Polarized light in a Zeiss LSM510 Meta
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Carlos Sanchez-8 |
Polarized light in a Zeiss LSM510 Meta
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Search the CONFOCAL archive at
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Hello everybody.
We would like to know your expert opinion about a problem with polarized transmitted light which we have been observing in our Zeiss LSM510 Meta confocal system since Summer-2007.
Please, download examples from: http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madrid_2008.zip
The Zeiss database include three “*.lsm” files with some timelapse images taken with the transmission detector and using the 488nm laser line in channel 1, the 543nm one in channel two and 633nm in the third one: 1- METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the polarizer (never the analyzer since lasers are already polarized) inserted in the condenser, 2- withoutpolarizer.lsm- exactly the same as before but without inserting the polarizer, 3- Withpolarizerinnewconfocal0.lsm- the same, inserting the polarizer, but in another newer Zeiss LSM510 confocal system.
As you will see the signal is rather stable in samples 2 and 3 with all laser lines. However, when the polarizer is inserted in the condenser of the Zeiss LSM510 Meta system we get fluctuations with slow cycles of around two minutes. Zeiss technicians changed the optical fiber and aligned lasers with the fiber several times. In fact they were able to solve some additional strange line pattern got with the 543nm laser line but slow light fluctuations still persist. We would be very pleased if you could send us your ideas, impressions, similar experiences. Thank you very much in advance for your help and time. Regards,
Carlos Sánchez Martín Servicio de Microscopía Optica y Confocal (Lab.310) Centro de Biología Molecular "Severo Ochoa" (CSIC-UAM) C/Nicolás Cabrera,1 Universidad Autónoma de Madrid. Cantoblanco. 28049. Madrid Tlfs. 91 196 4613/4643 Fax. 91 196 4420 E-mail: [hidden email] Web: http://www.cbm.uam.es/confocal
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simon walker (BI) |
Re: Polarized light in a Zeiss LSM510 Meta
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Search the CONFOCAL archive at
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It looks like there is a similar effect
with the other confocal system, it’s just more pronounced on the original
system – perhaps due to the settings on the transmitted light detector. Presumably
these are live cells in some sort of buffer. Are they under flow? Do you see
the same effect when imaging fixed cells? Simon From:
Hello everybody. We
would like to know your expert opinion about a problem with polarized
transmitted light which we have been observing in our Zeiss LSM510 Meta
confocal system since Summer-2007. Please, download examples from: http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madrid_2008.zip The Zeiss database
include three “*.lsm” files with some timelapse images taken with
the transmission detector and using the 488nm laser line in channel 1, the
543nm one in channel two and 633nm in the third one: 1- METAwithpolarizer.lsm-
taken in the LSM510 Meta system with just the polarizer (never the analyzer
since lasers are already polarized) inserted in the condenser, 2- withoutpolarizer.lsm-
exactly the same as before but without inserting the polarizer, 3- Withpolarizerinnewconfocal0.lsm-
the same, inserting the polarizer, but in another newer Zeiss LSM510 confocal
system. As
you will see the signal is rather stable in samples 2 and 3 with all laser
lines. However, when the polarizer is inserted in the condenser of the Zeiss
LSM510 Meta system we get fluctuations with slow cycles of around two minutes.
Zeiss technicians changed the optical fiber and aligned lasers with the fiber
several times. In fact they were able to solve some additional strange line
pattern got with the 543nm laser line but slow light fluctuations still
persist. We would be very pleased
if you could send us your ideas, impressions, similar experiences. Thank you very much in advance for your help and time. Regards, Carlos Sánchez Martín Servicio de Microscopía Optica y Confocal (Lab.310) Centro de Biología Molecular "Severo Ochoa"
(CSIC-UAM) C/Nicolás Cabrera,1 Universidad Autónoma de Madrid. Cantoblanco. 28049. Madrid Tlfs. 91 196 4613/4643 Fax. 91 196 4420 E-mail: [hidden email] Web: http://www.cbm.uam.es/confocal |
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Carol Norris |
Re: Polarized light in a Zeiss LSM510 Meta
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In reply to this post by Carlos Sanchez-8
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
We had a similar problem on our Leica SP2, except that the cycle occurred over a couple of hours. With the 488 line, background intensity in the DIC channel varied from 10 to about 250 and back down; there were similar changes with the 458 line off the same laser. Transmitted light images using the 543 laser over the same time period showed a steady decrease from 200 down to 20. All this happened while fluorescent images showed essentially no variation. Our tech person fixed it by adjusting the polarization of the lasers.
