Re: Ana alarming amount of image manipulation

[Mario-2]

(I apologize if people are getting this message twice. My problem is that my last couple of posts have not been acknowledged or sent back to me as part of the list. Thanks for your patience)

Oh Criminy,

I have tried to stay out of this. I now feel compelled to comment:

1. the human visual dynamic range under normal viewing conditions is much less that 8-bits (256 levels); its more like 6-bits (64 levels or worse). Thus, it is inevitable that microscopy images confocal or W.F. will require manipulation in intensity and contrast (and gamma correction) in order to generate prints or downloadable images that demonstrate the experimenters claims. It is rediculous that anyone should retract a paper because of minor intensity or contrast changes. It would be nice to know what procedures were carried out on an important data set and could be described in a few sentences. Further, I agree with advocates that original raw image data be made available by the journal and/or researchers upon request. In fact, uncompressed raw image files should be provided to the reviewers of a paper. Might save a lot time. As for Catherine Verfaillie and colleagues, without knowing the details, it seems like a black eye on common sense on the part of U of Minn. sorry don't mean to offend but consider item 2.

2.As for journals, different publishers use different settings for printing images. In my opinion, they must bear some of the responsibility for getting the images to print as the authors wish, but much of the time they tell the authors merely to enhance contrast because the journal prints tend to lose contrast and images appear washed out. I have experienced that even enhancing contrast to an absurd degree resulting in an image that is grossly inappropriate on my computer monitor, can still lead to inadequate contrast when printed in a journal! On the other hand, my experience with the journal Science was quite positive in that their Photoshop layout person worked with me to get the right print quality. I think it would be beneficial if all microscopy journals and others employing images did the same.

3. When it comes to creating fraudulent images free of manipulation artifacts, it is actually very time consuming and difficult, and I don't know why anyone would even bother. (Well actually, I do, but besides being immoral and unethical, it is not worth the trouble. You will be found out and your career will go down the drain). Anyway, before anybody gets paranoid, I know this from doing digital reconstructions of damaged historically important photographs (e.g., 90-100+ year old photogravures, metalochromes, platinum prints, etc). I try to create replicas of images that are as close to the originals as possible. Depending on the degree of damage or artifacts in an original print, it could take several weeks to be successful with a single 11"x17" original using the full image processing arsenal including Fourier analysis, building notch filters, etc.

Early in this thread it was mentioned that a histogram analysis can often reveal selective image tampering. I also think that this is a useful tool especially when it is applied with noise analysis on subsets of an image. Areas that have been manipulated by sharpening, blurring, selective intensity changes, and other types of local manipulation will create tell tale features that might not be visible to the eye but can be revealed by disproportionate statistical inconsistencies within the image as a whole. Systematic errors due to improperly adjusted PMT voltages or problems with amplifier gain dependent noise in a CCD camera can also be detected.

Mario

The issue of the recent request by the University of Minnesota to have Catherine Verfaillie retract her publication is worse than tragic: it has all the elements of a Puritanical witch-hunt, and all for the charge of changing the brightness level of three images.   Let me be quick to say that Morayma Reyes, the graduate student who was picked out for the "offense," changed brightness levels to conform the image for publication, exactly what a printer at the printing press would do (but somehow comes clean), and exactly what I advised her to do.

Believe me, no list of ethical rules by the Microscopy Society could EVER be written to have changed this outcome--which was clearly an investigation for political reasons--except one rule: no post-processing period (which is done anyway at the press: are they accountable, too?).

It is especially disheartening because Dr. Reyes is among the best researchers I have worked with.  I cannot forget the day when she called me into a room long after regular working hours and asked me to look through the microscope.  I saw a beating heart before this had ever been done with the use of stem cells.  I turned to her and commented, "This is too large for a rat heart and too small for a rabbit.  What is it?"  She smiled triumphantly and said, "We made it on a scaffold."

I agree wholeheartedly with Johan: fight back.  Especially for this so-called violation on research that has since been corroborated by other labs.

Jerry

-- 

________________________________________________________________________________

Mario M. Moronne, Ph.D.

[hidden email]
[hidden email]
[hidden email]

[mahogny]

Mario wrote:
...
> upon request. In fact, uncompressed raw image files should be provided
> to the reviewers of a paper. Might save a lot time. As for Catherine
> Verfaillie and colleagues, without knowing the details, it seems like
> a black eye on common sense on the part of U of Minn. sorry don't mean
> to offend but consider item 2.
it's about time to get this straight (mario might have just a typo here,
not claiming anything for him):

       compression does NOT imply reduced image quality

but it does for sure increase the speed of disk transfer, and if you
know what you are doing, some but not all compression algorithms can
*optionally* trade high frequency information (noise) for disk space.
avoid using the terminology that "compression" destroys images because
it confuses non-experts into thinking they should be storing the images
uncompressed. there's a factor 2-10 to gain in disk space/speed for
normal images.

/Johan (who is very tired of re-teaching about compression)

--
--
------------------------------------------------
Johan Henriksson
MSc Engineering
PhD student, Karolinska Institutet
http://mahogny.areta.org http://www.endrov.net

[Martin Wessendorf-2]

Dear Johan--

Johan Henriksson wrote:

>        compression does NOT imply reduced image quality
 >
> but it does for sure increase the speed of disk transfer, and if you
> know what you are doing, some but not all compression algorithms can
> *optionally* trade high frequency information (noise) for disk space.
> avoid using the terminology that "compression" destroys images because
> it confuses non-experts into thinking they should be storing the images
> uncompressed. there's a factor 2-10 to gain in disk space/speed for
> normal images.

Could you please elaborate about storing images compressed?  Are you
talking about lossless compression or (as it sounds like from the
context) lossy compression?

Thanks--

Martin Wessendorf
--
Martin Wessendorf, Ph.D.                   office: [phone]
Assoc Prof, Dept Neuroscience                 lab: [phone]
University of Minnesota             Preferred FAX: [phone]
6-145 Jackson Hall, 321 Church St. SE    Dept Fax: [phone]
Minneapolis, MN  55455
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[Mario-2]

Johan,

I have no problem with lossless compression, LZW etc. I do have a
problem with people using "high quality" jpegs which might look the
same, but introduce high frequency pixel level artifacts.
Colocalization and other correlation dependent imaging methods can
suffer. Resolution can suffer. As far as transfer rates, I am patient
and have no problem waiting 20 min to download a gigabyte image file
to my home office. Storage is an issue but I am of the opinion that
it is especially important to keep at least two copies of the
original unmanipulated data with at least one on a read only memory
format such as a DVD. Patents (or academic reputations) can succeed
or fail on being able to justify a claim.

