Southeastern Microscopy Society meeting
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Amanda M. Lawrence |
Southeastern Microscopy Society meeting
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On behalf of the Southeastern Microscopy Society (SEMS) Local
Arrangements and Program Committees I would like to invite you to the SEMS annual meeting which will be held from May 24-26, 2010 at the Francis Marion hotel in historical Charleston, South Carolina. Microscopy Society of America, Microbeam Analysis Society, and SEMS sponsored speakers include: Dr. David Piston (MSA President) * Vanderbilt University Dr. David Joy * University of Tennessee Dr. Paul Kotula * Sandia National Laboratories Dr. Michael Postek * NIST Dr. Elaine Schumacher * McCrone Associates, Inc Dr. John Lemasters * Medical University of South Carolina Dr. Sue Lessner * University of South Carolina Dr. Heather Evans-Anderson * Winthrop University Dr. Anand Ramamurthi * Clemson University Assistance for travel and meeting costs for students competing in the SEMS Ruska Award and technologists is available. There are also opportunities to participate in pre-meeting workshops on Confocal Microscopy and Digital Imaging, a tour of the Robert Bosch Company Facility, and post-meeting tours of the Yorktown and Fort Sumter. For further information on abstract submission, registration, and participation in the above workshops and tours please visit: http://www.southeasternmicroscopy.org/index-2.html and http://www.southeasternmicroscopy.org/meetings_doc/SEMSBro_web.pdf The deadline for abstract submission is Thursday, April 1, 2010. I hope all of you will be able to attend. Sincerely, Robert L. Price President, SEMS |
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(I apologize if this is a second post, I got some strange errors from the first one)
Dear List It's not a confocal question, it's strictly a wide-field question... When people talk about spherical aberration they typically start with a parallel beam falling on an objective, and then, for whatever reason, the objective fails to bring all the rays into a common focus. In wide-field, one has a luminous spot in the object space, and the effect of spherical aberration would be a failure of the objective to collect all the rays into a single parallel beam. Some rays will form a converging (or diverging) cone instead. Then, this cone will be received (I suspect that in the case of a diverging cone, some light may even get lost on its way) by a tube lens and form a blurry spot on the image plane. My question is, Is this situation really equivalent to the standard one considered in all books? (I suspect, it's not because even the distance to the tube lens should make a difference). But I would be very interested in the opinion of those who understand optics better than I do. Thanks! Mike Model Michael Model, Ph.D. Confocal Microscopy, Dpt Biological Sciences, 1275 University Esplanade, Kent State University, Kent, OH 44242 tel. 330-672-2874 |
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Andreas Bruckbauer |
Re: tube lens and spherical aberration
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Just go the other way, from a perfect spot in the image plane to the diffuse spot in the object plane, then you are back to the standard situation. You can always reverse the light path in linear optics.
best wishes Andreas -----Original Message-----
From: MODEL, MICHAEL <[hidden email]> To: [hidden email] Sent: Fri, 26 Feb 2010 20:54 Subject: tube lens and spherical aberration (I apologize if this is a second post, I got some strange errors from the first |
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Andreas, thanks for your response. I understand that the path can be reversed for a ray but it seems to me that the size of the blur in the image will depend, for example, on the distance between the
objective and the tube lens. Which is not part of the standard formula.
Mike
From: Confocal Microscopy List [[hidden email]] On Behalf Of Andreas Bruckbauer [[hidden email]] Sent: Friday, February 26, 2010 5:06 PM To: [hidden email] Subject: Re: tube lens and spherical aberration Just go the other way, from a perfect spot in the image plane to the diffuse spot in the object plane, then you are back to the standard situation. You can always reverse the light path in linear
optics.