Carol Norris On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Carol E. Norris, Ph.D Facility Scientist Flow Cytometry/Confocal Microscopy Facility Biotechnology/Bioservices Center University of Connecticut Unit 3149 91 N. Eagleville Rd Storrs, CT 06269-3149 Phone (860) 486-3080 Fax (860) 486-5005 |
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Urs Utzinger |
Re: Polarized light in a Zeiss LSM510 Meta
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In reply to this post by Carlos Sanchez-8
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
Temperature fluctuations in your microscopes objective lens are
most likely affecting the polarization state of your laser at the sample. If you couple the laser through a fiber optic cable into the
scan head you might want to check that the output of the cable is polarized and
that it remains stable (measure through linear polarizer on power meter). You could post-process your image and keep the intensity
constant in an “empty” area of your image. Alternatively you could take two images with the linear
polarizer rotating by 90 degree and calculate a composite image. Urs Utzinger University of Arizona From: Confocal Microscopy
List [mailto:[hidden email]] On Behalf Of Carlos Sanchez Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Hello everybody.
We would like to know your expert opinion about a problem with polarized
transmitted light which we have been observing in our Zeiss LSM510 Meta
confocal system since Summer-2007. Please, download examples from: http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madrid_2008.zip The Zeiss
database include three “*.lsm” files with some timelapse images
taken with the transmission detector and using the 488nm laser line in channel
1, the 543nm one in channel two and 633nm in the third one: 1-
METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the polarizer
(never the analyzer since lasers are already polarized) inserted in the
condenser, 2-
withoutpolarizer.lsm- exactly the same as before but
without inserting the polarizer, 3-
Withpolarizerinnewconfocal0.lsm- the same, inserting
the polarizer, but in another newer Zeiss LSM510 confocal system.
As you will see the signal is rather stable in samples 2 and 3 with all laser
lines. However, when the polarizer is inserted in the condenser of the Zeiss
LSM510 Meta system we get fluctuations with slow cycles of around two minutes.
Zeiss technicians changed the optical fiber and aligned lasers with the fiber
several times. In fact they were able to solve some additional strange line
pattern got with the 543nm laser line but slow light fluctuations still
persist. We would be very
pleased if you could send us your ideas, impressions, similar experiences. Thank you very much in advance for your
help and time. Regards, Carlos Sánchez Martín Servicio de Microscopía Optica y Confocal
(Lab.310) Centro de Biología Molecular "Severo
Ochoa" (CSIC-UAM) C/Nicolás Cabrera,1 Universidad Autónoma de Madrid. Cantoblanco. 28049. Madrid Tlfs. 91 196 4613/4643 Fax. 91 196 4420 E-mail: [hidden email] Web: http://www.cbm.uam.es/confocal |
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JOEL B. SHEFFIELD |
Re: Polarized light in a Zeiss LSM510 Meta
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In reply to this post by Carol Norris
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal We had a similar situation with our Leica SP1, also with the Argon laser, and fixed by realignment, just as with Carol Joel > Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi- > bin/wa?S1=confocal We had a similar problem on our Leica SP2, except > that the cycle occurred over a couple of hours. With the 488 line, > background intensity in the DIC channel varied from 10 to about 250 > and back down; there were similar changes with the 458 line off the > same laser. Transmitted light images using the 543 laser over the > same time period showed a steady decrease from 200 down to 20. All > this happened while fluorescent images showed essentially no > variation. Our tech person fixed it by adjusting the polarization > of the lasers. > Carol Norris > > > On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: > > Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi- > bin/wa?S1=confocal Hello everybody. > > > > > > -- Joel B. Sheffield, Ph.D. Biology Department, Temple University 1900 North 12th Street Philadelphia, PA 19122 [hidden email] (215) 204 8839, fax (215) 204 0486 http://astro.temple.edu/~jbs |
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Marc Thibault |
Re: Polarized light in a Zeiss LSM510 Meta
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In reply to this post by Carol Norris
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
HI, I have nothing to add but maybe another question: Is there a
preffered laser line for transmitted light DIC images of cells (with no concerns
with photoxicity) ?
I
also have weird intensity variations with channel D (transmitted) on the
LSM.
Thanks
Marc
------------------------------------------------ De : Carol Norris [mailto:[hidden email]] Envoyé : 5 février 2008 10:18 À : [hidden email] Objet : Re: Polarized light in a Zeiss LSM510 Meta Carol Norris On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Carol E. Norris, Ph.D Facility Scientist Flow Cytometry/Confocal Microscopy Facility Biotechnology/Bioservices Center University of Connecticut Unit 3149 91 N. Eagleville Rd Storrs, CT 06269-3149 Phone (860) 486-3080 Fax (860) 486-5005 |
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Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
Dear Marc,
they all work well (on a good day at least) and you
can get some interesting information by looking at the transmission of light
through specimens though I believe that the systems are generally set up using
the 488nm line.
One nice effect is detailed in an application note
from Werner Knebel at Leica on "false" "true colour imaging" using sequential
imaging and the transmitted light detector. Not confocal as such, but can give
nice colour relief of different structures in a sample.
all the best
Darran Clements
From: [hidden email]
Sent: Tuesday, February 05, 2008 8:59 PM
To: [hidden email]
Subject: Re: Polarized light in a Zeiss LSM510
Meta HI, I have nothing to add but maybe another question: Is there a
preffered laser line for transmitted light DIC images of cells (with no concerns
with photoxicity) ?
I
also have weird intensity variations with channel D (transmitted) on the
LSM.