>Mario wrote:
>...
>>  upon request. In fact, uncompressed raw image files should be provided
>>  to the reviewers of a paper. Might save a lot time. As for Catherine
>>  Verfaillie and colleagues, without knowing the details, it seems like
>>  a black eye on common sense on the part of U of Minn. sorry don't mean
>>  to offend but consider item 2.
>it's about time to get this straight (mario might have just a typo here,
>not claiming anything for him):
>
>        compression does NOT imply reduced image quality
>
>but it does for sure increase the speed of disk transfer, and if you
>know what you are doing, some but not all compression algorithms can
>*optionally* trade high frequency information (noise) for disk space.
>avoid using the terminology that "compression" destroys images because
>it confuses non-experts into thinking they should be storing the images
>uncompressed. there's a factor 2-10 to gain in disk space/speed for
>normal images.
>
>/Johan (who is very tired of re-teaching about compression)
>
>--
>--
>------------------------------------------------
>Johan Henriksson
>MSc Engineering
>PhD student, Karolinska Institutet
>http://mahogny.areta.org http://www.endrov.net

--
________________________________________________________________________________
Mario M. Moronne, Ph.D.

[hidden email]
[hidden email]
[hidden email]

[mahogny]

Martin Wessendorf wrote:

> Dear Johan--
>
> Johan Henriksson wrote:
>
>>        compression does NOT imply reduced image quality
> >
>> but it does for sure increase the speed of disk transfer, and if you
>> know what you are doing, some but not all compression algorithms can
>> *optionally* trade high frequency information (noise) for disk space.
>> avoid using the terminology that "compression" destroys images because
>> it confuses non-experts into thinking they should be storing the images
>> uncompressed. there's a factor 2-10 to gain in disk space/speed for
>> normal images.
>
> Could you please elaborate about storing images compressed?  Are you
> talking about lossless compression or (as it sounds like from the
> context) lossy compression?

I can sum up the current situation

======

there are 3 major open formats in use now:

TIFF. uncompressed or lossy JPEG. I think there is a rather bad lossless
format included that essentially no one supports
JPEG. lossy
PNG. lossless

TIFF supports 16bit but is rather awful overall. I recommend against it
whenever possible, because it is loosely specified. the result is that
some programs write images other programs cannot read. the problem with
JPEG and PNG is that they only support 8-bit images as far as I know,
and there are no real lossless formats competing with PNG. I would be
happy to see that someone extended PNG, the current algorithm with just
a larger word size would do the trick.

my benchmarks show that fluorescent images compress extremely well with
the PNG lossless algorithm so the only valid excuse not to use it at the
moment is if you have 12 or 16 bits. about a factor 10 in compression.
PNG does not work well on DIC images, about a factor 2.

JPEG, or any lossy compression, always wins over lossless since it in
practice applies a lowpass filter before storing down the data. JPEG
gives a factor 10 or more on DIC images, depending on your tradeoff.

======

in our lab, we use lossy compression (JPEG) on DIC images and lossless
(PNG) on fluorescent. motivation:
* DIC images give a qualitative impression, and as long as a human can
see what is there, we're fine.
* fluorescent images are used in quantification, and you have to be
careful with removing information.
* we cannot store all images with just lossless compression. we have
over 100 000 000 image planes, closing in on 2TB data.

under normal conditions we add 6GB data per day with the above scheme
(time lapse), as opposed to 50GB before I improved the setup. the
alternative to save space is to not capture an image at all, but I
consider this to be by far worse for research than storing images
without the noise.

======

the future: I would like to see some applied computer scientists work on
standardizing new formats. there is for example JPEG2000 which is by far
better than JPEG (better base functions), but cannot really be used
since no one implements it. likewise, it would be trivial to extend PNG
to higher pixel depths. but given that we now work with 3D datasets, or
even 5D, lossless compression rates can be improved even further, so
this should really be investigated.

there are more issues to be resolved. any format that stores all images
to one single file has all sorts of issues. some formats even store
images as XML, I won't even get started about these. then there is the
whole metadata issue. we started our own format standardization
initiative (for OST) since no format delivered what our lab required.

/Johan

--
--
------------------------------------------------
Johan Henriksson
MSc Engineering
PhD student, Karolinska Institutet
http://mahogny.areta.org http://www.endrov.net

[Turner, Scott]

While I agree that a certain amount of image processing should be allowed and in fact ought to be expected, the news story reported here actually misrepresents the conclusions of the University of Minnesota's ethics panel.  It is not simply that image brightness and contrast were changed; according to the conclusions of the panel (attached below) there were manipulations of images that could be construed as falsification of data.  These included "elimination of bands on blots, altered orientation of bands, introduction of lanes not included in the original figure, and covering objects or image density in certain lanes."  

I think that we can all agree that processing images is often desirable, and in some cases even necessary.  What should not happen, and according to the University of Minnesota did happen in this case, is the manipulation of images to present data in a way that are not consistent with the original content of the image.  Obviously, eliminating unwanted bands from a blot or altering the orientation of a band or including lanes not in the original figure go beyond image processing into the realm of data manipulation.  While it might be acceptable to brighten an image to make it more legible for print, it is not acceptable to cut and paste bands or lanes in a gel or alter the orientation of a band in order to better represent your data.


Scott Turner
Scientist II
Schering-Plough Biopharma
Palo Alto, CA



Statement from the University of Minnesota

University Misconduct Panel Concludes That Certain Data in Stem Cell Paper Were Falsified

University of Minnesota Vice President for Research Tim Mulcahy has accepted the conclusions of an academic misconduct committee impaneled by the University which found that certain data published in the journal Blood in 2001 in connection with federally sponsored stem cell research at the University were falsified.  The University has asked the journal to retract the article.  Vice President Mulcahy also accepted the findings of discrepancies, but not falsification, in certain data published in the Journal of Clinical Investigation.

