
a
> well-understood reason for use, but they are systematically
manipulating
> pixel values. Flatfields and "fix-bad-pixels" are not array processes
in
> that they alter specific pixels in specific ways to make the image
look
> better: they are altering values to nominally fix errors that would
make
> the captured image not appear as the observed image. Will these
> manipulations be detected and flagged by the sleuths checking images?

As far as I understand it (and I may be totally wrong) white balance
uses
the same algorithms as, say, Photoshop Levels, a channel at a time, so
it's "legitimate" (unreportable) by our rules. Flatfield correction
would
sort of be Levels on a gradient, doesn't fit the criteria, so how about
reporting it? Bad pixels, well, a bad pixel obscurs data, so an
arbitrary
fix based on neighboring pixels is about the same, except that now you
expect that pixel to be "true". How do you report that for an individual
pixel?

This is useful, though. Let's hash this out and come up with a
consensus,
if possible.

Aloha, Tina

>
> Dale
>
> Tina Carvalho wrote:
> > Search the CONFOCAL archive at
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> >> 1. Often, in response to the referees, I have to change figures.
> >> Maybe group them differently (which means changing letters) or
> >> alter the labelling.  I suppose I should go back to the original
> >> images and re-create the entire plate, but if the changes are
> >> minor it's much easier to (for example) black out 'a' and paste
> >> in 'b', or take out an arrow and put in a letter, etc.  All these
> >> things would presumably show up as 'fraud' ....
> >
> > Yup. Sigh. This is the education part, and I guide people through

this a
> > lot. The trick is to learn to use Layers in Photoshop, and save
every
> > iteration (disk space is cheap) so that you can go back and move the
> > images, letters, arrows, etc. on a figure plate. Once you do this
one
> > time, you realize how valuable it is!
> >
> >> 2.  A point nobody has yet raised is deconvolution imaging
> >> systems.  Clinicians love them, but they have no training in
physics
> >> (and, to be honest, not much in science).  None of them has the
> >> first idea what the software is doing, just that it's giving them
> >> the pictures they want.  How does anyone handle that?
> >
> > Education. Give them a cheat sheet that lays out how it works. Give
them a
> > test. Whatever. I know it can be a pain, but researchers should (in
an
> > ideal world) understand what it is that they are doing. I can go on
a rant
> > about labeling "kits" where you follow directions, but never have
any idea

> > what is actually happening. I can go on lots of rants...
> >
> > Aloha, Tina
> >
> >> Optical Imaging Techniques in Cell Biology
> >> by Guy Cox    CRC Press / Taylor & Francis
> >>     http://www.guycox.com/optical.htm
> >> ______________________________________________
> >> Associate Professor Guy Cox, MA, DPhil(Oxon)
> >> Electron Microscope Unit, Madsen Building F09,
> >> University of Sydney, NSW 2006
> >> ______________________________________________
> >> Phone [phone]     Fax [phone]
> >> Mobile [phone]
> >> ______________________________________________
> >>      http://www.guycox.net
> >> -----Original Message-----
> >> From: Confocal Microscopy List

[mailto:[hidden email]] On Behalf Of Michael Cammer
> >> Sent: Monday, 23 June 2008 2:30 AM
> >> To: [hidden email]
> >> Subject: Re: An alarming amount of image manipulation
> >>
> >> Search the CONFOCAL archive at
> >> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>
> >> In my experience, when somebody shows up with micrographs that have
cells pasted in or background "distractions" stamp tooled out and I
suggest maybe taking more pictures or doing the experiment again, they
complain that this would cost more and delay them.  "The data are what
they are and I'm just showing people and if I don't make it look good,
then they won't publish it."  Sometimes junior lab members will go back
to their PIs and say that I wasn't helpful; I become the problem.  I
will not apologize for refusing to use the lasso tool around a band on a
gel and S curve adjust it in Photoshop Curves or for refusing to paint
out "an anomalous bad cell off in the corner of the image".  Yes, both
these requests have come through our facility along with a bunch of
other egregious ones.  Some scientists really feel justified because the
competition does it.  "Hey, everybody does it," they say.  Although,
more often (and a search will show that we've discussed this before),!
  p
> eople simply don;t understand what they are doing.  They simply misuse
the tools for making figures.  As with example of film, before you can
begin to alter images in an intentional manner or make even reasonably
good pictures of anything, you need to be at least knowledgeable of
darkroom technique.  But with the new digital tools, any dummy can be a
photographer and can cut and paste and manipulate in complicated ways.
> >> -Michael
> >>
> >>> Search the CONFOCAL archive at
> >>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>>
> >>> Interesting - especially the statement that images on film cannot
be
> >>> doctored. I guess it's irrelevant nowadays, but that is so far
from
> >>> the truth.
> >>>
> >>> One funny story - which I've told before but some time ago.
> >>> Many years ago I (with colleagues) published a paper in a
well-known
> >>> journal which included a photo of some gels - taken by the
> >>> departmental photographer.  One of the gels had cracked when it
was
> >>> taken out of the tube, so there was a dark hairline across it in
the
> >>> photo.  In the published paper that line had disappeared.  You can

