Confocal Microscopy List

Zeiss Z1 optical table

Classic

List

Threaded

14 messages Options

Nuno Moreno

Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Dearz

this is not a confocal related question but I saw in a demo a small compressor for an active dumping optical table that comes with the Zeiss Z1. It had quite a low footprint and it was pretty silent. Does anyone have one around for sharing brand and model?

best,
Nuno Moreno

Julian Smith III

Re: Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

SilentAire Super-Silent 20A. We use one on our benchtop isolator and another to move the collection screen in/out of the column for our TEM digital camera.
Julian

-----Original Message-----
From: Confocal Microscopy List on behalf of Nuno Moreno
Sent: Mon 8/18/2014 11:47 AM
To: [hidden email]
Subject: Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
https://urldefense.proofpoint.com/v1/url?u=http://lists.umn.edu/cgi-bin/wa?A0%3Dconfocalmicroscopy&k=LzsPWH25gRR3YYS3VmwyEA%3D%3D%0A&r=Y%2BDJidFvffs1mPiUdTAbICUkqL90AXa4VdwC6vYS4%2Bg%3D%0A&m=Fh%2B4vTiAz8RS159qzZN%2B8qJB6V9Xsr1EO4NLEuTSe%2B8%3D%0A&s=e6dc2361cfe0940dc977d38cfb03bb11f1404d2a1031aca7f9f564fd25453c97
Post images on https://urldefense.proofpoint.com/v1/url?u=http://www.imgur.com/&k=LzsPWH25gRR3YYS3VmwyEA%3D%3D%0A&r=Y%2BDJidFvffs1mPiUdTAbICUkqL90AXa4VdwC6vYS4%2Bg%3D%0A&m=Fh%2B4vTiAz8RS159qzZN%2B8qJB6V9Xsr1EO4NLEuTSe%2B8%3D%0A&s=caf0148d2e59aa415e69652879ff497d323f5aae80463e098240eaaac35610a8 and include the link in your posting.
*****

Dearz

this is not a confocal related question but I saw in a demo a small compressor for an active dumping optical table that comes with the Zeiss Z1. It had quite a low footprint and it was pretty silent. Does anyone have one around for sharing brand and model?

best,
Nuno Moreno

Julio Vazquez

Re: Zeiss Z1 optical table

In reply to this post by Nuno Moreno

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

This may not be the same one, but we have one from Werther International S.p.A. , Model P 15-TC 110/60 (US 110 Volt model). I had seen this one at a Leica demo facility for one of their SP8 tables.

Haven't used it yet, but I remember it being very quiet.

http://www.werther.com/ClientProductDetails.asp?ProductID=1

best,

Julio Vazquez
Fred Hutchinson Cancer Research Center
Seattle, WA 98109

http://www.fhcrc.org/en.html

On Aug 18, 2014, at 8:47 AM, Nuno Moreno wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dearz
>
> this is not a confocal related question but I saw in a demo a small compressor for an active dumping optical table that comes with the Zeiss Z1. It had quite a low footprint and it was pretty silent. Does anyone have one around for sharing brand and model?
>
> best,
> Nuno Moreno

Smith, Benjamin E.

Re: Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

We also have the Werther International Panther silent air compressor running three airtables off the same compressor, one large table for the SP8, and two smaller tables for the FV500 and the ApoTome. The only sound is a release valve each time it finishes refilling the compressor tank. Maintenance involves popping the emergency release valve once a week, draining any water from the compressor tank once a month, and disassembling the compressor and replacing the compressor oil once per year.

Ben Smith
University of Oklahoma
________________________________________
From: Confocal Microscopy List [[hidden email]] on behalf of Julio Vazquez [[hidden email]]
Sent: Monday, August 18, 2014 11:36 AM
To: [hidden email]
Subject: Re: Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

This may not be the same one, but we have one from Werther International S.p.A. , Model P 15-TC 110/60 (US 110 Volt model). I had seen this one at a Leica demo facility for one of their SP8 tables.

