Analysis software
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Hi, I was wondering if confocal users have experience with VG studio max 3D analysis software and if so what they think of it compared to Image Pro Plus, metamorph, amira etc. DISCLAIMER: |
 
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Cameron Nowell |
Using FACS Dot Plots to Analyse Image Data
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Hi List, Has anyone out there ever tried to analyse morphology data
generated from HCS image analysis through a FACS software? We have a couple of
MetaMorph macros now that generate a scary amount of data. To be able to easily
visualise and analyse it, representing it like FACS data would be really beneficial.
We have the data in a spreadsheet with all the parameters listed for each
object/cell (size, shape, perimeter, width, length, texture, projected volume
etc). So the data is similar to FACS data but not in a format that can be
imported into FloJo or something similar. Cheers Cam Cameron J. Nowell Office: +61 3 9341 3155 |
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Csúcs Gábor |
Re: Using FACS Dot Plots to Analyse Image Data
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Hallo Cameron,
The Scan-R software from Olympus does exactly this although it is not a FACS sofware but a high-content imaging product. Cheers Gabor No commercial interest. > Hi List, > > > > Has anyone out there ever tried to analyse morphology data generated > from HCS image analysis through a FACS software? We have a couple of > MetaMorph macros now that generate a scary amount of data. To be able > to easily visualise and analyse it, representing it like FACS data > would be really beneficial. We have the data in a spreadsheet with all > the parameters listed for each object/cell (size, shape, perimeter, > width, length, texture, projected volume etc). So the data is similar > to FACS data but not in a format that can be imported into FloJo or > something similar. > > > > > > Cheers > > > > Cam > > > > > > *Cameron J. Nowell > *Microscopy Manager > Centre for Advanced Microscopy > Ludwig Institute for Cancer Research > PO Box 2008 > Royal Melbourne Hospital > Victoria, 3050 > AUSTRALIA > > *Office*: +61 3 9341 3155 > *Mobile*: +61422882700 > *Fax*: +61 3 9341 3104 > > Facility Website > <http://www.ludwig.edu.au/branch/research/platform/microscopy.htm> > > > > > -- Gabor Csucs Light Microscopy Centre, ETH Zurich Schafmattstrasse 18, HPM F16 CH-8093, Zurich, Switzerland Web: www.lmc.ethz.ch Phone: +41 44 633 6221 Mobile: +41 79 758 21 58 Fax: +41 44 632 1298 e-mail: [hidden email] |
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Martin Köhler |
Re: Using FACS Dot Plots to Analyse Image Data
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Hi Cameron and others,
BD Attovision also does exactly this, namely exporting of analyzed parameters in the fcs file format that can be further analyzed in the same way as FACS data. Attovision is the instrument software platform for the BD Pathway "high-content imaging products". Best, Martin No commercial interest. We are only users here. At 08:23 2009-09-02, you wrote: >Hallo Cameron, > >The Scan-R software from Olympus does exactly this although it is not a FACS sofware but a high-content imaging product. > >Cheers Gabor > >No commercial interest. > >>Hi List, >> >> >> >>Has anyone out there ever tried to analyse morphology data generated from HCS image analysis through a FACS software? We have a couple of MetaMorph macros now that generate a scary amount of data. To be able to easily visualise and analyse it, representing it like FACS data would be really beneficial. We have the data in a spreadsheet with all the parameters listed for each object/cell (size, shape, perimeter, width, length, texture, projected volume etc). So the data is similar to FACS data but not in a format that can be imported into FloJo or something similar. >> >> >> >> >> >>Cheers >> >> >> >>Cam >> >> >> >> >> >>*Cameron J. Nowell >>*Microscopy Manager >>Centre for Advanced Microscopy >>Ludwig Institute for Cancer Research >>PO Box 2008 >>Royal Melbourne Hospital >>Victoria, 3050 >>AUSTRALIA >> >>*Office*: +61 3 9341 3155 >>*Mobile*: +61422882700 >>*Fax*: +61 3 9341 3104 >> >>Facility Website <http://www.ludwig.edu.au/branch/research/platform/microscopy.htm> >> >> >>-- >Gabor Csucs Light Microscopy Centre, ETH Zurich >Schafmattstrasse 18, HPM F16 CH-8093, Zurich, Switzerland > >Web: www.lmc.ethz.ch >Phone: +41 44 633 6221 >Mobile: +41 79 758 21 58 >Fax: +41 44 632 1298 >e-mail: [hidden email] ------------------------------------------------------------- Martin Köhler The Rolf Luft Research Center for Diabetes and Endocrinology Karolinska Institutet Karolinska University Hospital Solna L1 SE-171 76 Stockholm, Sweden E-mail: [hidden email] Phone: +46 8 51775732 Fax: +46 8 51779450 ------------------------------------------------------------- |