> >>> hardly accuse me, or my two co-authors, of fraud since this was
done

> >>> by the journal's art department without any reference to us!
> >>>
> >>>                                                        Guy
> >>>
> >>>
> >>> Optical Imaging Techniques in Cell Biology
> >>> by Guy Cox    CRC Press / Taylor & Francis
> >>>     http://www.guycox.com/optical.htm
> >>> ______________________________________________
> >>> Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope

Unit,
> >>> Madsen Building F09, University of Sydney, NSW 2006
> >>> ______________________________________________
> >>> Phone [phone]     Fax [phone]
> >>> Mobile [phone]
> >>> ______________________________________________
> >>>      http://www.guycox.net
> >>> -----Original Message-----
> >>> From: Confocal Microscopy List
[mailto:[hidden email]]

> >>> On Behalf Of John Oreopoulos
> >>> Sent: Sunday, 22 June 2008 10:57 PM
> >>> To: [hidden email]
> >>> Subject: An alarming amount of image manipulation
> >>>
> >>> Search the CONFOCAL archive at
> >>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >>>
> >>> I was a bit surprised at the statistics cited in this article:
> >>>
> >>> http://chronicle.com/free/2008/05/3028n.htm
> >>>
> >>> Does this mean that all journals will start hiring image

manipulation

> >>> detectives someday? Could be an interesting career.
> >>>
> >>>
> >>> John Oreopoulos, BSc,
> >>> PhD Candidate
> >>> University of Toronto
> >>> Institute For Biomaterials and Biomedical Engineering Centre For
> >>> Studies in Molecular Imaging
> >>>
> >>> Tel: W:416-946-5022
> >>>
> >>> No virus found in this incoming message.
> >>> Checked by AVG.
> >>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> >>> 21/06/2008
> >>> 9:27 AM
> >>>
> >>>
> >>> No virus found in this outgoing message.
> >>> Checked by AVG.
> >>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> >>> 21/06/2008
> >>> 9:27 AM
> >>>
> >>>
> >>
> >> _________________________________________
> >> Michael Cammer   http://www.aecom.yu.edu/aif/
> >>
> >> No virus found in this incoming message.
> >> Checked by AVG.
> >> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:

21/06/2008 9:27 AM
> >>  
> >>
> >> No virus found in this outgoing message.
> >> Checked by AVG.
> >> Version: 7.5.524 / Virus Database: 270.4.1/1513 - Release Date:
22/06/2008 7:52 AM
> >>  
> >>
> >
> >
************************************************************************
****
> > * Tina (Weatherby) Carvalho               * [hidden email]
*
> > * Biological Electron Microscope Facility * [phone]
*
> > * University of Hawaii at Manoa           *
http://www.pbrc.hawaii.edu/bemf* 
> >
************************************************************************
****
>

************************************************************************
****
* Tina (Weatherby) Carvalho               * [hidden email]
*
* Biological Electron Microscope Facility * [phone]
*
* University of Hawaii at Manoa           *
http://www.pbrc.hawaii.edu/bemf* 
************************************************************************
****

[Stephen C. Kempf]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

I guess I'll add my two cents here. As I've said in the past (and  
this was true back in the darkroom days) it still comes down to the  
integrity of the investigator preparing the images. There are just  
about always ways to get around any safeguards against image fraud  
that are put in place. For example, if someone were to do an  
excellent job of producing a fraudulent image in photoshop, print it  
out, and then retake a digital picture of the figure, how easy would  
it be to detect manipulation? I realize that in certain cases this  
might be possible, but in other cases it would probably be very  
difficult.