Thanks
Marc
------------------------------------------------ De : Carol Norris [mailto:[hidden email]] Envoyé : 5 février 2008 10:18 À : [hidden email] Objet : Re: Polarized light in a Zeiss LSM510 Meta Carol Norris On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Carol E. Norris, Ph.D Facility Scientist Flow Cytometry/Confocal Microscopy Facility Biotechnology/Bioservices Center University of Connecticut Unit 3149 91 N. Eagleville Rd Storrs, CT 06269-3149 Phone (860) 486-3080 Fax (860) 486-5005 |
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Reece, Jeff (NIH/NIEHS) [V] |
Re: Polarized light in a Zeiss LSM510 Meta
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In reply to this post by Carlos Sanchez-8
Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Better resolution at shorter shorter wavelengths, but when working with thicker tissue slices, results are better at longer wavelengths because of dispersion. Cheers, Jeff ________________________________ From: Marc Thibault [mailto:[hidden email]] Sent: Tue 2/5/2008 3:59 PM To: [hidden email] Subject: Re: Polarized light in a Zeiss LSM510 Meta ---------------------- Information from the mail header ----------------------- Sender: Confocal Microscopy List <[hidden email]> Poster: Marc Thibault <[hidden email]> Subject: Re: Polarized light in a Zeiss LSM510 Meta ------------------------------------------------------------------------------- This is a multi-part message in MIME format. ------=_NextPart_000_0007_01C86810.13E46600 Content-Type: text/plain; charset="iso-8859-1" Content-Transfer-Encoding: quoted-printable Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal HI, I have nothing to add but maybe another question: Is there a = preffered laser line for transmitted light DIC images of cells (with no concerns = with photoxicity) ? I also have weird intensity variations with channel D (transmitted) on = the LSM. =20 Thanks =20 Marc =20 ------------------------------------------------ Marc Thibault, Ph.D. Postdoctoral Fellow NSERC/Biosynthech Chair in Hybrid Biomaterials for Innovative Regenerative Technologies Department of Chemical Engineering Ecole Polytechnique Montreal, Qc, Canada 514 340 4711 (4781) ------------------------------------------------ =20 _____ =20 De : Carol Norris [mailto:[hidden email]]=20 Envoy=E9 : 5 f=E9vrier 2008 10:18 =C0 : [hidden email] Objet : Re: Polarized light in a Zeiss LSM510 Meta Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal We had a = similar problem on our Leica SP2, except that the cycle occurred over a couple = of hours. With the 488 line, background intensity in the DIC channel = varied from 10 to about 250 and back down; there were similar changes with the = 458 line off the same laser. Transmitted light images using the 543 laser = over the same time period showed a steady decrease from 200 down to 20. All = this happened while fluorescent images showed essentially no variation. Our tech person fixed it by adjusting the polarization of the lasers. =20 Carol Norris On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal=20 Hello everybody. =20 We would like to know your expert opinion about a problem = with polarized transmitted light which we have been observing in our Zeiss = LSM510 Meta confocal system since Summer-2007. =20 Please, download examples from: http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madrid_2008.= zip =20 The Zeiss database include three =93*.lsm=94 files with some timelapse = images taken with the transmission detector and using the 488nm laser line in channel 1, the 543nm one in channel two and 633nm in the third one: 1- METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the polarizer (never the analyzer since lasers are already polarized) = inserted in the condenser, 2- withoutpolarizer.lsm- exactly the same as before but without inserting the polarizer, 3- Withpolarizerinnewconfocal0.lsm- the same, inserting the = polarizer, but in another newer Zeiss LSM510 confocal system. =20 As you will see the signal is rather stable in samples 2 and = 3 with all laser lines. However, when the polarizer is inserted in the condenser of the Zeiss LSM510 Meta system we get fluctuations with slow cycles of around two minutes. Zeiss technicians changed the optical = fiber and aligned lasers with the fiber several times. In fact they were able = to solve some additional strange line pattern got with the 543nm laser line = but slow light fluctuations still persist. We would be very pleased if you could send us your ideas, impressions, similar experiences. Thank you very much in advance for your help and time. Regards, =20 Carlos S=E1nchez Mart=EDn Servicio de Microscop=EDa Optica y Confocal (Lab.310) Centro de Biolog=EDa Molecular "Severo Ochoa" (CSIC-UAM) C/Nicol=E1s Cabrera,1 Universidad Aut=F3noma de Madrid. Cantoblanco. 