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Emmanuel Gustin |
Re: Using FACS Dot Plots to Analyse Image Data
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In reply to this post by Cameron Nowell
Hello Cameron, With a bit of programming skill it is not too difficult to
generate FCS formatted files from a cell list in another format, because the FCS format is well documented:
The specification is on the site of the ISAC, www.isac-net.org.
The files might even be generated from an Excel macro. The spec
also allows a variation in which the data is represented as ASCII text instead of binary, which
is what I have used to write FCS files directly from PE Acapella – Acapella provides
a scripting environment with some basic file I/O tools, so it was possible to add this myself.
(A limitation of the FCS ASCII data representation is that the data values have to be integers,
but that can be more or less resolved by introducing scaling factors.) The existing FCS spec is very flexible, allowing you to define ‘channels’
representing any named feature, but I don’t know to what extent the existing
FACS analysis packages support that. I have written our own analysis software for
FCS data sets in 96 or 384 well format, so that I can fairly easily tweak the analysis software
for any specific requirements of the HCS data... Also, when exporting HCS data for a flow package, you probably have
to be careful to choose the best data representation. For example, if the distribution
of the values is lognormal rather than normal, it would probably be better to use
the logarithmic scaling option of the FCS spec (as many flow cytometers do for intensity
values), because that will generate better histograms. ISAC, who defined the FSC standard, is now involved in attempts
to define standards for HCS, and I hope that will produce a similar file format for cell
level data. Best Regards, Emmanuel -- From: Confocal Microscopy
List [mailto:[hidden email]] On Behalf Of Cameron
Nowell Hi List, Has anyone out there ever tried to analyse morphology data
generated from HCS image analysis through a FACS software? We have a couple of
MetaMorph macros now that generate a scary amount of data. To be able to easily
visualise and analyse it, representing it like FACS data would be really
beneficial. We have the data in a spreadsheet with all the parameters listed
for each object/cell (size, shape, perimeter, width, length, texture, projected
volume etc). So the data is similar to FACS data but not in a format that can
be imported into FloJo or something similar. Cheers Cam Cameron
J. Nowell Office: +61
3 9341 3155
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David Basiji |
Re: Using FACS Dot Plots to Analyse Image Data
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Hi Cameron,
Our IDEAS data analysis software does what you're trying to do without an export step,
but IDEAS can also export any image parameter in FCS format and we've learned a
few things trying to get other software to read our FCS-compliant
output.
To expand on Emmanuel's comments, we've found that the
most common issues with FCS export have to do with data linearity,
continuity, and range. FCS analysis has historically been performed on
integer-valued data with a range of 0-255 or 0-1023, so many of the FCS packages
default to that assumption. As Emmanuel mentioned, if you've got log-transformed
data, you need encode that in the FCS output so it's handled properly. You
should also do the same with floating point data and be particularly careful
with discrete (e.g. spot count), and unbounded distributions. If you run in to
problems, don't hesitate to contact the software vendor for advice, particularly
if you're using FlowJo or FCS Express. Both companies are highly responsive, in
my experience.
