*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
Hi all,
I would like to hear a little about where this data is coming from.
Am I right in assuming that it is related to a photon image of some
sort. Probably not confocal, where we virtually never get as many as
4,096 photons/pixel.
But even if it is the output from some widefield LM on a CCD or
sCMOS, and the saturation level really does correspond to 65k photons
(or at least 65k photoelectrons), I can't get all that worried about
whether it is 65,000 or 65,016, when the Poisson Noise on such a
signal is +/- 256 photoelectrons. Even if you manage somehow to get
your staining to be so specific that there is almost no background
(say 100:1), then the darkest pixel would be 650 but the Poison Noise
will still be +/-25.6 photoelectrons.
Usually in fluorescence microscopy, we are working at the other end
of the scale: maybe 2,000 PE/pixel peak, 20 PE in the darkest area.
In this case, I don't see a lot of reason to worry about the
interpolation levels, as long as you have the camera-gain set so that
one count in the memory equals about half the read noise of the
camera in e/pixel.
Cheers,
Jim Pawley
***************************************************************************
Prof. James B. Pawley, Ph.
608-238-3953
21. N. Prospect Ave. Madison, WI 53726 USA
[log in to unmask]
3D Microscopy of Living Cells Course, June 11-23, 2011, UBC, Vancouver Canada
Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2011
"If it ain't diffraction, it must be statistics." Anon.
>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>*****
>
>Metamorph also maintains the 12 bit data leaving the 4 extra bits empty.
>
>The software in the Zeiss LSM 510 gives you the option of doing that
>(12 bit Tiffs) or expanding the scale to fill up the 16 bit range.
>
>I don't think there is interpolation or "data creation" involved,
>just a simple linear scaling, right? x 16?
>
>I think the motivation is precisely to deal with programs like
>photoshop that do not scale the displays and confuse the users... I
>have dealt with that in our imaging core for too long, some users
>would not hesitate to throw away information by converting all their
>12 bit images to 8 bits just so they display right in photoshop...
>dumb
>
>Leoncio A. Vergara MD
>Assistant Director
>Center for Biomedical Engineering
>Assistant Professor
>Microbiology and Immunology
>University of Texas Medical Branch
>409-750-2153 (cell)
>
>
>-----Original Message-----
>From: Confocal Microscopy List
>[mailto:[log in to unmask]] On Behalf Of Jerry
>(Gerald) Sedgewick
>Sent: Thursday, February 03, 2011 11:12 AM
>To: [log in to unmask]
>Subject: Re: 12- to 16-bit images
>
>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>*****
>
>I would like to make a table of manufacturer's image formats that
>separate scaled 16-bit images from those image formats in which 12-bits
>of tonal data (0-4095 tones) are placed in a 16-bit (0-65,535) "package."
>
>As an example, images saved as 16-bit TIFF files in SPOT software are
>scaled from 12-bits to 16-bits, so these contain the full range of tones
>(at least in earlier versions of the software). Olympus .oib and
>exported TIFF files, on the other hand, contain 12-bits of tonal range
>within a 16-bit image, leaving 4 "empty" bits.
>
>In programs that auto-scale for display purposes, like Image J, the
>12-bit inside 16-bit looks great on the screen; but, in programs like
>Photoshop in which images are NOT auto-scaled for display purposes, the
>images are completely black, because only 1/16th of the 16-bit tonal
>range is displayed.
>
>I don't have access to all image formats that retain the 12-bit original
>tonal range within a 16-bit image: I know of only Olympus and the DVC
>company.
>
>I would greatly appreciate it if participants on this list could let me
>know of other manufacturer's image formats that retain 12-bit tonal
>range inside a 16-bit image. Or maybe a good source for downloading
>Zeiss, Leica, Nikon, Andor, Hamamatsu, etc., sample images. This table
>will be posted on the web for those who are interested.
>
>Thanks!
>
>Jerry
>
>--
>IMAGING: Image Integrity, Quantitation, Digital Imaging Instruction
>
>Jerry Sedgewick
>Sedgewick Initiatives
>965 Cromwell Avenue
>Saint Paul, MN 55114
>651-788-2261
>
>[log in to unmask]
>[log in to unmask]
>
>http://www.imagingandanalysis.com
>http://www.quickphotoshop.com
>
>Author of: "Scientific Imaging with Photoshop: Methods, Measurement, and
>Output"
--
***************************************************************************
Prof. James B. Pawley, Ph.
608-238-3953
21. N. Prospect Ave. Madison, WI 53726 USA
[log in to unmask]
3D Microscopy of Living Cells Course, June 11-23, 2011, UBC, Vancouver Canada
Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2011
"If it ain't diffraction, it must be statistics." Anon.
|