28049. Madrid Tlfs. 91 196 4613/4643 Fax. 91 196 4420 E-mail: [hidden email] Web: http://www.cbm.uam.es/confocal =20 Carol E. Norris, Ph.D Facility Scientist Flow Cytometry/Confocal Microscopy Facility Biotechnology/Bioservices Center University of Connecticut Unit 3149 91 N. Eagleville Rd Storrs, CT 06269-3149 Phone (860) 486-3080 Fax (860) 486-5005 ------=_NextPart_000_0007_01C86810.13E46600 Content-Type: text/html; charset="iso-8859-1" Content-Transfer-Encoding: quoted-printable Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal <!DOCTYPE HTML PUBLIC "-//W3C//DTD HTML 4.0 Transitional//EN"> <HTML><HEAD> <META http-equiv=3DContent-Type content=3D"text/html; = charset=3Diso-8859-1"> <META content=3D"MSHTML 6.00.6000.16544" name=3DGENERATOR></HEAD> <BODY=20 style=3D"WORD-WRAP: break-word; -webkit-nbsp-mode: space; = -webkit-line-break: after-white-space"> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial=20 size=3D2>HI, I have nothing to add but maybe another question: Is there = a=20 preffered laser line for transmitted light DIC images of cells (with no = concerns=20 with photoxicity) ?</FONT></SPAN></DIV> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial size=3D2>I=20 also have weird intensity variations with channel D (transmitted) on the = LSM.</FONT></SPAN></DIV> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial=20 size=3D2></FONT></SPAN> </DIV> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial=20 size=3D2>Thanks</FONT></SPAN></DIV> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial=20 size=3D2></FONT></SPAN> </DIV> <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = face=3DArial=20 size=3D2>Marc</FONT></SPAN></DIV> <DIV> </DIV><!-- Converted from text/plain format --> <P><FONT = size=3D2>------------------------------------------------<BR>Marc=20 Thibault, Ph.D.<BR><BR>Postdoctoral Fellow<BR>NSERC/Biosynthech Chair in = Hybrid=20 Biomaterials<BR>for Innovative Regenerative Technologies<BR>Department = of=20 Chemical Engineering<BR>Ecole Polytechnique<BR>Montreal, Qc, = Canada<BR>514 340=20 4711=20 (4781)<BR>------------------------------------------------<BR><BR><BR></F= ONT></P> <DIV> </DIV><BR> <DIV class=3DOutlookMessageHeader lang=3Dfr dir=3Dltr align=3Dleft> <HR tabIndex=3D-1> <FONT face=3DTahoma size=3D2><B>De :</B> Carol Norris=20 [mailto:[hidden email]] <BR><B>Envoy=E9 :</B> 5 f=E9vrier = 2008=20 10:18<BR><B>=C0 :</B> = [hidden email]<BR><B>Objet :</B>=20 Re: Polarized light in a Zeiss LSM510 Meta<BR></FONT><BR></DIV> <DIV></DIV>Search the CONFOCAL archive at=20 http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal We had a = similar problem=20 on our Leica SP2, except that the cycle occurred over a couple of hours. = With the 488 line, background intensity in the DIC channel varied = from 10=20 to about 250 and back down; there were similar changes with the 458 line = off the=20 same laser. Transmitted light images using the 543 laser over the same = time=20 period showed a steady decrease from 200 down to 20. All this = happened=20 while fluorescent images showed essentially no variation. Our = tech person=20 fixed it by adjusting the polarization of the lasers. <DIV><BR class=3Dwebkit-block-placeholder></DIV> <DIV>Carol Norris<BR> <DIV><BR class=3Dwebkit-block-placeholder></DIV> <DIV><BR> <DIV> <DIV>On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote:</DIV><BR=20 class=3DApple-interchange-newline> <BLOCKQUOTE type=3D"cite"><SPAN class=3DApple-style-span=20 style=3D"WORD-SPACING: 0px; FONT: 12px Helvetica; TEXT-TRANSFORM: = none; COLOR: rgb(0,0,0); TEXT-INDENT: 0px; WHITE-SPACE: normal; = LETTER-SPACING: normal; BORDER-COLLAPSE: separate; orphans: 2; widows: = 2; -webkit-border-horizontal-spacing: 0px; = -webkit-border-vertical-spacing: 0px; = -webkit-text-decorations-in-effect: none; -webkit-text-size-adjust: = auto; -webkit-text-stroke-width: 0">Search=20 the CONFOCAL archive at <A=20 = href=3D"http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal">http:/= /listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal</A> <DIV class=3DSection1> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">Hello=20 everybody.</SPAN></FONT></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: = 12pt"> = We would like to know your expert opinion about a problem with = polarized=20 transmitted light which we have been observing in our Zeiss LSM510 = Meta=20 confocal system since Summer-2007.</SPAN></FONT></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">Please,=20 download examples from:</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Courier New" size=3D2><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 10pt; FONT-FAMILY: 'Courier New'"><A=20 style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 = href=3D"http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madr= id_2008.zip">http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM= _Madrid_2008.zip</A></SPAN></FONT></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: 35.4pt; = FONT-FAMILY: 'Times New Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">The=20 Zeiss database include three =93*.lsm=94 files with some timelapse = images taken=20 with the transmission detector and using the 488nm laser line in = channel 1,=20 the 543nm one in channel two and 633nm in the third = one:</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">1-=20 METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the = polarizer=20 (never the analyzer since lasers are already polarized) inserted in = the=20 condenser,</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: -18pt; = FONT-FAMILY: 'Times New Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">2-<FONT=20 face=3D"Times New Roman" size=3D1><SPAN=20 style=3D"FONT: 7pt 'Times New Roman'"> <SPAN=20 = class=3DApple-converted-space> </SPAN></SPAN></FONT></SPAN></FONT><S= PAN=20 lang=3DEN-GB>withoutpolarizer.lsm- exactly the same as before but = without=20 inserting the polarizer,</SPAN></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: -18pt; = FONT-FAMILY: 'Times New Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">3-<FONT=20 face=3D"Times New Roman" size=3D1><SPAN=20 style=3D"FONT: 7pt 'Times New Roman'"> <SPAN=20 = class=3DApple-converted-space> </SPAN></SPAN></FONT></SPAN></FONT><S= PAN=20 lang=3DEN-GB>Withpolarizerinnewconfocal0.lsm- the same, inserting the = polarizer,=20 but in another newer Zeiss LSM510 confocal system.