I tend to think that good education and training of investigators,  
and subsequently, their students is the best route to go.

Just some thoughts,

Steve

On Jun 23, 2008, at 4:17 PM, Csucs Gabor wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Dear Tina,
>
> Do you think that your guidelines are sufficient? If someone is  
> allowed to change the brightness/contrast and gamma settings for  
> the different fluorescent channels separately and save those "new",  
> edited images (especially if I do repeated cycles of this  
> procedure) - it is pretty easy to create completely new images -  
> showing e.g. more or less co-localization. The problem is that also  
> the above mentioned procedures are able to manipulate the  
> information content of an image... (OK they are always just  
> reducing it - but still, one could use them to "cut out" the  
> unwanted stuff). Don't you think so?
>
> Greetings   Gabor
>
> --
> Gabor Csucs Light Microscopy Centre, ETH Zurich
> Schafmattstrasse 18, HPM F16 CH-8093, Zurich, Switzerland
>
> Web: www.lmc.ethz.ch
> Phone: [phone]
> Fax: [phone]
> e-mail: [hidden email]

[Christopher Gilpin]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

All,
I recently attended a workshop at NIST
http://www.nist.gov/public_affairs/confpage/080501.htm where the open forum
discussion centered around image metadata. The OME folks were there (amongst
others) and there was a consensus that there needs to be a formal way to
record metadata. Interesting suggestions were that each instrument in future
would contain a unique ID and this would be recorded along with images.
Unique codes embedded in images could identify "raw data". There could be a
way to record all image manipulations - at the instrument and subsequently -
by careful use of metadata. Some of the discussion was concerned with
availability of raw data for others to access and mine as new methods become
available or a new perspective was desired. There are of course many issues
surrounding this but note the way that protein structures have to be lodged
with the pdb as a condition of e.g. NIH funding.

Representatives from the microscopy community who attended also agreed that
NIST may have a role to play with the production of "best practice"
guidelines for microscopists. Whilst NIST does not have a mandate to enforce
standards it is their core business and they have a lot of experience.

Another thread to the discussion was the development of the "semantic web"
see e.g. the wiki at http://en.wikipedia.org/wiki/Semantic_Web

Eventually, data mining will be web-based but it will rely entirely on
having confidence in and records of how data was produced.

I am sure there will be wider discussion over time.

Chris


Christopher J Gilpin Ph.D.
Assistant Professor
Director, Molecular and Cellular Imaging Facility
K1.A04
UT Southwestern Medical Center at Dallas
5323 Harry Hines Boulevard
Dallas, TX 75390-9039
Phone [phone]
Fax [phone]

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi, Gabor-

> Do you think that your guidelines are sufficient? If someone is allowed
> to change the brightness/contrast and gamma settings for the different
> fluorescent channels separately and save those "new", edited images
> (especially if I do repeated cycles of this procedure) - it is pretty
> easy to create completely new images - showing e.g. more or less
> co-localization. The problem is that also the above mentioned procedures
> are able to manipulate the information content of an image... (OK they
> are always just reducing it - but still, one could use them to "cut out"
> the unwanted stuff). Don't you think so?

Yes, I do think it's pretty easy to enhance or cut out the different
channels. (I usually find myself bringing up a channel rather than
reducing a channel.) But you are doing this with your choice of filters or
your voltage on your PMTs of any number of other ways already. So the
manipulation can begin before saving the image. Changing the levels on
each channel after the fact can be really useful to bring out information
that *is* really there but not captured and then mixed well because of
your type of camera or the health of your PMTs, the software is weird, or
that monitor is deficient, or whatever, so from that standpoint it is a
good thing. Or you can make up false colocalization, which is a big can of
worms anyway. There are all kinds of pitfalls along the way. In every case
I feel it is incumbent on the researcher to understand how the system is
working and what manipulations they are making, the consequences, and then
know what to report so that it can be checked or reproduced.

Are the guidelines sufficient? No, but we couldn't think of a way to make
them any more easy to understand or implement. Unless you say that ANY
post-processing must be reported, including contrast and brightness and
levels. What do you think?