Two current or former University employees were the subject of a complaint.  Dr. Catherine Verfaillie was previously a full-time tenured faculty member at the University of Minnesota. Dr. Verfaillie is currently the Director of the Stem Cell Institute at the Catholic University in Leuven, Belgium, and retains a 10 percent faculty appointment at the University of Minnesota. Dr. Morayma Reyes was a University of Minnesota student in the combined M.D./Ph.D. program who worked in Dr. Verfaillie's laboratory. Dr. Reyes is currently an Assistant Professor of Pathology at the University of Washington. None of the co-authors of the papers or other laboratory personnel were subjects of any complaints or findings.

The complaint was investigated by an investigation committee, chaired by Dr. David Bernlohr, Professor of Biochemistry, Molecular Biology and Biophysics at the University of Minnesota, and included Dr. Karen Reue, Professor of Human Genetics at the David Geffen School of Medicine - UCLA, and Dr. William Smith, Professor of Biological Chemistry at the University of Michigan. The panel was charged with investigating complaints against the respondents pursuant to federal regulation 42 C.F.R. § 93.310 and the University's Academic Misconduct Policy after an earlier inquiry conducted by Senior Administrator Charles Muscoplat concluded that there had been sufficient questions raised about the research to warrant a full investigation.

The investigation panel submitted its final report to the Senior Administrator on September 5, 2008.  The panel concluded that parts of four figures in the Blood paper were falsified.  Allegations against Dr. Verfaillie were unsubstantiated.  The findings with respect to Dr. Reyes are private student data and cannot be released under the Minnesota Government Data Practices Act and the federal Family Educational Rights and Privacy Act (FERPA).

The Senior Administrator accepted the panel's report on September 12, 2008, and forwarded it to the Vice President for Research, Tim Mulcahy, who is the senior University official responsible for oversight of academic misconduct proceedings.  Vice President Mulcahy reviewed the report, accepted the panel's conclusions and issued the University's final decision on September 24, 2008.  On September 25, 2008, Vice President Mulcahy transmitted the investigation panel's report and other required materials to the federal Office for Research Integrity for its review and action as required under federal rules governing research supported by the Public Health Service (PHS).

In four of seven figures in the Blood paper, the panel concluded that aspects of the figures were altered in such a way that the manipulation misrepresented experimental data and sufficiently altered the original research record to constitute falsification under federal regulations and University policy.  Manipulations identified by the panel included: elimination of bands on blots, altered orientation of bands, introduction of lanes not included in the original figure, and covering objects or image density in certain lanes.

In one case all exposures of the source data for the published image were missing.  While the panel could not conclude misconduct in this case, it concluded that the figure should be withdrawn as it cannot be substantiated by the existing experimental record.  The panel found no academic misconduct in the remaining two figures in the Blood paper.

The panel also considered three duplications of Fluorescent Activated Cells Sorting (FACS) data and incorrect labeling included in the article published in Blood, as well as two duplications of FACS data and incorrect labeling in a 2002 article published in the Journal of Clinical Investigation (JCI).  These latter discrepancies were self-reported to the University and JCI by Dr. Verfaillie prior to the initiation of the University's investigation.  In all cases, the panel concluded that no academic misconduct was associated with these FACS discrepancies.  With respect to the FACS discrepancies in the Blood paper, the panel noted poor scientific method and inadequate training and oversight for this research.  The panel made frequent reference to insufficient oversight throughout the report.

Based on the panel's findings, the University has requested that the article entitled "Purification and Ex Vivo Expansion of Postnatal Human Marrow and Mesodermal Progenitor Cells" published in the November 2001 edition of Blood be retracted.  Similarly, the University has notified the editorial office of the Journal of Clinical Investigation of the panel's findings in relation to the FACS discrepancies identified in the article entitled "Origin of Endothelial Progenitors in Human Post-Natal Bone Marrow" published in 2002.  As the panel did not find evidence of academic misconduct related to these figures, the University has not requested that the JCI paper be retracted.

The investigation panel also considered six discrepancies in two figures (Figures 6 and 10) included in an international patent application filed with the U.S. Patent and Trademark Office (USPTO) in August 2000 and again in a corresponding national stage filing dated August 2002.  While concluding that the figures were seriously flawed and not accurate data, there was insufficient evidence to conclude that misconduct occurred in connection with the patent applications.  Nevertheless, the panel recommended that the University notify the company holding the patent interests of these findings and cooperate with the company in making any appropriate disclosures to the USPTO.

The published version of Dr. Reyes' thesis contained all seven western blot discrepancies and three sets of FACS duplications included in the Blood paper.  The University's Student Conduct Code prohibits scholastic dishonesty and falsification in academic work. Student disciplinary proceedings are private, and information about student discipline can be released only in accordance with the Minnesota Government Data Practices Act and FERPA.
 


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Johan Henriksson
Sent: Wednesday, October 08, 2008 11:41 PM
To: [hidden email]
Subject: Re: An alarming amount of image manipulation - time to fight back

[hidden email] wrote:

> On Wed, 8 Oct 2008, John Oreopoulos wrote:
>
>> My apologies again if this discussion thread becomes heated. I
>> thought I'd pass on another news bit about image manipulation, this
>> time revolving around brightness and contrast:
>>
>> http://www.ctv.ca/servlet/ArticleNews/story/CTVNews/20081008/stemcell
>> _study_081008/20081008?hub=Health
>>
>>
>> Is this not something the journal should specify and the reviewers
>> should be looking for in the first place? It seems to me again that
>> all of this could be avoided if the authors simply state and describe
>> all image manipulations when submitting for publication
>>
>> John Oreopoulos
>>
>
> From the story you reference, the researcher did some minimal image
> processing that in no way altered the conclusions of the paper, and
> was hit on a religious objection that has no practical basis.  Doesn't
> sound to clever to me.
>
> The position these purists are taking is simply silly.  Were the same
> criteria in place before digital imaging, it would be "unethical" to
> produce prints from negatives -- or for that matter to even *develop*
> negatives at all -- since  all development and all printing
> *necessarily* involve "image processing."  When was the last time you
> created a print without affecting contrast and brightness?  Never?
> Hmmm....

by this logic, the only "ethical" way to include images would be as a table of sensor readouts in the supplementary material. I wonder who would be happy about that.

my stand point here is firm, any image manipulation is allowed. it is better by *default* to assume image processing. but the original image should always be made available in that case. the method description should come in form of a script to redo the operation using an open source package. I think this is how biologists should work with their data anyway because it allows them to redo the operation very easily on other images. as an additional advantage, checking correctedness can be almost automatic, the journal simply reruns the script. the only thing left to argue about is the choice of operations, left to the reviewers.

currently we have too many black boxes; deconvolution operations is one group of very important but hard to describe algorithms (the number of biologists here who has implemented it, raise your hand). I would not in any way be satisfied with a method description "was deconvolved with XXX" because I most likely do not have the package. you cannot expect a reviewer to suddenly shell out 10k usd just to verify a picture.