</SPAN></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: = 12pt"> = As you will see the signal is rather stable in samples 2 and 3 with = all laser=20 lines. However, when the polarizer is inserted in the condenser of the = Zeiss=20 LSM510 Meta system we get fluctuations with slow cycles of around two = minutes.=20 Zeiss technicians changed the optical fiber and aligned lasers with = the fiber=20 several times. In fact they were able to solve some additional strange = line=20 pattern got with the 543nm laser line but slow light fluctuations = still=20 persist.</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: 35.4pt; = FONT-FAMILY: 'Times New Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">We=20 would be very pleased if you could send us your ideas, impressions, = similar=20 experiences.</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = style=3D"FONT-SIZE: 12pt">Thank=20 you very much in advance for your help and time.</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN=20 style=3D"FONT-SIZE: 12pt">Regards,</SPAN></FONT></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN=20 style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">Carlos S=E1nchez=20 Mart=EDn</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">Servicio de=20 Microscop=EDa Optica y Confocal (Lab.310)</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">Centro de Biolog=EDa=20 Molecular "Severo Ochoa" (CSIC-UAM)</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">C/Nicol=E1s=20 Cabrera,1</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">Universidad=20 Aut=F3noma de Madrid. Cantoblanco.</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">28049.=20 Madrid</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">Tlfs. 91 196=20 4613/4643</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: 12pt">Fax. = 91 196=20 4420</SPAN></FONT></DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = 12pt">E-mail:<SPAN=20 class=3DApple-converted-space> </SPAN><A=20 style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 = href=3D"mailto:[hidden email]">[hidden email]</A></SPAN></FONT>= </DIV> <DIV=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: 12pt">Web:<SPAN = class=3DApple-converted-space> </SPAN><A=20 style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 = href=3D"http://www.cbm.uam.es/confocal">http://www.cbm.uam.es/confocal</A= ></SPAN></FONT></DIV> <P class=3DMsoNormal=20 style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = Roman'"><FONT=20 face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 style=3D"FONT-SIZE: = 12pt"></SPAN></FONT> </P></DIV></SPAN></BLOCKQUOTE></DIV><BR></DIV><= /DIV><BR><BR> <DIV> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Carol E. Norris, Ph.D</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Facility Scientist</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Flow Cytometry/Confocal Microscopy Facility</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Biotechnology/Bioservices Center</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>University of Connecticut Unit 3149</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>91 N. Eagleville Rd</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Storrs, CT 06269-3149</FONT></P> <P style=3D"MIN-HEIGHT: 14px; MARGIN: 0px; FONT: 12px = Helvetica"><BR></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Phone (860) 486-3080</FONT></P> <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = face=3DHelvetica=20 size=3D3>Fax (860) 486-5005</FONT></P></DIV><BR></BODY></HTML> ------=_NextPart_000_0007_01C86810.13E46600-- |
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Mayandi Sivaguru |
Re: Polarized light in a Zeiss LSM510 Meta
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Search the CONFOCAL archive at
http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Carlos, you can check the polarizer position as to whether it is sitting at 0 degree position or at 70 degrees, the zeiss polarizer has this much span. As you know the position 0 degree leaves only the linearly polarized wavefront either positive or negative polarization would leave either left or righthanded wavefronts together with the linearly polarized fronts. Check whether this make a diffence. Also I think your sample is a fixed convularia test slide from zeiss, I don't know the reason, you should insert the polarizer in the light path for such a thick specimen as though you should use the DIC optics. In case, if this is happening in every other sample even in thin cells then it is a concern. As Jeff mentioned below, I always use DIC using the 633nm, if you note the blips are much less in that channel, because, for a transmitted light DIC image, you might mostly use live cells and you would be better of with long wavelength and 5mW laser than the argon. I