Haven't used it yet, but I remember it being very quiet.

http://www.werther.com/ClientProductDetails.asp?ProductID=1

best,

Julio Vazquez
Fred Hutchinson Cancer Research Center
Seattle, WA 98109

http://www.fhcrc.org/en.html

On Aug 18, 2014, at 8:47 AM, Nuno Moreno wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Dearz
>
> this is not a confocal related question but I saw in a demo a small compressor for an active dumping optical table that comes with the Zeiss Z1. It had quite a low footprint and it was pretty silent. Does anyone have one around for sharing brand and model?
>
> best,
> Nuno Moreno

Craig Brideau

Re: Zeiss Z1 optical table

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

I also use the Werther Panther P15. It produces less noise that a typical 4
deg-C fridge. The first time we turned it on we thought we forgot to plug
it in because it was so quiet. We had to touch it to tell if it was on or
not. I've noticed the same valve release sounds as Ben. We have a very dry
environment so we don't have water in the compressor tank that often if
ever. We change out the oil roughly yearly.

Craig

On Mon, Aug 18, 2014 at 10:58 AM, Smith, Benjamin E. <[hidden email]>
wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> We also have the Werther International Panther silent air compressor
> running three airtables off the same compressor, one large table for the
> SP8, and two smaller tables for the FV500 and the ApoTome. The only sound
> is a release valve each time it finishes refilling the compressor tank.
> Maintenance involves popping the emergency release valve once a week,
> draining any water from the compressor tank once a month, and disassembling
> the compressor and replacing the compressor oil once per year.
>
> Ben Smith
> University of Oklahoma
> ________________________________________
> From: Confocal Microscopy List [[hidden email]] on
> behalf of Julio Vazquez [[hidden email]]
> Sent: Monday, August 18, 2014 11:36 AM
> To: [hidden email]
> Subject: Re: Zeiss Z1 optical table
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> This may not be the same one, but we have one from Werther International
> S.p.A. , Model P 15-TC 110/60 (US 110 Volt model). I had seen this one at a
> Leica demo facility for one of their SP8 tables.
>
> Haven't used it yet, but I remember it being very quiet.
>
> http://www.werther.com/ClientProductDetails.asp?ProductID=1
>
> best,
>
>
> Julio Vazquez
> Fred Hutchinson Cancer Research Center
> Seattle, WA 98109
>
> http://www.fhcrc.org/en.html
>
>
>
> On Aug 18, 2014, at 8:47 AM, Nuno Moreno wrote:
>
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Dearz
> >
> > this is not a confocal related question but I saw in a demo a small
> compressor for an active dumping optical table that comes with the Zeiss
> Z1. It had quite a low footprint and it was pretty silent. Does anyone have
> one around for sharing brand and model?
> >
> > best,
> > Nuno Moreno
>

Arvydas Matiukas

strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hello listers,

Recently a user complained about a square shadow in blue
channel (pseudocolor for far red). Below is the link
to the image
http://imgur.com/422pyYS
This image is a frame-sequential scan of three colors (GFP, RFP, and Cy5).
Three different lasers (488, 543, 633) were used for the excitation,
but only two detectors (the same one for RFP, and Cy5). I was not able
to reproduce this shadow artefact by reusing original settings. The user
claims that after the appearance the shadow in blue channel stays until
software and/or hardware is restarted (other channels/detectors are not affected).
The artefact was observed at least twice on different days.
I never saw anything similar during ten years of confocal imaging.

Now the system is off contract so I would prefer to identify myself
the cause of this weird behavior. Any comments and insights are
highly appreciated.

Thanks,
Arvydas
======================

Arvydas Matiukas, Ph.D.
Director of Confocal&Two-Photon Core
SUNY Upstate Medical University

mmodel

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Arvydas,
Are you talking about the absence of visible nuclei in the top right corner? The first thing I would do is collect a transmission image using the same laser to see if something is blocking excitation.