best wishes Andreas -----Original Message-----
From: MODEL, MICHAEL <[hidden email]> To: [hidden email] Sent: Fri, 26 Feb 2010 20:54 Subject: tube lens and spherical aberration (I apologize if this is a second post, I got some strange errors from the first |
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Lutz Schaefer |
Re: tube lens and spherical aberration
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In reply to this post by mmodel
Michael,
spherical aberration (SA) is mostly caused by refractive index mismatches between immersion and embedding. Also, using cover slips and immersion media that are different from the manufacturers recommendations will cause more or less SA. Often we model the system between front element and sample with 3 parallel layers of RI. Optimally, the manufacturer has corrected the objective with aberrations of the opposite sign, so your induced SA cancels out. Still, for example whenever the embedding has different RI than the immersion, the best way to have the least amount of SA is to stay close to the cover slip with your sample. Now, if you have no ICS optics (older microscopes) you may have a chance to do some SA compensation by changing that tube length, which will also slightly change the working distance. The same is true for changing other parameters in the 3 layer system. In an ICS you can only do the latter. Although SA depends linear on some of the model parameters, you cant really say that the wave front generally diverges or converges in presence of SA, because that could rather somewhat better explain defocus. Because it is hard to describe in a few simple words how a wave front behaves, one usually uses diffraction theory (e.g. "Principles of optics", Born & Wolf). This is a very well understood, rigorous theory, enabling to quantify complex amplitudes and their polarization, or if you wish just intensities. I would understand, when not all aspects of diffraction theory are detailed in a book that focuses on general microscopy, but then I am not sure what books you were referring to. Regards Lutz ____________________________________ Lutz Schaefer Advanced Imaging Methodology Consultation 16-715 Doon Village Rd. Kitchener, Ontario, N2P 2A2 Phone/Fax: (519) 894 8870 Email: [hidden email] Website: http://home.golden.net/~lschafer/ ____________________________________ -------------------------------------------------- From: "MODEL, MICHAEL" <[hidden email]> Sent: Friday, February 26, 2010 3:54 PM To: <[hidden email]> Subject: tube lens and spherical aberration > (I apologize if this is a second post, I got some strange errors from the > first one) > > Dear List > > It's not a confocal question, it's strictly a wide-field question... When > people talk about spherical aberration they typically start with a > parallel beam falling on an objective, and then, for whatever reason, the > objective fails to bring all the rays into a common focus. In wide-field, > one has a luminous spot in the object space, and the effect of spherical > aberration would be a failure of the objective to collect all the rays > into a single parallel beam. Some rays will form a converging (or > diverging) cone instead. Then, this cone will be received (I suspect that > in the case of a diverging cone, some light may even get lost on its way) > by a tube lens and form a blurry spot on the image plane. My question is, > Is this situation really equivalent to the standard one considered in all > books? (I suspect, it's not because even the distance to the tube lens > should make a difference). But I would be very interested in the opinion > of those who understand optics better than I do. Thanks! > > Mike Model > > Michael Model, Ph.D. > Confocal Microscopy, > Dpt Biological Sciences, > 1275 University Esplanade, > Kent State University, Kent, OH 44242 > tel. 330-672-2874 |
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Shalin Mehta |
Re: tube lens and spherical aberration
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In reply to this post by mmodel
Hi Michael,
I think your misgivings are true. If I understood correctly - you are wondering whether illumination-side spherical aberration is different from imaging-side spherical aberration. I think they are different. Plus they do different things when one uses confocal scanning geometry vs. widefield geometry. Perhaps different sources of spherical aberration can be explained thus:
- The case considered in most of the microscopy text is of RI mismatch between embedding and immersion media in confocal imaging. The mismatch causes the focused illumination spot to be worse than the diffraction-limited hour-glass, reducing the signal that can be collected by PMT after the pin-hole. So in confocal, spherical aberration = bad SNR. Plus, the signal appears to be coming from the wrong place as spherical aberration shifts the peak intensity in the focal volume.
Since in confocal, both illumination and detection perform imaging and we pick-up the intensity in the center of the imaging volume by using a pin-hole --- I wonder what does the PSF of spherically aberrated confocal look like. I think there are papers by Sheppard and Min Gu that need to be consulted.
- In widefield imaging, one uses Kohler type of arrangement to bring light from mercury filament onto the specimen. Otherwise, one will see filament imprinted on the specimen. The source of widefield illumination being incohernet (unlike coherent laser illumination of confocal), the aberrations seen by it on the way to specimen are unimportant. The PSF of the widefield system is strictly that of the imaging path - the 'mere' use of illumination is to excite fluorophores uniformly.