Aloha, Tina

> Greetings   Gabor
>
> --
> Gabor Csucs
> Light Microscopy Centre, ETH Zurich
> Schafmattstrasse 18, HPM F16
> CH-8093, Zurich, Switzerland
>
> Web: www.lmc.ethz.ch
> Phone: [phone]
> Fax: [phone]
> e-mail: [hidden email]
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi, Steve-

One reason I got into this is that I can make an entirely new species of
insect or copepod in Photoshop and probably you wouldn't be able to detect
it...!

It does boil down to the integrity of the researcher, but increasingly the
researcher has no idea what their actions during image acquisition and
post-processing for publication mean for the integrity of the data. Most
of the problems I see here are from people being uninformed, not
malicious. So I agree education and training are the solution. I'm doing
my personal best, but I can't vouch for anyone else on this campus! We've
got people who did not grow up with digital images mentoring students who
have never known anything different and are used to just clicking on
filters to get a pretty picture. If they come through my facility, they
get to hear me rant about it and, I hope, go away a little more
cautious. How to do this on a large scale? More explicit guidelines for
submissions to journals? Anyone want to help out with this? Volunteers
welcome. Join the Committee on the Ethics of Digital Imaging in the
Microscopy Society of America.

Aloha, Tina

> I guess I'll add my two cents here. As I've said in the past (and  
> this was true back in the darkroom days) it still comes down to the  
> integrity of the investigator preparing the images. There are just  
> about always ways to get around any safeguards against image fraud  
> that are put in place. For example, if someone were to do an  
> excellent job of producing a fraudulent image in photoshop, print it  
> out, and then retake a digital picture of the figure, how easy would  
> it be to detect manipulation? I realize that in certain cases this  
> might be possible, but in other cases it would probably be very  
> difficult.
>
> I tend to think that good education and training of investigators,  
> and subsequently, their students is the best route to go.
>
> Just some thoughts,
>
> Steve
>
> On Jun 23, 2008, at 4:17 PM, Csucs Gabor wrote:
>
> > Search the CONFOCAL archive at
> > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
> >
> > Dear Tina,
> >
> > Do you think that your guidelines are sufficient? If someone is  
> > allowed to change the brightness/contrast and gamma settings for  
> > the different fluorescent channels separately and save those "new",  
> > edited images (especially if I do repeated cycles of this  
> > procedure) - it is pretty easy to create completely new images -  
> > showing e.g. more or less co-localization. The problem is that also  
> > the above mentioned procedures are able to manipulate the  
> > information content of an image... (OK they are always just  
> > reducing it - but still, one could use them to "cut out" the  
> > unwanted stuff). Don't you think so?
> >
> > Greetings   Gabor
> >
> > --
> > Gabor Csucs Light Microscopy Centre, ETH Zurich
> > Schafmattstrasse 18, HPM F16 CH-8093, Zurich, Switzerland
> >
> > Web: www.lmc.ethz.ch
> > Phone: [phone]
> > Fax: [phone]
> > e-mail: [hidden email]
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Oh, good, some specific suggestions! I like both the NIST and wiki
ideas. Let's keep this going.

Mahalo,
Tina

> All,
> I recently attended a workshop at NIST
> http://www.nist.gov/public_affairs/confpage/080501.htm where the open forum
> discussion centered around image metadata. The OME folks were there (amongst
> others) and there was a consensus that there needs to be a formal way to
> record metadata. Interesting suggestions were that each instrument in future
> would contain a unique ID and this would be recorded along with images.
> Unique codes embedded in images could identify "raw data". There could be a
> way to record all image manipulations - at the instrument and subsequently -
> by careful use of metadata. Some of the discussion was concerned with
> availability of raw data for others to access and mine as new methods become
> available or a new perspective was desired. There are of course many issues
> surrounding this but note the way that protein structures have to be lodged
> with the pdb as a condition of e.g. NIH funding.
>
> Representatives from the microscopy community who attended also agreed that
> NIST may have a role to play with the production of "best practice"
> guidelines for microscopists. Whilst NIST does not have a mandate to enforce
> standards it is their core business and they have a lot of experience.
>
> Another thread to the discussion was the development of the "semantic web"
> see e.g. the wiki at http://en.wikipedia.org/wiki/Semantic_Web
>
> Eventually, data mining will be web-based but it will rely entirely on
> having confidence in and records of how data was produced.
>
> I am sure there will be wider discussion over time.
>
> Chris
>
>
> Christopher J Gilpin Ph.D.
> Assistant Professor
> Director, Molecular and Cellular Imaging Facility
> K1.A04
> UT Southwestern Medical Center at Dallas
> 5323 Harry Hines Boulevard
> Dallas, TX 75390-9039
> Phone [phone]
> Fax [phone]
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[ian gibbins]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