Mike

-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Arvydas Matiukas
Sent: Thursday, August 21, 2014 2:52 PM
To: [hidden email]
Subject: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hello listers,

Recently a user complained about a square shadow in blue channel (pseudocolor for far red). Below is the link to the image http://imgur.com/422pyYS This image is a frame-sequential scan of three colors (GFP, RFP, and Cy5).
Three different lasers (488, 543, 633) were used for the excitation, but only two detectors (the same one for RFP, and Cy5). I was not able to reproduce this shadow artefact by reusing original settings. The user claims that after the appearance the shadow in blue channel stays until software and/or hardware is restarted (other channels/detectors are not affected).
The artefact was observed at least twice on different days.
I never saw anything similar during ten years of confocal imaging.

Now the system is off contract so I would prefer to identify myself the cause of this weird behavior. Any comments and insights are highly appreciated.

Thanks,
Arvydas
======================

Arvydas Matiukas, Ph.D.
Director of Confocal&Two-Photon Core
SUNY Upstate Medical University

John Oreopoulos

Re: strange shadow in one ch

In reply to this post by Arvydas Matiukas

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Could that be a previously photobleached region of the sample in the blue channel, in a slightly different field of view? Or maybe the user used the scan zoom in that area previously and photobleached, then zoomed out to a larger field of view?

John Oreopoulos
Staff Scientist
Spectral Applied Research Inc.
A Division of Andor Technology
Richmond Hill, Ontario
Canada
www.spectral.ca

On 2014-08-21, at 2:52 PM, Arvydas Matiukas wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Hello listers,
>
> Recently a user complained about a square shadow in blue
> channel (pseudocolor for far red). Below is the link
> to the image
> http://imgur.com/422pyYS
> This image is a frame-sequential scan of three colors (GFP, RFP, and Cy5).
> Three different lasers (488, 543, 633) were used for the excitation,
> but only two detectors (the same one for RFP, and Cy5). I was not able
> to reproduce this shadow artefact by reusing original settings. The user
> claims that after the appearance the shadow in blue channel stays until
> software and/or hardware is restarted (other channels/detectors are not affected).
> The artefact was observed at least twice on different days.
> I never saw anything similar during ten years of confocal imaging.
>
> Now the system is off contract so I would prefer to identify myself
> the cause of this weird behavior. Any comments and insights are
> highly appreciated.
>
> Thanks,
> Arvydas
> ======================
>
>
>
>
>
>
>
>
> Arvydas Matiukas, Ph.D.
> Director of Confocal&Two-Photon Core
> SUNY Upstate Medical University

Arvydas Matiukas

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Thanks John, the suggestion about previously photobleached region is quite likely,
and yes, scan zoom was varied during imaging (final zoom=2) .
It also would explain why in another slide (not shown) the square shadow
does not completely blank the signal and is at shifted position.

Good news then would be that the shadow is not related to hardware malfunction.
I will ask the user about her samples and how easily they can be photobleached.
Arvydas

>>> John Oreopoulos <[hidden email]> 8/21/2014 3:19 PM >>>
*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Could that be a previously photobleached region of the sample in the blue channel, in a slightly different field of view? Or maybe the user used the scan zoom in that area previously and photobleached, then zoomed out to a larger field of view?

John Oreopoulos
Staff Scientist
Spectral Applied Research Inc.
A Division of Andor Technology
Richmond Hill, Ontario
Canada
www.spectral.ca