Consequently, as you say, the above RI mismatch will give rise to converging or diverging wavefront from the focal volume between the tube-lens and the objective *in the detection path*. The imaging path's PSF will be therefore have distorted look like the standard spherical aberrated PSF that we see in Born and Wolf or Hecht.
best Shalin On Sat, Feb 27, 2010 at 7:52 AM, MODEL, MICHAEL <[hidden email]> wrote:
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Shalin Mehta |
Re: tube lens and spherical aberration
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And also Jim Pawley's experimental papers and the confocal book should be looked up. best Shalin |
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In reply to this post by mmodel
I don't think that is true. A lot of published versions start out from
an objective forming an image of a point object. Many years ago I got a bit incensed at a diagram in a textbook, purporting to show SA introduced by a coverslip. The rays leaving the coverslip were not parallel to those entering it! I wanted a diagram to show my students, and this obviously wasn't it. So I sat down with pencil, paper and ruler. First of all I drew a perfect lens forming an image of a point. Then I drew a coverslip in the way, looked up the sines of the angles as measured, and constructed the ray paths. Lo and behold, there was the classic caustic curve. (And, need I say, the rays exited the coverslip at the same angle they entered.) So I traced it on OHP film and used it in my lectures thereafter (long before Powerpoint!). And the students were much more willing to take it on board since it was constructed without any complex formulae - nothing more than Snell's Law. Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of MODEL, MICHAEL Sent: Saturday, 27 February 2010 7:55 AM To: [hidden email] Subject: tube lens and spherical aberration (I apologize if this is a second post, I got some strange errors from the first one) Dear List It's not a confocal question, it's strictly a wide-field question... When people talk about spherical aberration they typically start with a parallel beam falling on an objective, and then, for whatever reason, the objective fails to bring all the rays into a common focus. In wide-field, one has a luminous spot in the object space, and the effect of spherical aberration would be a failure of the objective to collect all the rays into a single parallel beam. Some rays will form a converging (or diverging) cone instead. Then, this cone will be received (I suspect that in the case of a diverging cone, some light may even get lost on its way) by a tube lens and form a blurry spot on the image plane. My question is, Is this situation really equivalent to the standard one considered in all books? (I suspect, it's not because even the distance to the tube lens should make a difference). But I would be very interested in the opinion of those who understand optics better than I do. Thanks! Mike Model Michael Model, Ph.D. Confocal Microscopy, Dpt Biological Sciences, 1275 University Esplanade, Kent State University, Kent, OH 44242 tel. 330-672-2874 No virus found in this incoming message. Checked by AVG - www.avg.com Version: 9.0.733 / Virus Database: 271.1.1/2710 - Release Date: 02/26/10 06:57:00 |
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James Pawley |
Re: tube lens and spherical aberration
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In reply to this post by mmodel
Hi Mike,
I agree with Guy. In fact going back to the "mental optical diagram" in your original post, the "blurry spot" that you suggest occurs in the intermediate image plane (and, larger, on your retina) is just the caustic that he describes. This is not only because the rays "fail to come to a common focus," but because they do this in a specific way: the rays from around the periphery of the optic come to a focus that is either closer to it or farther away from from it than the focus of the paraxial rays near the optical axis. However, regarding: "(I suspect, it's not because even the distance to the tube lens should make a difference)", in the case the tube lens is essentially part of the objective. Together, (and only together) they will form an aberration-free image a certain distance behind the tube lens. Wrong tube lens, NG. Wrong distance: NG. In short, one can ONLY design an aberration-free optical system once one has first defined the positions of the object and final image planes (In incoherent imaging such as fluorescence, the optical system is everything between the object and final image plane). This is why we have objectives designed for different magnifications. One could use a 100x objective is a lens with a focal length of about 2 mm. Despite what is written on the barrel, it could be used to form an image that was, say, only 15x larger than the object. But if you positioned the objective to make this occur, the plane at which this this 15x image appears will be near the back of the objective itself and it would be highly aberrated (and aberrated not just because the light had yet to pass through the tube lens. It would even be aberrated if the objective was a fixed-conjugate, rather than an infinity, objective). When I learned microscopy, I was taught that, although SA could be completely corrected in the LM, as all EM (cylindrical) lenses intrinsically had positive SA it could not be corrected in EM. Now