And then there are the scanners and image acquisition software that
tend to have default settings that include unsharp masking, contrast
stretching, automatic assignment of RGB channels, many of which pass
into the "data" completely unobserved by naive users... ther eis no
solution by eternal vigilance, I fear...


IAN


On Monday, June 23, 2008, at 08:52  PM, Guy Cox wrote:

> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> Two quite separate comments on this thread.
>
> 1. Often, in response to the referees, I have to change figures.
> Maybe group them differently (which means changing letters) or
> alter the labelling.  I suppose I should go back to the original
> images and re-create the entire plate, but if the changes are
> minor it's much easier to (for example) black out 'a' and paste
> in 'b', or take out an arrow and put in a letter, etc.  All these
> things would presumably show up as 'fraud' ....
>
> 2.  A point nobody has yet raised is deconvolution imaging
> systems.  Clinicians love them, but they have no training in physics
> (and, to be honest, not much in science).  None of them has the
> first idea what the software is doing, just that it's giving them
> the pictures they want.  How does anyone handle that?
>
>                                                    Guy
>
>
>
> Optical Imaging Techniques in Cell Biology
> by Guy Cox    CRC Press / Taylor & Francis
>     http://www.guycox.com/optical.htm
> ______________________________________________
> Associate Professor Guy Cox, MA, DPhil(Oxon)
> Electron Microscope Unit, Madsen Building F09,
> University of Sydney, NSW 2006
> ______________________________________________
> Phone [phone]     Fax [phone]
> Mobile [phone]
> ______________________________________________
>      http://www.guycox.net
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]]
> On Behalf Of Michael Cammer
> Sent: Monday, 23 June 2008 2:30 AM
> To: [hidden email]
> Subject: Re: An alarming amount of image manipulation
>
> Search the CONFOCAL archive at
> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
> In my experience, when somebody shows up with micrographs that have
> cells pasted in or background "distractions" stamp tooled out and I
> suggest maybe taking more pictures or doing the experiment again, they
> complain that this would cost more and delay them.  "The data are what
> they are and I'm just showing people and if I don't make it look good,
> then they won't publish it."  Sometimes junior lab members will go
> back to their PIs and say that I wasn't helpful; I become the problem.
>  I will not apologize for refusing to use the lasso tool around a band
> on a gel and S curve adjust it in Photoshop Curves or for refusing to
> paint out "an anomalous bad cell off in the corner of the image".  
> Yes, both these requests have come through our facility along with a
> bunch of other egregious ones.  Some scientists really feel justified
> because the competition does it.  "Hey, everybody does it," they say.  
> Although, more often (and a search will show that we've discussed this
> before), people simply don;t understand what they are doing.  They
> simply misuse the tools for making figures.  As with example of film,
> before you can begin to alter images in an intentional manner or make
> even reasonably good pictures of anything, you need to be at least
> knowledgeable of darkroom technique.  But with the new digital tools,
> any dummy can be a photographer and can cut and paste and manipulate
> in complicated ways.
> -Michael
>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> Interesting - especially the statement that images on film cannot be
>> doctored. I guess it's irrelevant nowadays, but that is so far from
>> the truth.
>>
>> One funny story - which I've told before but some time ago.
>> Many years ago I (with colleagues) published a paper in a well-known
>> journal which included a photo of some gels - taken by the
>> departmental photographer.  One of the gels had cracked when it was
>> taken out of the tube, so there was a dark hairline across it in the
>> photo.  In the published paper that line had disappeared.  You can
>> hardly accuse me, or my two co-authors, of fraud since this was done
>> by the journal's art department without any reference to us!
>>
>>                                                        Guy
>>
>>
>> Optical Imaging Techniques in Cell Biology
>> by Guy Cox    CRC Press / Taylor & Francis
>>     http://www.guycox.com/optical.htm
>> ______________________________________________
>> Associate Professor Guy Cox, MA, DPhil(Oxon) Electron Microscope Unit,
>> Madsen Building F09, University of Sydney, NSW 2006
>> ______________________________________________
>> Phone [phone]     Fax [phone]
>> Mobile [phone]
>> ______________________________________________
>>      http://www.guycox.net
>> -----Original Message-----
>> From: Confocal Microscopy List [mailto:[hidden email]]
>> On Behalf Of John Oreopoulos
>> Sent: Sunday, 22 June 2008 10:57 PM
>> To: [hidden email]
>> Subject: An alarming amount of image manipulation
>>
>> Search the CONFOCAL archive at
>> http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>>
>> I was a bit surprised at the statistics cited in this article:
>>
>> http://chronicle.com/free/2008/05/3028n.htm
>>
>> Does this mean that all journals will start hiring image manipulation
>> detectives someday? Could be an interesting career.
>>
>>
>> John Oreopoulos, BSc,
>> PhD Candidate
>> University of Toronto
>> Institute For Biomaterials and Biomedical Engineering Centre For
>> Studies in Molecular Imaging
>>
>> Tel: W:416-946-5022
>>
>> No virus found in this incoming message.
>> Checked by AVG.
>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
>> 21/06/2008
>> 9:27 AM
>>
>>
>> No virus found in this outgoing message.
>> Checked by AVG.
>> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
>> 21/06/2008
>> 9:27 AM
>>
>>
>
>
> _________________________________________
> Michael Cammer   http://www.aecom.yu.edu/aif/
>
> No virus found in this incoming message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1512 - Release Date:
> 21/06/2008 9:27 AM
>
>
> No virus found in this outgoing message.
> Checked by AVG.
> Version: 7.5.524 / Virus Database: 270.4.1/1513 - Release Date:
> 22/06/2008 7:52 AM
>
>
>