have seen this kind of blips, that is what I call in both Biorad and Zeiss systems. Can happen, when the laser anode current goes up and laser power drops (meaning either a tuning or replacement is imminent). If your lasers are pretty new, I think then you need the argon to be tuned to achieve maximum power and the 543 HeNe too. Alternatively, if everything above is good, then the laser signal integrator board in the scan head is heating too much and may need a better ventilation, if this attempt fails, then the integrator board (around $26k USD) in the scan head needs to be replaced (hope you have service contract). This should be done by the Zeiss service engineer. Let us know when you overcome this issue and how, appreciate a summary of response to this thread, would help people in the future. Thanks Shiv Mayandi Sivaguru Microscopy Facility Manager Institute for Genomic Biology University of Illinois at Urbana-Champaign Urbana, IL, 61801 USA > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal > > > Better resolution at shorter shorter wavelengths, but when working with > thicker tissue slices, results are better at longer wavelengths because of > dispersion. > > Cheers, > Jeff > > ________________________________ > > From: Marc Thibault [mailto:[hidden email]] > Sent: Tue 2/5/2008 3:59 PM > To: [hidden email] > Subject: Re: Polarized light in a Zeiss LSM510 Meta > > > > ---------------------- Information from the mail header > ----------------------- > Sender: Confocal Microscopy List <[hidden email]> > Poster: Marc Thibault <[hidden email]> > Subject: Re: Polarized light in a Zeiss LSM510 Meta > ------------------------------------------------------------------------------- > > This is a multi-part message in MIME format. > > ------=_NextPart_000_0007_01C86810.13E46600 > Content-Type: text/plain; > charset="iso-8859-1" > Content-Transfer-Encoding: quoted-printable > > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal > > HI, I have nothing to add but maybe another question: Is there a = > preffered > laser line for transmitted light DIC images of cells (with no concerns = > with > photoxicity) ? > I also have weird intensity variations with channel D (transmitted) on = > the > LSM. > =20 > Thanks > =20 > Marc > =20 > > ------------------------------------------------ > Marc Thibault, Ph.D. > > Postdoctoral Fellow > NSERC/Biosynthech Chair in Hybrid Biomaterials > for Innovative Regenerative Technologies > Department of Chemical Engineering > Ecole Polytechnique > Montreal, Qc, Canada > 514 340 4711 (4781) > ------------------------------------------------ > > > > > =20 > > _____ =20 > > De : Carol Norris [mailto:[hidden email]]=20 > Envoy=E9 : 5 f=E9vrier 2008 10:18 > =C0 : [hidden email] > Objet : Re: Polarized light in a Zeiss LSM510 Meta > > > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal We had a = > similar > problem on our Leica SP2, except that the cycle occurred over a couple = > of > hours. With the 488 line, background intensity in the DIC channel = > varied > from 10 to about 250 and back down; there were similar changes with the = > 458 > line off the same laser. Transmitted light images using the 543 laser = > over > the same time period showed a steady decrease from 200 down to 20. All = > this > happened while fluorescent images showed essentially no variation. Our > tech person fixed it by adjusting the polarization of the lasers. =20 > > Carol Norris > > > > On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote: > > > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal=20 > Hello everybody. > > =20 > > We would like to know your expert opinion about a problem = > with > polarized transmitted light which we have been observing in our Zeiss = > LSM510 > Meta confocal system since Summer-2007. > > =20 > > Please, download examples from: > http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madrid_2008.= > zip > > =20 > > The Zeiss database include three =93*.lsm=94 files with some timelapse = > images > taken with the transmission detector and using the 488nm laser line in > channel 1, the 543nm one in channel two and 633nm in the third one: > 1- METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the > polarizer (never the analyzer since lasers are already polarized) = > inserted > in the condenser, > 2- withoutpolarizer.lsm- exactly the same as before but without > inserting the polarizer, > 3- Withpolarizerinnewconfocal0.lsm- the same, inserting the = > polarizer, > but in another newer Zeiss LSM510 confocal system. > > =20 > > As you will see the signal is rather stable in samples 2 and = > 3 > with all laser lines. However, when the polarizer is inserted in the > condenser of the Zeiss LSM510 Meta system we get fluctuations with slow > cycles of around two minutes. Zeiss technicians changed the optical = > fiber > and aligned lasers with the fiber several times. In fact they were able = > to > solve some additional strange line pattern got with the 543nm laser line = > but > slow light fluctuations still persist. > We would be very pleased if you could send us your ideas, impressions, > similar experiences. > Thank you very much in advance for your help and time. > Regards, > > =20 > > Carlos S=E1nchez Mart=EDn > Servicio de Microscop=EDa Optica y Confocal (Lab.310) > Centro de Biolog=EDa Molecular "Severo Ochoa" (CSIC-UAM) > C/Nicol=E1s