it is the EM guys who have corrected SA (by employing multipole lenses) and the LM'ers now have a serious SA problems because they have started to look at thick specimens that have the "wrong" RI. Must be time to retire. Jim Pawley ********************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 223, Zoology Research Building, FAX 608-265-5315 1117 Johnson Ave., Madison, WI, 53706 [hidden email] 3D Microscopy of Living Cells Course, June 12-24, 2010, UBC, Vancouver Canada Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2010 "If it ain't diffraction, it must be statistics." Anon. >I don't think that is true. A lot of published versions start out from >an objective forming an image of a point object. > >Many years ago I got a bit incensed at a diagram in a textbook, >purporting to show SA introduced by a coverslip. The rays leaving the >coverslip were not parallel to those entering it! I wanted a diagram to >show my students, and this obviously wasn't it. > >So I sat down with pencil, paper and ruler. First of all I drew a >perfect lens forming an image of a point. Then I drew a coverslip in >the way, looked up the sines of the angles as measured, and constructed >the ray paths. Lo and behold, there was the classic caustic curve. >(And, need I say, the rays exited the coverslip at the same angle they >entered.) So I traced it on OHP film and used it in my lectures >thereafter (long before Powerpoint!). And the students were much more >willing to take it on board since it was constructed without any complex >formulae - nothing more than Snell's Law. > Guy > >-----Original Message----- >From: Confocal Microscopy List [mailto:[hidden email]] >On Behalf Of MODEL, MICHAEL >Sent: Saturday, 27 February 2010 7:55 AM >To: [hidden email] >Subject: tube lens and spherical aberration > >(I apologize if this is a second post, I got some strange errors from >the first one) > >Dear List > >It's not a confocal question, it's strictly a wide-field question... >When people talk about spherical aberration they typically start with a >parallel beam falling on an objective, and then, for whatever reason, >the objective fails to bring all the rays into a common focus. In >wide-field, one has a luminous spot in the object space, and the effect >of spherical aberration would be a failure of the objective to collect >all the rays into a single parallel beam. Some rays will form a >converging (or diverging) cone instead. Then, this cone will be received >(I suspect that in the case of a diverging cone, some light may even get >lost on its way) by a tube lens and form a blurry spot on the image >plane. My question is, Is this situation really equivalent to the >standard one considered in all books? (I suspect, it's not because even >the distance to the tube lens should make a difference). But I would be >very interested in the opinion of those who understand optics better >than I do. Thanks! > >Mike Model > >Michael Model, Ph.D. >Confocal Microscopy, >Dpt Biological Sciences, >1275 University Esplanade, >Kent State University, Kent, OH 44242 >tel. 330-672-2874 > >No virus found in this incoming message. >Checked by AVG - www.avg.com >Version: 9.0.733 / Virus Database: 271.1.1/2710 - Release Date: 02/26/10 >06:57:00 -- ********************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 223, Zoology Research Building, FAX 608-265-5315 1117 Johnson Ave., Madison, WI, 53706 [hidden email] 3D Microscopy of Living Cells Course, June 12-24, 2010, UBC, Vancouver Canada Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2010 "If it ain't diffraction, it must be statistics." Anon. -- James and Christine Pawley, 21 N. Prospect Ave. Madison, WI, 53726 Phone: 608-238-3953 |
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Thank you, Jim and others who replied to my question! - Mike
-----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of James Pawley Sent: Tuesday, March 02, 2010 11:55 PM To: [hidden email] Subject: Re: tube lens and spherical aberration Hi Mike, I agree with Guy. In fact going back to the "mental optical diagram" in your original post, the "blurry spot" that you suggest occurs in the intermediate image plane (and, larger, on your retina) is just the caustic that he describes. This is not only because the rays "fail to come to a common focus," but because they do this in a specific way: the rays from around the periphery of the optic come to a focus that is either closer to it or farther away from from it than the focus of the paraxial rays near the optical axis. However, regarding: "(I suspect, it's not because even the distance to the tube lens should make a difference)", in the case the tube lens is essentially part of the objective. Together, (and only together) they will form an aberration-free image a certain distance behind the tube lens. Wrong tube lens, NG. Wrong distance: NG. In short, one can ONLY design an aberration-free optical system once one has first defined the positions of the object and final image planes (In incoherent imaging such as fluorescence, the optical system is everything between the object and final image plane). This is why we have objectives designed for different magnifications. One could use a 100x objective is a lens with a focal length of about 2 mm. Despite what is written on the barrel, it could be used to form an image that was, say, only 15x larger than the object. But if you positioned the objective to make this