* * * * * * * * * * *
Prof Ian Gibbins
Anatomy & Histology
Flinders University
GPO Box 2100
Adelaide SA 5001
AUSTRALIA

[hidden email]
voice: [phone]
fax: [phone]

http://som.flinders.edu.au/FUSA/Anatomy/
http://www.flinders.edu.au/neuroscience

[Julio Vazquez]

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal =

As they say in Hungary (or so I was told), every good action gets its punishment. I think that people with a PhD, or about to get one, should know the difference between enhancing an image for display or publication, and misrepresentation (or fraud). If they are in doubt, they should also be smart enough to ask for advice. By trying to protect everybody against themselves, I am somewhat worried that out of all this, NIH or other agency will come up with a 1,500-page manual full with regulations and guidelines that will only make our lives just a little bit more complicated, probably with little impact on the amount of misconduct or cluelessness...  

I am currently analyzing cells labeled by FISH, and counting those that have nuclear and/or cytoplasmic staining. Nuclear spots are maybe 20-40 times brighter than the diffuse cytoplasmic signal. My eyes, camera, and computer monitor, all have different dynamic ranges and response curves. To see on the monitor what I see at the microscope, or to be able to print it, I need a pretty severe gamma adjustment to enhance the low intensities, otherwise I just will miss a lot of cells. This procedure however will change pixel intensities non-linearly, will not preserve intensity ratios between different regions of the image, and is pretty irreversible once applied. But that's OK... I know what I am doing and why I am doing it (and keeping the original data). On the other hand, without this gamma adjustment, the pictures I get on the screen (or paper) will just not match what I see at the microscope. We certainly don't want regulators telling us that non-linear contrast adjustment is no longer allowed.  

--

Julio Vazquez, PhD

Fred Hutchinson Cancer Research Center

1100 Fairview Ave. N.,  mailstop DE-512

Seattle, WA 98109-1024

http://www.fhcrc.org/

[Tina Carvalho]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> As they say in Hungary (or so I was told), every good action gets its  
> punishment. I think that people with a PhD, or about to get one,  
> should know the difference between enhancing an image for display or  
> publication, and misrepresentation (or fraud). If they are in doubt,  
> they should also be smart enough to ask for advice.