/Johan

--
--
------------------------------------------------
Johan Henriksson
MSc Engineering
PhD student, Karolinska Institutet
http://mahogny.areta.org http://www.endrov.net
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[Bill Oliver-3]

On Thu, 9 Oct 2008, Turner, Scott wrote:

> While I agree that a certain amount of image processing should be allowed and in fact ought to be expected, the news story reported here actually misrepresents the conclusions of the University of Minnesota's ethics panel.  It is not simply that image brightness and contrast were changed; according to the conclusions of the panel (attached below) there were manipulations of images that could be construed as falsification of data.  These included "elimination of bands on blots, altered orientation of bands, introduction of lanes not included in the original figure, and covering objects or image density in certain lanes."
>

Having been playing this game in the forensics arena for a couple of decades, I note the wiggle words are pertinent.  Of course it is possible to "construe" something as falsification of data.  I know expert witnesses that will "construe" anything you want for $400/hr.

Further, they are "construing" malpractice in a circular manner -- by their own definitions.  If you define changing brightness as "unethical" then changing brightness is "unethical."  It's not a matter of some absolute.

Let me give you an illustration from my primary specialty -- Forensic Pathology.  I apologize for the length of this, but I think it's important when faced with these kinds of appeals to authority.

So, here's the story:

A young man in Bay County, FL was caught joyriding in a stolen car.  He was convicted of car theft and was sentenced to a penal boot camp with the idea that a little discipline would put him on the right track.

One of the first things that the guards do when a young offender comes to one of these camps is make them exercise.  It wears them out and determines issues of authority.  The standard exercise regimen is the old Army standard of as many situps as one can do in two minutes, as many pushups as one can do in two minutes, and a two-mile run.

The young man did the situps and pushups, and ran about 1.5 miles.  He then stopped and refused to run any more.  The guards coerced him into running another lap, at which point he stopped and refused to run again.  The guards again attempted to coerce him, but he collapsed.  At the hospital he was found to be suffering from rhabdomyolysis, disseminated intravascular coagulation, and multiorgan failure.  He died.

At autopsy, he was found to have many sickled cells, and was found on evaluation to have sickle cell trait.  The Medical Examiner called it an exertional sickle cell trait death (a well-documented cause of death among young recruits in the military and in young athletes in training).

The NAACP and black caucus claimed that this was a race-based death due to beating (though, oddly, some of the guards were also black), and filed a complaint with the State Attorney General (who was running for governor).   A well-known TV pathologist flown in by Fox News opined that exertional sickle cell trait deaths did not exist.  Another expert opined a cause that had no previous example in history.

The Governor directed the Medical Examiner Commission (MEC) to review the cases done by this Medical Examiner and look for evidence of malpractice.

The MEC reviewed 700 of the Medical Examiner's cases and found 35 examples of "negligence," including such things as typing "2cm" instead of "2 cm" in a report, and the failure to measure the size of the base of a bullet (which is, in fact, a bad thing for a ME to do).  Further, at least three of the errors of omission were things that were explicitly stated in the state guidelines as things MEs should *not* do.  The MEC claimed to use standards that did not exist, but were created ad hoc for the purpose of removing this particular Medical Examiner.

Most important, however, after reviewing *700* cases of Dr. Siebert's the MEC could find *no* errors in diagnosis.  However, they claimed that this was unimportant, and that this kind of "negligence" was unacceptable.

On the basis of these ad hoc criteria, the MEC "construed" ethical violations and, at the order of the Governor, moved to fire the medical examiner.  The political basis for this was so egregious that the National Association of Medical Examiners wrote to the MEC that:

"As an organization, we believe that Dr. Siebert has met [the NAME] autopsy standards, and he continues to be a NAME member in good standing. By continuing to imply that Dr. Siebert does not meet the aforementioned nonexistent "NAME guidelines" or the NAME Autopsy Standards, the MEC is dishonestly misrepresenting the facts.  Furthermore, as these errors have not been publicly acknowledged by the MEC, the MEC is discrediting not only Dr. Siebert but NAME itself. Since the MEC apparently believes that falsely invoking the imprimatur of the National Association of Medical Examiners in this fashion is acceptable, the Executive Committee of NAME demands that the MEC officially acknowledge and make public retraction of the inconsistencies noted above..."

Of course, Dr. Siebert was still fired, and the guards were put on trial for manslaughter.  The medical testimony by the prosecution experts was so self-contradictory and the support for Dr. Siebert's diagnosis was so strong that the prosecution ended up asking the jury to *ignore* the medical testimony altogether.  After 90 minutes the jury found the guards not guilty.  The NAACP still insisted that Dr. Siebert be fired, and he was.  He now has a very successfull practice in his original home state of New Jersey.


See: http://en.wikipedia.org/wiki/Martin_Anderson_controversy
and http://www.billoblog.com/?p=271


Accordingly, I think that folk should be very careful about these kinds of appeals to authority.  Just because a committee holds a finding, it doesn't mean that it's valid. Just as it was telling that out of 700 cases, there were no errors in diagnosis, I think it is telling that out of all the frou frou in this case, nothing was changed that affected any of the conclusions.  Somehow that gets lost in these kinds of inquisitions.

Remember, when a commission decides to get you on the basis of process -- in any field of endeavor --  there are *no* innocents.  There are only the untargeted.

If one wants to cast stones at people for doing image processing, I think that one big criterion is whether or not it substantively affects the conclusion.  In this case it did not.