occur, the plane at which this this 15x image appears will be near the back of the objective itself and it would be highly aberrated (and aberrated not just because the light had yet to pass through the tube lens. It would even be aberrated if the objective was a fixed-conjugate, rather than an infinity, objective). When I learned microscopy, I was taught that, although SA could be completely corrected in the LM, as all EM (cylindrical) lenses intrinsically had positive SA it could not be corrected in EM. Now it is the EM guys who have corrected SA (by employing multipole lenses) and the LM'ers now have a serious SA problems because they have started to look at thick specimens that have the "wrong" RI. Must be time to retire. Jim Pawley ********************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 223, Zoology Research Building, FAX 608-265-5315 1117 Johnson Ave., Madison, WI, 53706 [hidden email] 3D Microscopy of Living Cells Course, June 12-24, 2010, UBC, Vancouver Canada Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2010 "If it ain't diffraction, it must be statistics." Anon. >I don't think that is true. A lot of published versions start out from >an objective forming an image of a point object. > >Many years ago I got a bit incensed at a diagram in a textbook, >purporting to show SA introduced by a coverslip. The rays leaving the >coverslip were not parallel to those entering it! I wanted a diagram to >show my students, and this obviously wasn't it. > >So I sat down with pencil, paper and ruler. First of all I drew a >perfect lens forming an image of a point. Then I drew a coverslip in >the way, looked up the sines of the angles as measured, and constructed >the ray paths. Lo and behold, there was the classic caustic curve. >(And, need I say, the rays exited the coverslip at the same angle they >entered.) So I traced it on OHP film and used it in my lectures >thereafter (long before Powerpoint!). And the students were much more >willing to take it on board since it was constructed without any complex >formulae - nothing more than Snell's Law. > Guy > >-----Original Message----- >From: Confocal Microscopy List [mailto:[hidden email]] >On Behalf Of MODEL, MICHAEL >Sent: Saturday, 27 February 2010 7:55 AM >To: [hidden email] >Subject: tube lens and spherical aberration > >(I apologize if this is a second post, I got some strange errors from >the first one) > >Dear List > >It's not a confocal question, it's strictly a wide-field question... >When people talk about spherical aberration they typically start with a >parallel beam falling on an objective, and then, for whatever reason, >the objective fails to bring all the rays into a common focus. In >wide-field, one has a luminous spot in the object space, and the effect >of spherical aberration would be a failure of the objective to collect >all the rays into a single parallel beam. Some rays will form a >converging (or diverging) cone instead. Then, this cone will be received >(I suspect that in the case of a diverging cone, some light may even get >lost on its way) by a tube lens and form a blurry spot on the image >plane. My question is, Is this situation really equivalent to the >standard one considered in all books? (I suspect, it's not because even >the distance to the tube lens should make a difference). But I would be >very interested in the opinion of those who understand optics better >than I do. Thanks! > >Mike Model > >Michael Model, Ph.D. >Confocal Microscopy, >Dpt Biological Sciences, >1275 University Esplanade, >Kent State University, Kent, OH 44242 >tel. 330-672-2874 > >No virus found in this incoming message. >Checked by AVG - www.avg.com >Version: 9.0.733 / Virus Database: 271.1.1/2710 - Release Date: 02/26/10 >06:57:00 -- ********************************************** Prof. James B. Pawley, Ph. 608-263-3147 Room 223, Zoology Research Building, FAX 608-265-5315 1117 Johnson Ave., Madison, WI, 53706 [hidden email] 3D Microscopy of Living Cells Course, June 12-24, 2010, UBC, Vancouver Canada Info: http://www.3dcourse.ubc.ca/ Applications due by March 15, 2010 "If it ain't diffraction, it must be statistics." Anon. -- James and Christine Pawley, 21 N. Prospect Ave. Madison, WI, 53726 Phone: 608-238-3953 |
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James Pawley |
Re: tube lens and spherical aberration
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Thank you, Jim and others who replied to my question! - Mike You are welcome.
I forgot one more point that you might find interesting.
As the proper correction of SA depends on the image plane being a
specific distance behind the tube lens, it follows that one can change
the SA correction by changing this distance. Indeed, in older
microscopes, one could adjust the tube length specifically to give the
best SA correction under whatever imaging conditions were in
use.
Of course, changing the tube length also changes the
magnification and this is presumably the main reason why modern
instruments lack this convenient adjustment.
Cheers,
Jim Pawley
**********************************************
Prof. James B. Pawley, Room 223, Zoology Research Building, 1117 Johnson Ave., Madison, WI, 53706 3D Microscopy of Living Cells Course, June 12-24, 2010, UBC, Vancouver Canada Info:
http://www.3dcourse.ubc.ca/
-----Original Message----- Info: http://www.3dcourse.ubc.ca/ -- |
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