Cabrera,1 > Universidad Aut=F3noma de Madrid. Cantoblanco. > 28049. Madrid > Tlfs. 91 196 4613/4643 > Fax. 91 196 4420 > E-mail: [hidden email] > Web: http://www.cbm.uam.es/confocal > > =20 > > > > > Carol E. Norris, Ph.D > > Facility Scientist > > Flow Cytometry/Confocal Microscopy Facility > > Biotechnology/Bioservices Center > > University of Connecticut Unit 3149 > > 91 N. Eagleville Rd > > Storrs, CT 06269-3149 > > > > > Phone (860) 486-3080 > > Fax (860) 486-5005 > > > > ------=_NextPart_000_0007_01C86810.13E46600 > Content-Type: text/html; > charset="iso-8859-1" > Content-Transfer-Encoding: quoted-printable > > Search the CONFOCAL archive at > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal > <!DOCTYPE HTML PUBLIC "-//W3C//DTD HTML 4.0 Transitional//EN"> > <HTML><HEAD> > <META http-equiv=3DContent-Type content=3D"text/html; = > charset=3Diso-8859-1"> > <META content=3D"MSHTML 6.00.6000.16544" name=3DGENERATOR></HEAD> > <BODY=20 > style=3D"WORD-WRAP: break-word; -webkit-nbsp-mode: space; = > -webkit-line-break: after-white-space"> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial=20 > size=3D2>HI, I have nothing to add but maybe another question: Is there = > a=20 > preffered laser line for transmitted light DIC images of cells (with no = > concerns=20 > with photoxicity) ?</FONT></SPAN></DIV> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial size=3D2>I=20 > also have weird intensity variations with channel D (transmitted) on the = > > LSM.</FONT></SPAN></DIV> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial=20 > size=3D2></FONT></SPAN> </DIV> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial=20 > size=3D2>Thanks</FONT></SPAN></DIV> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial=20 > size=3D2></FONT></SPAN> </DIV> > <DIV dir=3Dltr align=3Dleft><SPAN class=3D451365420-05022008><FONT = > face=3DArial=20 > size=3D2>Marc</FONT></SPAN></DIV> > <DIV> </DIV><!-- Converted from text/plain format --> > <P><FONT = > size=3D2>------------------------------------------------<BR>Marc=20 > Thibault, Ph.D.<BR><BR>Postdoctoral Fellow<BR>NSERC/Biosynthech Chair in = > Hybrid=20 > Biomaterials<BR>for Innovative Regenerative Technologies<BR>Department = > of=20 > Chemical Engineering<BR>Ecole Polytechnique<BR>Montreal, Qc, = > Canada<BR>514 340=20 > 4711=20 > (4781)<BR>------------------------------------------------<BR><BR><BR></F= > ONT></P> > <DIV> </DIV><BR> > <DIV class=3DOutlookMessageHeader lang=3Dfr dir=3Dltr align=3Dleft> > <HR tabIndex=3D-1> > <FONT face=3DTahoma size=3D2><B>De :</B> Carol Norris=20 > [mailto:[hidden email]] <BR><B>Envoy=E9 :</B> 5 f=E9vrier = > 2008=20 > 10:18<BR><B>=C0 :</B> = > [hidden email]<BR><B>Objet :</B>=20 > Re: Polarized light in a Zeiss LSM510 Meta<BR></FONT><BR></DIV> > <DIV></DIV>Search the CONFOCAL archive at=20 > http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal We had a = > similar problem=20 > on our Leica SP2, except that the cycle occurred over a couple of hours. = > > With the 488 line, background intensity in the DIC channel varied = > from 10=20 > to about 250 and back down; there were similar changes with the 458 line = > off the=20 > same laser. Transmitted light images using the 543 laser over the same = > time=20 > period showed a steady decrease from 200 down to 20. All this = > happened=20 > while fluorescent images showed essentially no variation. Our = > tech person=20 > fixed it by adjusting the polarization of the lasers. > <DIV><BR class=3Dwebkit-block-placeholder></DIV> > <DIV>Carol Norris<BR> > <DIV><BR class=3Dwebkit-block-placeholder></DIV> > <DIV><BR> > <DIV> > <DIV>On Feb 5, 2008, at 3:25 AM, Carlos Sanchez wrote:</DIV><BR=20 > class=3DApple-interchange-newline> > <BLOCKQUOTE type=3D"cite"><SPAN class=3DApple-style-span=20 > style=3D"WORD-SPACING: 0px; FONT: 12px Helvetica; TEXT-TRANSFORM: = > none; COLOR: rgb(0,0,0); TEXT-INDENT: 0px; WHITE-SPACE: normal; = > LETTER-SPACING: normal; BORDER-COLLAPSE: separate; orphans: 2; widows: = > 2; -webkit-border-horizontal-spacing: 0px; = > -webkit-border-vertical-spacing: 0px; = > -webkit-text-decorations-in-effect: none; -webkit-text-size-adjust: = > auto; -webkit-text-stroke-width: 0">Search=20 > the CONFOCAL archive at <A=20 > = > href=3D"http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal">http:/= > /listserv.acsu.buffalo.edu/cgi-bin/wa?S1=3Dconfocal</A> > <DIV class=3DSection1> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">Hello=20 > everybody.</SPAN></FONT></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: = > 12pt"> = > > We would like to know your expert opinion about a problem with = > polarized=20 > transmitted light which we have been observing in our Zeiss LSM510 = > Meta=20 > confocal system since Summer-2007.