There's an old principle in law of "no harm, no foul."  That's not such a bad rule to use elsewhere.  There is no allegation of "harm,"  and in the absence of that, then "foul" is pretty much in the eye of the inquisitor.

billo

[James Pawley]

>Johan,
>
>I have no problem with lossless compression, LZW etc. I do have a
>problem with people using "high quality" jpegs which might look the
>same, but introduce high frequency pixel level artifacts.
>Colocalization and other correlation dependent imaging methods can
>suffer. Resolution can suffer. As far as transfer rates, I am
>patient and have no problem waiting 20 min to download a gigabyte
>image file to my home office. Storage is an issue but I am of the
>opinion that it is especially important to keep at least two copies
>of the original unmanipulated data with at least one on a read only
>memory format such as a DVD. Patents (or academic reputations) can
>succeed or fail on being able to justify a claim.
>

About the travesty in Minnesota: Shame! Shame!!

About saving storage space.

It is true that noisy images have a lot of high-frequency info in
them and that efforts to compress them must necessarily introduce
some changes that may confuse later efforts at image analysis.
However, if the brightest pixel in you data represents only16
photons, you lose nothing by clipping the data to 16 levels or 4
bits. This too will speed retrieval and husband disk space.

Of you could do what NASA used to do with its images and store only
the sqrt of the intensity number (expressed in photons or
photoelectrons). This means that you only store gray levels that are
significantly different from each other, but it also takes half the
disk space (4 bits for the signal above).

In any case, assuming that you have Nyquist-sampled your data, you
should always deconvolve it before viewing it. This not only averages
out a lot of that fuzzy noise, it also allows you to meet the Nyquist
reconstruction condition: that the bandwidth of the output device
(i.e., the computer plus the LCD screen) is the same as that of the
input the device (i.e, the diffraction-limited microscope)

Hope that you are all surviving better than your hedge funds.

Jim P.

>>Mario wrote:
>>...
>>>  upon request. In fact, uncompressed raw image files should be provided
>>>  to the reviewers of a paper. Might save a lot time. As for Catherine
>>>  Verfaillie and colleagues, without knowing the details, it seems like
>>>  a black eye on common sense on the part of U of Minn. sorry don't mean
>>>  to offend but consider item 2.
>>it's about time to get this straight (mario might have just a typo here,
>>not claiming anything for him):
>>
>>        compression does NOT imply reduced image quality
>>
>>but it does for sure increase the speed of disk transfer, and if you
>>know what you are doing, some but not all compression algorithms can
>>*optionally* trade high frequency information (noise) for disk space.
>>avoid using the terminology that "compression" destroys images because
>>it confuses non-experts into thinking they should be storing the images
>>uncompressed. there's a factor 2-10 to gain in disk space/speed for
>>normal images.
>>
>>/Johan (who is very tired of re-teaching about compression)
>>
>>--
>>--
>>------------------------------------------------
>>Johan Henriksson
>>MSc Engineering
>>PhD student, Karolinska Institutet
>>http://mahogny.areta.org http://www.endrov.net
>
>
>--
>________________________________________________________________________________
>Mario M. Moronne, Ph.D.
>
>[hidden email]
>[hidden email]
>[hidden email]

--
               ****************************************
Prof. James B. Pawley,                 Ph.  608-263-3147
Room 223, Zoology Research Building,                         FAX  [phone]
250 N. Mills St., Madison, WI, 53706  [hidden email]
"A scientist is not one who can answer questions but one who can
question answers."  Theodore Schick Jr., Skeptical Enquirer, 21-2:39

[Mark Cannell]

I totally absolutely with Scott's point wrote. I'd like to suggest that
a simpler definition of acceptable image manipulation might be: That
there exists a continuous transfer function between input and output?

Regards Mark Cannell

Turner, Scott wrote:

> While I agree that a certain amount of image processing should be allowed and in fact ought to be expected, the news story reported here actually misrepresents the conclusions of the University of Minnesota's ethics panel.  It is not simply that image brightness and contrast were changed; according to the conclusions of the panel (attached below) there were manipulations of images that could be construed as falsification of data.  These included "elimination of bands on blots, altered orientation of bands, introduction of lanes not included in the original figure, and covering objects or image density in certain lanes."  
>
> I think that we can all agree that processing images is often desirable, and in some cases even necessary.  What should not happen, and according to the University of Minnesota did happen in this case, is the manipulation of images to present data in a way that are not consistent with the original content of the image.  Obviously, eliminating unwanted bands from a blot or altering the orientation of a band or including lanes not in the original figure go beyond image processing into the realm of data manipulation.  While it might be acceptable to brighten an image to make it more legible for print, it is not acceptable to cut and paste bands or lanes in a gel or alter the orientation of a band in order to better represent your data.
>
>
> Scott Turner
> Scientist II
> Schering-Plough Biopharma
> Palo Alto, CA
>
>
>
> Statement from the University of Minnesota
>
> University Misconduct Panel Concludes That Certain Data in Stem Cell Paper Were Falsified
>
> University of Minnesota Vice President for Research Tim Mulcahy has accepted the conclusions of an academic misconduct committee impaneled by the University which found that certain data published in the journal Blood in 2001 in connection with federally sponsored stem cell research at the University were falsified.  The University has asked the journal to retract the article.  Vice President Mulcahy also accepted the findings of discrepancies, but not falsification, in certain data published in the Journal of Clinical Investigation.
>
> Two current or former University employees were the subject of a complaint.  Dr. Catherine Verfaillie was previously a full-time tenured faculty member at the University of Minnesota. Dr. Verfaillie is currently the Director of the Stem Cell Institute at the Catholic University in Leuven, Belgium, and retains a 10 percent faculty appointment at the University of Minnesota. Dr. Morayma Reyes was a University of Minnesota student in the combined M.D./Ph.D. program who worked in Dr. Verfaillie's laboratory. Dr. Reyes is currently an Assistant Professor of Pathology at the University of Washington. None of the co-authors of the papers or other laboratory personnel were subjects of any complaints or findings.
>
> The complaint was investigated by an investigation committee, chaired by Dr. David Bernlohr, Professor of Biochemistry, Molecular Biology and Biophysics at the University of Minnesota, and included Dr. Karen Reue, Professor of Human Genetics at the David Geffen School of Medicine - UCLA, and Dr. William Smith, Professor of Biological Chemistry at the University of Michigan. The panel was charged with investigating complaints against the respondents pursuant to federal regulation 42 C.F.R. § 93.310 and the University's Academic Misconduct Policy after an earlier inquiry conducted by Senior Administrator Charles Muscoplat concluded that there had been sufficient questions raised about the research to warrant a full investigation.
>
> The investigation panel submitted its final report to the Senior Administrator on September 5, 2008.  The panel concluded that parts of four figures in the Blood paper were falsified.  Allegations against Dr. Verfaillie were unsubstantiated.  The findings with respect to Dr. Reyes are private student data and cannot be released under the Minnesota Government Data Practices Act and the federal Family Educational Rights and Privacy Act (FERPA).
>
> The Senior Administrator accepted the panel's report on September 12, 2008, and forwarded it to the Vice President for Research, Tim Mulcahy, who is the senior University official responsible for oversight of academic misconduct proceedings.  Vice President Mulcahy reviewed the report, accepted the panel's conclusions and issued the University's final decision on September 24, 2008.  On September 25, 2008, Vice President Mulcahy transmitted the investigation panel's report and other required materials to the federal Office for Research Integrity for its review and action as required under federal rules governing research supported by the Public Health Service (PHS).
>
> In four of seven figures in the Blood paper, the panel concluded that aspects of the figures were altered in such a way that the manipulation misrepresented experimental data and sufficiently altered the original research record to constitute falsification under federal regulations and University policy.  Manipulations identified by the panel included: elimination of bands on blots, altered orientation of bands, introduction of lanes not included in the original figure, and covering objects or image density in certain lanes.
>
> In one case all exposures of the source data for the published image were missing.  While the panel could not conclude misconduct in this case, it concluded that the figure should be withdrawn as it cannot be substantiated by the existing experimental record.  The panel found no academic misconduct in the remaining two figures in the Blood paper.
>
> The panel also considered three duplications of Fluorescent Activated Cells Sorting (FACS) data and incorrect labeling included in the article published in Blood, as well as two duplications of FACS data and incorrect labeling in a 2002 article published in the Journal of Clinical Investigation (JCI).  These latter discrepancies were self-reported to the University and JCI by Dr. Verfaillie prior to the initiation of the University's investigation.  In all cases, the panel concluded that no academic misconduct was associated with these FACS discrepancies.  With respect to the FACS discrepancies in the Blood paper, the panel noted poor scientific method and inadequate training and oversight for this research.  The panel made frequent reference to insufficient oversight throughout the report.
>
> Based on the panel's findings, the University has requested that the article entitled "Purification and Ex Vivo Expansion of Postnatal Human Marrow and Mesodermal Progenitor Cells" published in the November 2001 edition of Blood be retracted.  Similarly, the University has notified the editorial office of the Journal of Clinical Investigation of the panel's findings in relation to the FACS discrepancies identified in the article entitled "Origin of Endothelial Progenitors in Human Post-Natal Bone Marrow" published in 2002.  As the panel did not find evidence of academic misconduct related to these figures, the University has not requested that the JCI paper be retracted.
>
> The investigation panel also considered six discrepancies in two figures (Figures 6 and 10) included in an international patent application filed with the U.S. Patent and Trademark Office (USPTO) in August 2000 and again in a corresponding national stage filing dated August 2002.  While concluding that the figures were seriously flawed and not accurate data, there was insufficient evidence to conclude that misconduct occurred in connection with the patent applications.  Nevertheless, the panel recommended that the University notify the company holding the patent interests of these findings and cooperate with the company in making any appropriate disclosures to the USPTO.
>
> The published version of Dr. Reyes' thesis contained all seven western blot discrepancies and three sets of FACS duplications included in the Blood paper.  The University's Student Conduct Code prohibits scholastic dishonesty and falsification in academic work. Student disciplinary proceedings are private, and information about student discipline can be released only in accordance with the Minnesota Government Data Practices Act and FERPA.
>  
>
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Johan Henriksson
> Sent: Wednesday, October 08, 2008 11:41 PM
> To: [hidden email]
> Subject: Re: An alarming amount of image manipulation - time to fight back
>
> [hidden email] wrote:
>  
>> On Wed, 8 Oct 2008, John Oreopoulos wrote:
>>
>>    
>>> My apologies again if this discussion thread becomes heated. I
>>> thought I'd pass on another news bit about image manipulation, this
>>> time revolving around brightness and contrast:
>>>
>>> http://www.ctv.ca/servlet/ArticleNews/story/CTVNews/20081008/stemcell
>>> _study_081008/20081008?hub=Health
>>>
>>>
>>> Is this not something the journal should specify and the reviewers
>>> should be looking for in the first place? It seems to me again that
>>> all of this could be avoided if the authors simply state and describe
>>> all image manipulations when submitting for publication
>>>
>>> John Oreopoulos
>>>
>>>      
>> From the story you reference, the researcher did some minimal image
>> processing that in no way altered the conclusions of the paper, and
>> was hit on a religious objection that has no practical basis.  Doesn't
>> sound to clever to me.
>>
>> The position these purists are taking is simply silly.  Were the same
>> criteria in place before digital imaging, it would be "unethical" to
>> produce prints from negatives -- or for that matter to even *develop*
>> negatives at all -- since  all development and all printing
>> *necessarily* involve "image processing."  When was the last time you
>> created a print without affecting contrast and brightness?  Never?
>> Hmmm....
>>    
> by this logic, the only "ethical" way to include images would be as a table of sensor readouts in the supplementary material. I wonder who would be happy about that.
>
> my stand point here is firm, any image manipulation is allowed. it is better by *default* to assume image processing. but the original image should always be made available in that case. the method description should come in form of a script to redo the operation using an open source package. I think this is how biologists should work with their data anyway because it allows them to redo the operation very easily on other images. as an additional advantage, checking correctedness can be almost automatic, the journal simply reruns the script. the only thing left to argue about is the choice of operations, left to the reviewers.
>
> currently we have too many black boxes; deconvolution operations is one group of very important but hard to describe algorithms (the number of biologists here who has implemented it, raise your hand). I would not in any way be satisfied with a method description "was deconvolved with XXX" because I most likely do not have the package. you cannot expect a reviewer to suddenly shell out 10k usd just to verify a picture.
>
> /Johan
>
> --
> --
> ------------------------------------------------
> Johan Henriksson
> MSc Engineering
> PhD student, Karolinska Institutet
> http://mahogny.areta.org http://www.endrov.net
> *********************************************************************
> This message and any attachments are solely for the
> intended recipient. If you are not the intended recipient,
> disclosure, copying, use or distribution of the information
> included in this message is prohibited -- Please
> immediately and permanently delete.
>  

[Koo, Lily (NIH/NIAID) [E]]

Hello all,

One of my colleagues is conducting an experiment in which she studies
the diffusion of her membrane protein tagged with Cerulean; also present
in her cell is another membrane protein tagged with Venus.  Her control
experiments seem to suggest a possibility that Cerulean and Venus may
bind to each other.  