I agree here, as long as whomever they ask knows...

By trying to  
> protect everybody against themselves, I am somewhat worried that out  
> of all this, NIH or other agency will come up with a 1,500-page  
> manual full with regulations and guidelines that will only make our  
> lives just a little bit more complicated, probably with little impact  
> on the amount of misconduct or cluelessness...

Which is why MSA is trying (or at least I am, anyway) to come up with
guidelines that are easy to implement and make sense, to stave off further
complicated regulations. Input invited.

> I am currently analyzing cells labeled by FISH, and counting those  
> that have nuclear and/or cytoplasmic staining. Nuclear spots are  
> maybe 20-40 times brighter than the diffuse cytoplasmic signal. My  
> eyes, camera, and computer monitor, all have different dynamic ranges  
> and response curves. To see on the monitor what I see at the  
> microscope, or to be able to print it, I need a pretty severe gamma  
> adjustment to enhance the low intensities, otherwise I just will miss  
> a lot of cells. This procedure however will change pixel intensities  
> non-linearly, will not preserve intensity ratios between different  
> regions of the image, and is pretty irreversible once applied. But  
> that's OK... I know what I am doing and why I am doing it (and  
> keeping the original data). On the other hand, without this gamma  
> adjustment, the pictures I get on the screen (or paper) will just not  
> match what I see at the microscope. We certainly don't want  
> regulators telling us that non-linear contrast adjustment is no  
> longer allowed.

Same here, which is why I still think that our guidelines of being able to
adjust contrast, brightness, and levels/gamma makes sense.

Recap: The Microscopy Society of America's whitepaper says you can adjust
brightness and contrast, and levels/gamma over the entire image. Anything
else should be reported as manipulation or enhancement. And you need to
store the "original" as uncompressed TIFF on archival media. We haven't
lately looked at other formats, like RAW, so the latter may change. Again,
if anyone wants to make an argument for another format, just dive in.

Aloha, Tina

>
> --
> Julio Vazquez, PhD
> Fred Hutchinson Cancer Research Center
> 1100 Fairview Ave. N.,  mailstop DE-512
> Seattle, WA 98109-1024
>
>
> http://www.fhcrc.org/
>
>

****************************************************************************
* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
****************************************************************************

[Peter Humphreys]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

  It would be quite helpful if journals could decide a clear and simple
standard for figures. Many papers are a little light on detail of the
manipulations used, but we frequently see vague and contradictory
information in the instructions for authors.
Since the journals usually insist on CMYK and we usually image RGB, the
colourspace conversion also alters the image. As it is almost impossible to
find which colour profile a journal uses, it is very difficult to get and
accurate conversion (photoshop usually does the conversion poorly); surely
the journals should take some of the responsibility for this part of the
publication process. Over the years I've seen the journals require more and
more formatting from our end.

Peter


Peter Humphreys
Imaging Facility
Centre for Stem Cell Research
Cambridge
CB2 1QA


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On
Behalf Of Tina Carvalho
Sent: 24 June 2008 01:37
To: [hidden email]
Subject: Re: An alarming amount of image manipulation

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

> As they say in Hungary (or so I was told), every good action gets its  
> punishment. I think that people with a PhD, or about to get one,  
> should know the difference between enhancing an image for display or  
> publication, and misrepresentation (or fraud). If they are in doubt,  
> they should also be smart enough to ask for advice.

I agree here, as long as whomever they ask knows...

By trying to  
> protect everybody against themselves, I am somewhat worried that out  
> of all this, NIH or other agency will come up with a 1,500-page  
> manual full with regulations and guidelines that will only make our  
> lives just a little bit more complicated, probably with little impact  
> on the amount of misconduct or cluelessness...

Which is why MSA is trying (or at least I am, anyway) to come up with
guidelines that are easy to implement and make sense, to stave off further
complicated regulations. Input invited.

