*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
HI
WE get horizontal lines with our zeiss LSM 510 when using transmission mode mostly. They increase/change frequency at faster scanning speeds and higher laser energy. We were told by our tech that it is stray light getting into the system. We have had to put blackout behind the existing black blinds we had across windows in our lab when doing transmission scans sometimes the light from the touch panel on the side of the zeiss is enough light but we turn it off if we are having real issues. WE don't have an issue if we do this and don't put the laser power/transmission up too high.
Liz Nickless
Microscopy and Imaging Centre
Senior Technical Officer
Analytical Development Section
Fonterra Innovations
Palmerston North
New Zealand
06 350 4649 x64527
-----Original Message-----
From: Confocal Microscopy List [mailto:[log in to unmask]] On Behalf Of JOEL B. SHEFFIELD
Sent: Monday, 3 September 2012 2:56 a.m.
To: [log in to unmask]
Subject: Fwd: horizontal dark lines appearing
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
*****
---------- Forwarded message ----------
From: JOEL B. SHEFFIELD <[log in to unmask]>
Date: Sun, Sep 2, 2012 at 10:46 AM
Subject: Re: horizontal dark lines appearing
To: [log in to unmask]
Hi Aryeh,
We have a Leica SP1 with an argon laser, and we too see these horizontal
lines at times, mostly with transmitted DIC images. In general, we remove
them with the FFT routines in ImageJ. Our Leica tech suggested that the
problem is that the polarization in the laser "wobbles" a bit, so that in
concert with the other polarized elements of the optical path, such as the
Wolleston filters, we get these horizontal patterns. Have you tried
removing all polarizing filters from the optical path?
Joel
On Sun, Sep 2, 2012 at 9:34 AM, Aryeh Weiss <[log in to unmask]> wrote:
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/**wa?A0=confocalmicroscopy<http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy>
> *****
>
> Thank you for many list members who responded on and off-list.
>
> I can summarize the responses as follows:
>
> 1. Some suspect the AOTF and/or the AOBS. Although other lasers do not
> exhibit the problem, these devices are tuned for each line. However, the
> fact that this problem occurs for each of the 4 argon lines, and not for
> 561 , 594 and 633, makes me doubt it is the AOTF.
>
> 2. Some suggest thermal issues. I checked the fans, and they all work (if
> they did not, that laser would shut down pretty fast). However, I cannot
> rule out a faulty thermal sensor, and am contacting Lasos to see how this
> can be checked.
>
> 3. I ran some scans with a fluorescent plastic slide. The results are
> uploaded to http://cl.ly/3K1L1S321925 . This is a LIF file, so you need a
> LIF reader for it. ImageJ will do, but it is easier to see the metadata
> with LAS Lite. Seems hard to find, but this link seems ok:
> http://cifweb.unil.ch/index.**php?option=com_docman&task=**
> doc_details&gid=26&Itemid=57<http://cifweb.unil.ch/index.php?option=com_docman&task=doc_details&gid=26&Itemid=57>
> This time I included the transmitted light images, which show the same
> effect. I think I can rule out photobleaching. There are various
> combinations of channels, simultaneous, and sequential scanning, using
> 488nm or 514nm (argon ion laser) and 561nm (DPSS laser).
>
> I again thank everyone who responded, and will update on the list as I
> obtain more information.
>
> --aryeh
> --
> Aryeh Weiss
> Faculty of Engineering
> Bar Ilan University
> Ramat Gan 52900 Israel
>
> Ph: 972-3-5317638
> FAX: 972-3-7384051
>
>
>
>> -----Original Message-----
>> From: Aryeh Weiss <[log in to unmask]>
>> Sender: Confocal Microscopy List <[log in to unmask]**EDU<[log in to unmask]>
>> >
>> Date: Fri, 31 Aug 2012 15:35:44
>> To: <[log in to unmask]**EDU<[log in to unmask]>
>> >
>> Reply-To: [log in to unmask]
>> Subject: horizontal dark lines appearing
>>
>>
>> We see (apparently) periodic horizontal dark lines with our Leica SP5,
>> when using the argon ion laser. Images that show these lines can be seen
>> at the following link:
>> http://cl.ly/2m2h0R1u1J2w
>>
>> There were some some posts on the list about this, with the suggestion
>> that the laser is dying. However, when last I checked (a few weeks ago),
>> that laser was putting out plenty of power (although the 488nm line was
>> relatively weak compared to the 514nm line). Also, the lines have some
>> periodicity (50Hz?). Faster scan rates increase the line spacing, but it
>> does not appear to be linear with scan speed.
>> Frame averaging will average it out, so it is not correlated with the
>> scanning system. So the lines are annoying, but average signal seems fine.
>>
>> None of the other lasers do this, and it is independent of detector.
>> Therefore, we assume that the problem is upstream of the vis laser
>> combiner/AOTF. That leaves the fiber, laser head, and power supply.
>>
>> We suspect the laser, but it would be more satisfying to understand what
>> causes this behavior. Understanding the cause may lead to knowing how to
>> fix it without replacing the laser, but even if not, I like to know why
>> things work (or dont work) the way they do.
>>
>> Has anyone has seen something similar?
>>
>> Thanks in advance
>> --aryeh
>>
>>
--
Joel B. Sheffield, Ph.D
Department of Biology
Temple University
Philadelphia, PA 19122
Voice: 215 204 8839
e-mail: [log in to unmask]
URL: http://astro.temple.edu/~jbs
--
Joel B. Sheffield, Ph.D
Department of Biology
Temple University
Philadelphia, PA 19122
Voice: 215 204 8839
e-mail: [log in to unmask]
URL: http://astro.temple.edu/~jbs
DISCLAIMER
This email contains information that is confidential and which may be legally privileged. If you have received this email in error, please notify the sender immediately and delete the email. This email is intended solely for the use of the intended recipient and you may not use or disclose this email in any way.
|