</SPAN></FONT></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">Please,=20 > download examples from:</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Courier New" size=3D2><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 10pt; FONT-FAMILY: 'Courier New'"><A=20 > style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 > = > href=3D"http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM_Madr= > id_2008.zip">http://bacterio.cbm.uam.es/confocal/Temp/Polarized_light_CBM= > _Madrid_2008.zip</A></SPAN></FONT></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: 35.4pt; = > FONT-FAMILY: 'Times New Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">The=20 > Zeiss database include three =93*.lsm=94 files with some timelapse = > images taken=20 > with the transmission detector and using the 488nm laser line in = > channel 1,=20 > the 543nm one in channel two and 633nm in the third = > one:</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">1-=20 > METAwithpolarizer.lsm- taken in the LSM510 Meta system with just the = > polarizer=20 > (never the analyzer since lasers are already polarized) inserted in = > the=20 > condenser,</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: -18pt; = > FONT-FAMILY: 'Times New Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">2-<FONT=20 > face=3D"Times New Roman" size=3D1><SPAN=20 > style=3D"FONT: 7pt 'Times New Roman'"> <SPAN=20 > = > class=3DApple-converted-space> </SPAN></SPAN></FONT></SPAN></FONT><S= > PAN=20 > lang=3DEN-GB>withoutpolarizer.lsm- exactly the same as before but = > without=20 > inserting the polarizer,</SPAN></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: -18pt; = > FONT-FAMILY: 'Times New Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">3-<FONT=20 > face=3D"Times New Roman" size=3D1><SPAN=20 > style=3D"FONT: 7pt 'Times New Roman'"> <SPAN=20 > = > class=3DApple-converted-space> </SPAN></SPAN></FONT></SPAN></FONT><S= > PAN=20 > lang=3DEN-GB>Withpolarizerinnewconfocal0.lsm- the same, inserting the = > polarizer,=20 > but in another newer Zeiss LSM510 confocal system.</SPAN></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: = > 12pt"> = > > As you will see the signal is rather stable in samples 2 and 3 with = > all laser=20 > lines. However, when the polarizer is inserted in the condenser of the = > Zeiss=20 > LSM510 Meta system we get fluctuations with slow cycles of around two = > minutes.=20 > Zeiss technicians changed the optical fiber and aligned lasers with = > the fiber=20 > several times. In fact they were able to solve some additional strange = > line=20 > pattern got with the 543nm laser line but slow light fluctuations = > still=20 > persist.</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; TEXT-INDENT: 35.4pt; = > FONT-FAMILY: 'Times New Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">We=20 > would be very pleased if you could send us your ideas, impressions, = > similar=20 > experiences.</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB = > style=3D"FONT-SIZE: 12pt">Thank=20 > you very much in advance for your help and time.</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN=20 > style=3D"FONT-SIZE: 12pt">Regards,</SPAN></FONT></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN=20 > style=3D"FONT-SIZE: 12pt"></SPAN></FONT> </P> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">Carlos S=E1nchez=20 > Mart=EDn</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">Servicio de=20 > Microscop=EDa Optica y Confocal (Lab.310)</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">Centro de Biolog=EDa=20 > Molecular "Severo Ochoa" (CSIC-UAM)</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">C/Nicol=E1s=20 > Cabrera,1</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">Universidad=20 > Aut=F3noma de Madrid. Cantoblanco.</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">28049.=20 > Madrid</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">Tlfs. 91 196=20 > 4613/4643</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: 12pt">Fax. = > 91 196=20 > 4420</SPAN></FONT></DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN style=3D"FONT-SIZE: = > 12pt">E-mail:<SPAN=20 > class=3DApple-converted-space> </SPAN><A=20 > style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 > = > href=3D"mailto:[hidden email]">[hidden email]</A></SPAN></FONT>= > </DIV> > <DIV=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: 12pt">Web:<SPAN = > class=3DApple-converted-space> </SPAN><A=20 > style=3D"COLOR: blue; TEXT-DECORATION: underline"=20 > = > href=3D"http://www.cbm.uam.es/confocal">http://www.cbm.uam.es/confocal</A= >></SPAN></FONT></DIV> > <P class=3DMsoNormal=20 > style=3D"FONT-SIZE: 12pt; MARGIN: 0cm 0cm 0pt; FONT-FAMILY: 'Times New = > Roman'"><FONT=20 > face=3D"Times New Roman" size=3D3><SPAN lang=3DEN-GB=20 > style=3D"FONT-SIZE: = > 12pt"></SPAN></FONT> </P></DIV></SPAN></BLOCKQUOTE></DIV><BR></DIV><= > /DIV><BR><BR> > <DIV> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Carol E. Norris, Ph.D</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Facility Scientist</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Flow Cytometry/Confocal Microscopy Facility</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Biotechnology/Bioservices Center</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>University of Connecticut Unit 3149</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>91 N. Eagleville Rd</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Storrs, CT 06269-3149</FONT></P> > <P style=3D"MIN-HEIGHT: 14px; MARGIN: 0px; FONT: 12px = > Helvetica"><BR></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Phone (860) 486-3080</FONT></P> > <P style=3D"MARGIN: 0px"><FONT style=3D"FONT: 12px Helvetica" = > face=3DHelvetica=20 > size=3D3>Fax (860) 486-5005</FONT></P></DIV><BR></BODY></HTML> > > ------=_NextPart_000_0007_01C86810.13E46600-- > |
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