Best,
David
David Basiji, Ph.D. From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Gustin, Emmanuel [TIBBE] Sent: Wednesday, September 02, 2009 2:34 AM To: [hidden email] Subject: Re: Using FACS Dot Plots to Analyse Image Data Hello Cameron, With a bit of programming skill it is not too difficult
to generate FCS formatted files from a cell list in another format, because the FCS format is
well documented: The specification is on the site of the ISAC, www.isac-net.org. The files might even be generated from an Excel macro.
The spec also allows a variation in which the data is represented as ASCII text instead of
binary, which is what I have used to write FCS files directly from PE Acapella – Acapella
provides a scripting environment with some basic file I/O tools, so it was possible to add this
myself. (A limitation of the FCS ASCII data representation is that the data values have to be
integers, but that can be more or less resolved by introducing scaling
factors.) The existing FCS spec is very flexible, allowing you to
define ‘channels’ representing any named feature, but I don’t know to what extent the
existing FACS analysis packages support that. I have written our own analysis
software for FCS data sets in 96 or 384 well format, so that I can fairly easily tweak the analysis
software for any specific requirements of the HCS data... Also, when exporting HCS data for a flow package, you
probably have to be careful to choose the best data representation. For example, if the
distribution of the values is lognormal rather than normal, it would probably be better
to use the logarithmic scaling option of the FCS spec (as many flow cytometers do for
intensity values), because that will generate better histograms. ISAC, who defined the FSC standard, is now involved in
attempts to define standards for HCS, and I hope that will produce a similar file format
for cell level data. Best Regards, Emmanuel -- From: Confocal
Microscopy List [mailto:[hidden email]] On Behalf Of
Cameron Nowell Hi List, Has anyone out there ever tried to analyse
morphology data generated from HCS image analysis through a FACS software? We
have a couple of MetaMorph macros now that generate a scary amount of data. To
be able to easily visualise and analyse it, representing it like FACS data would
be really beneficial. We have the data in a spreadsheet with all the parameters
listed for each object/cell (size, shape, perimeter, width, length, texture,
projected volume etc). So the data is similar to FACS data but not in a format
that can be imported into FloJo or something similar. Cheers Cam Cameron J. Nowell Office:
+61 3 9341 3155 |
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Daniel James White |
Re: Using FACS Dot Plots to Analyse Image Data
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In reply to this post by Cameron Nowell
Hi Cameron,
On Sep 2, 2009, at 7:03 AM, CONFOCALMICROSCOPY automatic digest system wrote: > Hi List, > > =20 > > Has anyone out there ever tried to analyse morphology data generated > from HCS image analysis through a FACS software? For sure you can use the freeware Fiji (imageJ) plugins 2D histogram and colour inspector 3D do do FACS style data representations. (use control - L to search and find the menu commands of you cant see them) see http://pacific.mpi-cbg.de FIJI - is just ImageJ (Batteries Included) cheers Dan Dr. Daniel James White BSc. (Hons.) PhD Senior Microscopist / Image Visualisation, Processing and Analysis Light Microscopy and Image Processing Facilities Max Planck Institute of Molecular Cell Biology and Genetics Pfotenhauerstrasse 108 01307 DRESDEN Germany +49 (0)15114966933 (German Mobile) +49 (0)351 210 2627 (Work phone at MPI-CBG) +49 (0)351 210 1078 (Fax MPI-CBG LMF) http://www.bioimagexd.net BioImageXD http://pacific.mpi-cbg.de FIJI - is just ImageJ (Batteries Included) http://www.chalkie.org.uk Dan's Homepages https://info.med.tu-dresden.de/MTZimaging/index.php/Main_Page Dresden Imaging Facility Network dan (at) chalkie.org.uk ( white (at) mpi-cbg.de ) |
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