On 2014-08-21, at 2:52 PM, Arvydas Matiukas wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Hello listers,
>
> Recently a user complained about a square shadow in blue
> channel (pseudocolor for far red). Below is the link
> to the image
> http://imgur.com/422pyYS
> This image is a frame-sequential scan of three colors (GFP, RFP, and Cy5).
> Three different lasers (488, 543, 633) were used for the excitation,
> but only two detectors (the same one for RFP, and Cy5). I was not able
> to reproduce this shadow artefact by reusing original settings. The user
> claims that after the appearance the shadow in blue channel stays until
> software and/or hardware is restarted (other channels/detectors are not affected).
> The artefact was observed at least twice on different days.
> I never saw anything similar during ten years of confocal imaging.
>
> Now the system is off contract so I would prefer to identify myself
> the cause of this weird behavior. Any comments and insights are
> highly appreciated.
>
> Thanks,
> Arvydas
> ======================
>
>
>
>
>
>
>
>
> Arvydas Matiukas, Ph.D.
> Director of Confocal&Two-Photon Core
> SUNY Upstate Medical University

Craig Brideau

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

We work with fairly photosensitive dyes, and our users have gotten used to
finding faint rectangles in their sample. If the dyes in your sample don't
have a particular reputation for photosensitivity your user may be
illuminating with too much power or dwell time. Sometimes antifade mounting
media and the like can help a little bit.

Craig

On Thu, Aug 21, 2014 at 2:44 PM, Arvydas Matiukas <[hidden email]>
wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Thanks John, the suggestion about previously photobleached region is quite
> likely,
> and yes, scan zoom was varied during imaging (final zoom=2) .
> It also would explain why in another slide (not shown) the square shadow
> does not completely blank the signal and is at shifted position.
>
> Good news then would be that the shadow is not related to hardware
> malfunction.
> I will ask the user about her samples and how easily they can be
> photobleached.
> Arvydas
>
> >>> John Oreopoulos <[hidden email]> 8/21/2014 3:19 PM >>>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Could that be a previously photobleached region of the sample in the blue
> channel, in a slightly different field of view? Or maybe the user used the
> scan zoom in that area previously and photobleached, then zoomed out to a
> larger field of view?
>
> John Oreopoulos
> Staff Scientist
> Spectral Applied Research Inc.
> A Division of Andor Technology
> Richmond Hill, Ontario
> Canada
> www.spectral.ca
>
>
> On 2014-08-21, at 2:52 PM, Arvydas Matiukas wrote:
>
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Hello listers,
> >
> > Recently a user complained about a square shadow in blue
> > channel (pseudocolor for far red). Below is the link
> > to the image
> > http://imgur.com/422pyYS
> > This image is a frame-sequential scan of three colors (GFP, RFP, and
> Cy5).
> > Three different lasers (488, 543, 633) were used for the excitation,
> > but only two detectors (the same one for RFP, and Cy5). I was not able
> > to reproduce this shadow artefact by reusing original settings. The user
> > claims that after the appearance the shadow in blue channel stays until
> > software and/or hardware is restarted (other channels/detectors are not
> affected).
> > The artefact was observed at least twice on different days.
> > I never saw anything similar during ten years of confocal imaging.
> >
> > Now the system is off contract so I would prefer to identify myself
> > the cause of this weird behavior. Any comments and insights are
> > highly appreciated.
> >
> > Thanks,
> > Arvydas
> > ======================
> >
> >
> >
> >
> >
> >
> >
> >
> > Arvydas Matiukas, Ph.D.
> > Director of Confocal&Two-Photon Core
> > SUNY Upstate Medical University
>

Jeff Reece

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

If you didn't notice yet, the other channels show slightly reduced signal for the same rectangular area. Perhaps the user was "live-scanning" the blue channel when the phone range, then switched to another channel, shifted the field of view a bit and snapped the image there.