Has anyone had similar observations or suspicion, at least in cell
membranes?

Thanks,

Lily

[vb-2]

Hi Lily,

Yes, they will bind each other at the membrane. Adding L221K, F223R
mutations to the Cerulean and all three mutations to venusYFP may reduce
this hetero-dimerization (Cerulean has the A206K mutation, as published).

Depending on the protein and the level of overexpression (as many young
folks are in a "blind love" with a very strong CMV promoter and/or COS-7 or
293T cells which are transgenic for T-large antigen), CeFP-venusYFP
co-aggregation is a well known phenomena.

If you have any specific questions, please contact me off-line.

Vitaly

NCI-Frederick,
301-846-6575

----- Original Message -----
From: "Koo, Lily (NIH/NIAID) [F]" <[hidden email]>
To: <[hidden email]>
Sent: Friday, October 10, 2008 9:15 AM
Subject: Cerulean Venus binding possible?

> Hello all,
>
> One of my colleagues is conducting an experiment in which she studies
> the diffusion of her membrane protein tagged with Cerulean; also present
> in her cell is another membrane protein tagged with Venus.  Her control
> experiments seem to suggest a possibility that Cerulean and Venus may
> bind to each other.
>
> Has anyone had similar observations or suspicion, at least in cell
> membranes?
>
> Thanks,
>
> Lily
>

[Turner, Scott]

I was not making "appeals to authority" (I don't know anything about the
qualifications of the committee at U of Minnesota) but simply making the
point that the news article gave a misleading impression of the
committee's findings.  If the committee had determined that image
contrast and brightness were the only adjustments made to the images
then I think we could and perhaps should be outraged.  If other
manipulations were made, then that opens another area of debate.  As
Rosemary pointed out, two images were determined by the committee to be
identical except for their orientation (one rotated and flipped relative
to the other).  This is fairly obvious when looking at figures 5c and 6b
in the paper.  Though this may not have been intentional, it is clearly
more egregious then simply omitting a space between the numeral "2" and
the unit "cm".

Scott Turner
Scientist II
Schering-Plough Biopharma
Palo Alto, CA




-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]]
On Behalf Of [hidden email]
Sent: Thursday, October 09, 2008 07:33 AM
To: [hidden email]
Subject: Re: An alarming amount of image manipulation - time to fight
back

On Thu, 9 Oct 2008, Turner, Scott wrote:

> While I agree that a certain amount of image processing should be
allowed and in fact ought to be expected, the news story reported here
actually misrepresents the conclusions of the University of Minnesota's
ethics panel.  It is not simply that image brightness and contrast were
changed; according to the conclusions of the panel (attached below)
there were manipulations of images that could be construed as
falsification of data.  These included "elimination of bands on blots,
altered orientation of bands, introduction of lanes not included in the
original figure, and covering objects or image density in certain
lanes."
>

Having been playing this game in the forensics arena for a couple of
decades, I note the wiggle words are pertinent.  Of course it is
possible to "construe" something as falsification of data.  I know
expert witnesses that will "construe" anything you want for $400/hr.

Further, they are "construing" malpractice in a circular manner -- by
their own definitions.  If you define changing brightness as "unethical"
then changing brightness is "unethical."  It's not a matter of some
absolute.

Let me give you an illustration from my primary specialty -- Forensic
Pathology.  I apologize for the length of this, but I think it's
important when faced with these kinds of appeals to authority.

So, here's the story:

A young man in Bay County, FL was caught joyriding in a stolen car.  He
was convicted of car theft and was sentenced to a penal boot camp with
the idea that a little discipline would put him on the right track.

One of the first things that the guards do when a young offender comes
to one of these camps is make them exercise.  It wears them out and
determines issues of authority.  The standard exercise regimen is the
old Army standard of as many situps as one can do in two minutes, as
many pushups as one can do in two minutes, and a two-mile run.

The young man did the situps and pushups, and ran about 1.5 miles.  He
then stopped and refused to run any more.  The guards coerced him into
running another lap, at which point he stopped and refused to run again.
The guards again attempted to coerce him, but he collapsed.  At the
hospital he was found to be suffering from rhabdomyolysis, disseminated
intravascular coagulation, and multiorgan failure.  He died.

At autopsy, he was found to have many sickled cells, and was found on
evaluation to have sickle cell trait.  The Medical Examiner called it an
exertional sickle cell trait death (a well-documented cause of death
among young recruits in the military and in young athletes in training).

The NAACP and black caucus claimed that this was a race-based death due
to beating (though, oddly, some of the guards were also black), and
filed a complaint with the State Attorney General (who was running for
governor).   A well-known TV pathologist flown in by Fox News opined
that exertional sickle cell trait deaths did not exist.  Another expert
opined a cause that had no previous example in history.

The Governor directed the Medical Examiner Commission (MEC) to review
the cases done by this Medical Examiner and look for evidence of
malpractice.

The MEC reviewed 700 of the Medical Examiner's cases and found 35
examples of "negligence," including such things as typing "2cm" instead
of "2 cm" in a report, and the failure to measure the size of the base
of a bullet (which is, in fact, a bad thing for a ME to do).  Further,
at least three of the errors of omission were things that were
explicitly stated in the state guidelines as things MEs should *not* do.
The MEC claimed to use standards that did not exist, but were created ad
hoc for the purpose of removing this particular Medical Examiner.