> I am currently analyzing cells labeled by FISH, and counting those  
> that have nuclear and/or cytoplasmic staining. Nuclear spots are  
> maybe 20-40 times brighter than the diffuse cytoplasmic signal. My  
> eyes, camera, and computer monitor, all have different dynamic ranges  
> and response curves. To see on the monitor what I see at the  
> microscope, or to be able to print it, I need a pretty severe gamma  
> adjustment to enhance the low intensities, otherwise I just will miss  
> a lot of cells. This procedure however will change pixel intensities  
> non-linearly, will not preserve intensity ratios between different  
> regions of the image, and is pretty irreversible once applied. But  
> that's OK... I know what I am doing and why I am doing it (and  
> keeping the original data). On the other hand, without this gamma  
> adjustment, the pictures I get on the screen (or paper) will just not  
> match what I see at the microscope. We certainly don't want  
> regulators telling us that non-linear contrast adjustment is no  
> longer allowed.

Same here, which is why I still think that our guidelines of being able to
adjust contrast, brightness, and levels/gamma makes sense.

Recap: The Microscopy Society of America's whitepaper says you can adjust
brightness and contrast, and levels/gamma over the entire image. Anything
else should be reported as manipulation or enhancement. And you need to
store the "original" as uncompressed TIFF on archival media. We haven't
lately looked at other formats, like RAW, so the latter may change. Again,
if anyone wants to make an argument for another format, just dive in.

Aloha, Tina

>
> --
> Julio Vazquez, PhD
> Fred Hutchinson Cancer Research Center
> 1100 Fairview Ave. N.,  mailstop DE-512
> Seattle, WA 98109-1024
>
>
> http://www.fhcrc.org/
>
>

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* Tina (Weatherby) Carvalho               * [hidden email]           *

* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf*

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[Ingela Parmryd]

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi,

The ease at which an operator can create colocalisation by changing the brightness/contrast settings is one of the reasons why the overlay method should not be used to assess colocalisation. Quantitation is a superior alternative, for instance by the Pearson correlation coefficient. The PCC is not sensitive to the actual intensities of the pixels but compares the overall correlation between pixels from two channels and like for any analysis, it is the raw images that should be analysed.

Regards,

Ingela Parmryd

-----Ursprungligt meddelande-----
Från: Confocal Microscopy List [mailto:[hidden email]] För Tina Carvalho
Skickat: den 24 juni 2008 00:07
Till: [hidden email]
Ämne: Re: An alarming amount of image manipulation

Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal

Hi, Gabor-

> Do you think that your guidelines are sufficient? If someone is allowed
> to change the brightness/contrast and gamma settings for the different
> fluorescent channels separately and save those "new", edited images
> (especially if I do repeated cycles of this procedure) - it is pretty
> easy to create completely new images - showing e.g. more or less
> co-localization. The problem is that also the above mentioned procedures
> are able to manipulate the information content of an image... (OK they
> are always just reducing it - but still, one could use them to "cut out"
> the unwanted stuff). Don't you think so?

Yes, I do think it's pretty easy to enhance or cut out the different
channels. (I usually find myself bringing up a channel rather than
reducing a channel.) But you are doing this with your choice of filters or
your voltage on your PMTs of any number of other ways already. So the
manipulation can begin before saving the image. Changing the levels on
each channel after the fact can be really useful to bring out information
that *is* really there but not captured and then mixed well because of
your type of camera or the health of your PMTs, the software is weird, or
that monitor is deficient, or whatever, so from that standpoint it is a
good thing. Or you can make up false colocalization, which is a big can of
worms anyway. There are all kinds of pitfalls along the way. In every case
I feel it is incumbent on the researcher to understand how the system is
working and what manipulations they are making, the consequences, and then
know what to report so that it can be checked or reproduced.

Are the guidelines sufficient? No, but we couldn't think of a way to make
them any more easy to understand or implement. Unless you say that ANY
post-processing must be reported, including contrast and brightness and
levels. What do you think?

Aloha, Tina

> Greetings   Gabor
>
> --
> Gabor Csucs
> Light Microscopy Centre, ETH Zurich
> Schafmattstrasse 18, HPM F16
> CH-8093, Zurich, Switzerland
>
> Web: www.lmc.ethz.ch
> Phone: [phone]
> Fax: [phone]
> e-mail: [hidden email]
>

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* Tina (Weatherby) Carvalho               * [hidden email]           *
* Biological Electron Microscope Facility * [phone]                 *
* University of Hawaii at Manoa           * http://www.pbrc.hawaii.edu/bemf* 
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