-Jeff

>________________________________
> From: Craig Brideau <[hidden email]>
>To: [hidden email]
>Sent: Thursday, August 21, 2014 7:11 PM
>Subject: Re: strange shadow in one ch
>
>
>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>Post images on http://www.imgur.com and include the link in your posting.
>*****
>
>We work with fairly photosensitive dyes, and our users have gotten used to
>finding faint rectangles in their sample. If the dyes in your sample don't
>have a particular reputation for photosensitivity your user may be
>illuminating with too much power or dwell time. Sometimes antifade mounting
>media and the like can help a little bit.
>
>Craig
>
>
>
>
>
>
>On Thu, Aug 21, 2014 at 2:44 PM, Arvydas Matiukas <[hidden email]>
>wrote:
>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> Post images on http://www.imgur.com and include the link in your posting.
>> *****
>>
>> Thanks John, the suggestion about previously photobleached region is quite
>> likely,
>> and yes, scan zoom was varied during imaging (final zoom=2) .
>> It also would explain why in another slide (not shown) the square shadow
>> does not completely blank the signal and is at shifted position.
>>
>> Good news then would be that the shadow is not related to hardware
>> malfunction.
>> I will ask the user about her samples and how easily they can be
>> photobleached.
>> Arvydas
>>
>> >>> John Oreopoulos <[hidden email]> 8/21/2014 3:19 PM >>>
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> Post images on http://www.imgur.com and include the link in your posting.
>> *****
>>
>> Could that be a previously photobleached region of the sample in the blue
>> channel, in a slightly different field of view? Or maybe the user used the
>> scan zoom in that area previously and photobleached, then zoomed out to a
>> larger field of view?
>>
>> John Oreopoulos
>> Staff Scientist
>> Spectral Applied Research Inc.
>> A Division of Andor Technology
>> Richmond Hill, Ontario
>> Canada
>> www.spectral.ca
>>
>>
>> On 2014-08-21, at 2:52 PM, Arvydas Matiukas wrote:
>>
>> > *****
>> > To join, leave or search the confocal microscopy listserv, go to:
>> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> > Post images on http://www.imgur.com and include the link in your
>> posting.
>> > *****
>> >
>> > Hello listers,
>> >
>> > Recently a user complained about a square shadow in blue
>> > channel (pseudocolor for far red). Below is the link
>> > to the image
>> > http://imgur.com/422pyYS
>> > This image is a frame-sequential scan of three colors (GFP, RFP, and
>> Cy5).
>> > Three different lasers (488, 543, 633) were used for the excitation,
>> > but only two detectors (the same one for RFP, and Cy5). I was not able
>> > to reproduce this shadow artefact by reusing original settings. The user
>> > claims that after the appearance the shadow in blue channel stays until
>> > software and/or hardware is restarted (other channels/detectors are not
>> affected).
>> > The artefact was observed at least twice on different days.
>> > I never saw anything similar during ten years of confocal imaging.
>> >
>> > Now the system is off contract so I would prefer to identify myself
>> > the cause of this weird behavior. Any comments and insights are
>> > highly appreciated.
>> >
>> > Thanks,
>> > Arvydas
>> > ======================
>> >
>> >
>> >
>> >
>> >
>> >
>> >
>> >
>> > Arvydas Matiukas, Ph.D.
>> > Director of Confocal&Two-Photon Core
>> > SUNY Upstate Medical University
>>
>
>
>

Feinstein, Timothy

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hi Jeff,

Not necessarily the case. People often use 405 nm to bleach everything in
a ROI. Blue-violet light has enough energy to nuke things well outside of
their typical absorption peak. I have found that 488 nm will bleach green
and most labile fluorophores further to the red, 561 nm will bleach red
and far red (though most far red dyes are pretty resistant) and so on. I
would consider a slight dimming in other channels as a good sign that they
just used too much blue.