Most important, however, after reviewing *700* cases of Dr. Siebert's
the MEC could find *no* errors in diagnosis.  However, they claimed that
this was unimportant, and that this kind of "negligence" was
unacceptable.

On the basis of these ad hoc criteria, the MEC "construed" ethical
violations and, at the order of the Governor, moved to fire the medical
examiner.  The political basis for this was so egregious that the
National Association of Medical Examiners wrote to the MEC that:

"As an organization, we believe that Dr. Siebert has met [the NAME]
autopsy standards, and he continues to be a NAME member in good
standing. By continuing to imply that Dr. Siebert does not meet the
aforementioned nonexistent "NAME guidelines" or the NAME Autopsy
Standards, the MEC is dishonestly misrepresenting the facts.
Furthermore, as these errors have not been publicly acknowledged by the
MEC, the MEC is discrediting not only Dr. Siebert but NAME itself. Since
the MEC apparently believes that falsely invoking the imprimatur of the
National Association of Medical Examiners in this fashion is acceptable,
the Executive Committee of NAME demands that the MEC officially
acknowledge and make public retraction of the inconsistencies noted
above..."

Of course, Dr. Siebert was still fired, and the guards were put on trial
for manslaughter.  The medical testimony by the prosecution experts was
so self-contradictory and the support for Dr. Siebert's diagnosis was so
strong that the prosecution ended up asking the jury to *ignore* the
medical testimony altogether.  After 90 minutes the jury found the
guards not guilty.  The NAACP still insisted that Dr. Siebert be fired,
and he was.  He now has a very successfull practice in his original home
state of New Jersey.


See: http://en.wikipedia.org/wiki/Martin_Anderson_controversy
and http://www.billoblog.com/?p=271


Accordingly, I think that folk should be very careful about these kinds
of appeals to authority.  Just because a committee holds a finding, it
doesn't mean that it's valid. Just as it was telling that out of 700
cases, there were no errors in diagnosis, I think it is telling that out
of all the frou frou in this case, nothing was changed that affected any
of the conclusions.  Somehow that gets lost in these kinds of
inquisitions.

Remember, when a commission decides to get you on the basis of process
-- in any field of endeavor --  there are *no* innocents.  There are
only the untargeted.

If one wants to cast stones at people for doing image processing, I
think that one big criterion is whether or not it substantively affects
the conclusion.  In this case it did not.

There's an old principle in law of "no harm, no foul."  That's not such
a bad rule to use elsewhere.  There is no allegation of "harm,"  and in
the absence of that, then "foul" is pretty much in the eye of the
inquisitor.

billo
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This message and any attachments are solely for the
intended recipient. If you are not the intended recipient,
disclosure, copying, use or distribution of the information
included in this message is prohibited -- Please
immediately and permanently delete.

[Bill Oliver-3]

On Fri, 10 Oct 2008, Turner, Scott wrote:

> I was not making "appeals to authority" (I don't know anything about the
> qualifications of the committee at U of Minnesota) but simply making the
> point that the news article gave a misleading impression of the
> committee's findings... [snip]


Well, I looked at the article, and you are right.  Looking at this as an image processing guy who only stumbled through a lab using electrophoresis now and then, it's pretty clear that some of those illustrations have been banged on pretty hard. It's more than simple color balancing and optimization.

And, yes, it's clear that 5c and 6b are the same data.

That's pretty sloppy.

However, in my lifetime, particularly in dealing with resident physicians and the occasional grad student, it's usually wrong to "construe" a lack of ethics when recognizing that stupid errors are much more common.

If everybody that made an error in an illustration was "construed" to be unethical, there aren't a whole lot of folk who would have jobs.

But once again, the issue is with the *illustration,* not the underlying findings.  If the authors had said something that *wasn't true* in their underlying data, I would would be more sympathetic with the committee's torch-and-pitchfork approach to dealing with such issues.  But this idea that a mistake in an illustration constitutes a severe ethical violation is dangerous ground.  How far do you really want to go in this kind of inquisition?  Ask not for whom the bell tolls, and all that.


billo

[Kurt Thorn]

It's been well documented that GFP and its derivatives will dimerize.  
The Kd is about 100 uM; mutating A206 to K or R abolishes this dimerization.

See http://www.ncbi.nlm.nih.gov/pubmed/11988576 for the details.

Kurt

Koo, Lily (NIH/NIAID) [F] wrote:

> Hello all,
>
> One of my colleagues is conducting an experiment in which she studies
> the diffusion of her membrane protein tagged with Cerulean; also present
> in her cell is another membrane protein tagged with Venus.  Her control
> experiments seem to suggest a possibility that Cerulean and Venus may
> bind to each other.  
>
> Has anyone had similar observations or suspicion, at least in cell
> membranes?
>
> Thanks,
>
> Lily
>
>  

--
Kurt Thorn, PhD
Director, Nikon Imaging Center
University of California San Francisco

UCSF MC 2140
Genentech Hall Room S252
600 16th St.
San Francisco, CA 94158-2517

http://nic.ucsf.edu
phone [phone]
fax   [phone]

[Blancaflor, Elison]

A colleague requested that I post the job opportunities at Monsanto for
cell biologist/confocal microscopist below. Please contact Dr. Mary
Fernandes at Monsanto directly if anyone has questions about the
advertised positions.


Dr. Mary Fernandes

Model Systems Lead

Monsanto Biotechnology

Chesterfield, MO 63017

e-mail: [hidden email]



MS position (microscopy)--# 8520

MONS - 00008520

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We are seeking a highly talented and experienced scientist with
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priority leads in our crop pipeline.  Successful applicants will work in
a team based environment, manage individual project(s) and interact
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Required Skills:

Applicants should have a degree in Plant Biology, Molecular Biology,
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with proteins and/or nucleic acids is required.  Demonstrated skills in
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advantageous.   Familiarity with routine bioinformatics tools used in
the above fields is desirable.  The successful candidate will be a
highly motivated and results-oriented team player with excellent oral
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PhD position Mol. Biologist - # 9321

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Responsibilities:

We are seeking a highly talented and experienced scientist with
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larger multi-disciplinary effort aimed at developing methods for
discovery and advancing the knowledge base around high priority leads in
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Required Skills:

Successful candidates will have a PhD degree in Molecular Biology, Cell
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