Cheers,

TF

Timothy Feinstein, Ph.D. | Confocal Manager
333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
Phone: 616-234-5819 | Email: [hidden email]

On 8/22/14, 7:48 AM, "Jeff Reece" <[hidden email]> wrote:

>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQdZw
>68T_VDA&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalmic
>roscopy
>Post images on
>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQdck
>7ozWBXA&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>posting.
>*****
>
>If you didn't notice yet, the other channels show slightly reduced signal
>for the same rectangular area. Perhaps the user was "live-scanning" the
>blue channel when the phone range, then switched to another channel,
>shifted the field of view a bit and snapped the image there.
>
>-Jeff
>
>
>
>>________________________________
>> From: Craig Brideau <[hidden email]>
>>To: [hidden email]
>>Sent: Thursday, August 21, 2014 7:11 PM
>>Subject: Re: strange shadow in one ch
>>
>>
>>*****
>>To join, leave or search the confocal microscopy listserv, go to:
>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQdZ
>>w68T_VDA&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfocalm
>>icroscopy
>>Post images on
>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQdc
>>k7ozWBXA&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>posting.
>>*****
>>
>>We work with fairly photosensitive dyes, and our users have gotten used
>>to
>>finding faint rectangles in their sample. If the dyes in your sample
>>don't
>>have a particular reputation for photosensitivity your user may be
>>illuminating with too much power or dwell time. Sometimes antifade
>>mounting
>>media and the like can help a little bit.
>>
>>Craig
>>
>>
>>
>>
>>
>>
>>On Thu, Aug 21, 2014 at 2:44 PM, Arvydas Matiukas <[hidden email]>
>>wrote:
>>
>>> *****
>>> To join, leave or search the confocal microscopy listserv, go to:
>>>
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>Zw68T_VDA&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfoca
>>>lmicroscopy
>>> Post images on
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>ck7ozWBXA&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>>posting.
>>> *****
>>>
>>> Thanks John, the suggestion about previously photobleached region is
>>>quite
>>> likely,
>>> and yes, scan zoom was varied during imaging (final zoom=2) .
>>> It also would explain why in another slide (not shown) the square
>>>shadow
>>> does not completely blank the signal and is at shifted position.
>>>
>>> Good news then would be that the shadow is not related to hardware
>>> malfunction.
>>> I will ask the user about her samples and how easily they can be
>>> photobleached.
>>> Arvydas
>>>
>>> >>> John Oreopoulos <[hidden email]> 8/21/2014 3:19 PM >>>
>>> *****
>>> To join, leave or search the confocal microscopy listserv, go to:
>>>
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>Zw68T_VDA&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfoca
>>>lmicroscopy
>>> Post images on
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>ck7ozWBXA&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>>posting.
>>> *****
>>>
>>> Could that be a previously photobleached region of the sample in the
>>>blue
>>> channel, in a slightly different field of view? Or maybe the user used
>>>the
>>> scan zoom in that area previously and photobleached, then zoomed out
>>>to a
>>> larger field of view?
>>>
>>> John Oreopoulos
>>> Staff Scientist
>>> Spectral Applied Research Inc.
>>> A Division of Andor Technology
>>> Richmond Hill, Ontario
>>> Canada
>>>
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>ZM_9mSGDg&u=http%3a%2f%2fwww%2espectral%2eca
>>>
>>>
>>> On 2014-08-21, at 2:52 PM, Arvydas Matiukas wrote:
>>>
>>> > *****
>>> > To join, leave or search the confocal microscopy listserv, go to:
>>> >
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>Zw68T_VDA&u=http%3a%2f%2flists%2eumn%2eedu%2fcgi-bin%2fwa%3fA0%3dconfoca
>>>lmicroscopy
>>> > Post images on
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>ck7ozWBXA&u=http%3a%2f%2fwww%2eimgur%2ecom and include the link in your
>>> posting.
>>> > *****
>>> >
>>> > Hello listers,
>>> >
>>> > Recently a user complained about a square shadow in blue
>>> > channel (pseudocolor for far red). Below is the link
>>> > to the image
>>> >
>>>http://scanmail.trustwave.com/?c=129&d=ha_30_eAhDRuscaXfT00aoYDawLfVmiQd
>>>Zo9q2XTDw&u=http%3a%2f%2fimgur%2ecom%2f422pyYS
>>> > This image is a frame-sequential scan of three colors (GFP, RFP, and
>>> Cy5).
>>> > Three different lasers (488, 543, 633) were used for the excitation,
>>> > but only two detectors (the same one for RFP, and Cy5). I was not
>>>able
>>> > to reproduce this shadow artefact by reusing original settings. The
>>>user
>>> > claims that after the appearance the shadow in blue channel stays
>>>until
>>> > software and/or hardware is restarted (other channels/detectors are
>>>not
>>> affected).
>>> > The artefact was observed at least twice on different days.
>>> > I never saw anything similar during ten years of confocal imaging.
>>> >
>>> > Now the system is off contract so I would prefer to identify myself
>>> > the cause of this weird behavior. Any comments and insights are
>>> > highly appreciated.
>>> >
>>> > Thanks,
>>> > Arvydas
>>> > ======================
>>> >
>>> >
>>> >
>>> >
>>> >
>>> >
>>> >
>>> >
>>> > Arvydas Matiukas, Ph.D.
>>> > Director of Confocal&Two-Photon Core
>>> > SUNY Upstate Medical University
>>>
>>
>>

Reece, Jeff (NIH/NIDDK) [E]

Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hi Tim,

"Perhaps" usually means "not necessarily". :-)

Cheers,
Jeff

________________________________________
From: Feinstein, Timothy [[hidden email]]
Sent: Friday, August 22, 2014 9:30 AM
To: [hidden email]
Subject: Re: strange shadow in one ch

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

Hi Jeff,

Not necessarily the case. People often use 405 nm to bleach everything in
a ROI. Blue-violet light has enough energy to nuke things well outside of
their typical absorption peak. I have found that 488 nm will bleach green
and most labile fluorophores further to the red, 561 nm will bleach red
and far red (though most far red dyes are pretty resistant) and so on. I
would consider a slight dimming in other channels as a good sign that they
just used too much blue.

Cheers,

TF

Timothy Feinstein, Ph.D. | Confocal Manager
333 Bostwick Ave., N.E., Grand Rapids, Michigan 49503
Phone: 616-234-5819 | Email: [hidden email]

On 8/22/14, 7:48 AM, "Jeff Reece" <[hidden email]> wrote:

Stanislav Vitha-2

Re: strange shadow in one ch

In reply to this post by Arvydas Matiukas

*****
To join, leave or search the confocal microscopy listserv, go to:
http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
Post images on http://www.imgur.com and include the link in your posting.
*****

As already noted, that shadow looks like a previously
scanned/photobleached ROI.
Besides antifades, using less laser power and checking that the zoom is not
excessive, I often suggest to my users to do all live view in a fast, low-
resolution mode (skip every other pixel) to preserve the specimen.

Stan

Microscopy and Imaging Center
Texas A&M University

On Thu, 21 Aug 2014 14:52:00 -0400, Arvydas Matiukas
<[hidden email]> wrote:

>*****
>To join, leave or search the confocal microscopy listserv, go to:
>http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>Post images on http://www.imgur.com and include the link in your
posting.

>*****
>
>Hello listers,
>
>Recently a user complained about a square shadow in blue
>channel (pseudocolor for far red). Below is the link
>to the image
>http://imgur.com/422pyYS
>This image is a frame-sequential scan of three colors (GFP, RFP, and Cy5).
>Three different lasers (488, 543, 633) were used for the excitation,
>but only two detectors (the same one for RFP, and Cy5). I was not able
>to reproduce this shadow artefact by reusing original settings. The user
>claims that after the appearance the shadow in blue channel stays until
>software and/or hardware is restarted (other channels/detectors are not

affected).

>The artefact was observed at least twice on different days.
>I never saw anything similar during ten years of confocal imaging.
>
>Now the system is off contract so I would prefer to identify myself
>the cause of this weird behavior. Any comments and insights are
>highly appreciated.
>
>Thanks,
>Arvydas
>======================
>
>
>
>
>
>
>
>
>Arvydas Matiukas, Ph.D.
>Director of Confocal&Two-Photon Core
>SUNY Upstate Medical University