*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques |
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** It would be nice to see such spectra on a logarithmic scale so that they could be compared to the optical density (OD) blocking performance of various filters. John Oreopoulos Research Assistant Spectral Applied Research Richmond Hill, Ontario Canada www.spectral.ca On 2012-02-24, at 8:35 AM, Guy Cox wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? > > Guy > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) > Sent: Friday, 24 February 2012 1:49 PM > To: [hidden email] > Subject: Re: LEDs > > "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." > > It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. > C > > > Carl A. Boswell > 520-742-6131 > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross > Sent: Thursday, February 23, 2012 6:14 PM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). > From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. > If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. > > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Julio Vazquez > Sent: Friday, 24 February 2012 4:14 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hello all, > > Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. > > Julio. > > == > > > On Feb 23, 2012, at 2:06 AM, jacques wrote: > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> Thanks for your help and comments ! >> >> Jacques |
In reply to this post by Guy Cox-2
Hi Guy,
My reference to lack of spectra came from the online documentation for API's InsightSSI http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no spectra) and X-Cite's XLED1's http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012.pdf separate LED spectra. Lumencor's SOLA light engine brochure http://www.lumencor.com/products/sola_product_sheet.html is the closest I've seen to a useable spectrum. In fact, thanks to Lumencor's openness the SOLA spectrum is on the database, www.spectra.arizona.edu. With the ability to juxtapose (and download) this spectrum with spectra from the listed filters, mirrors and labels, the value to the imaging community of making these light source spectra available becomes more obvious. Cheers, C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: Friday, February 24, 2012 6:36 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques |
In reply to this post by John Oreopoulos
I forgot to mention that www.spectra.arizona.edu has the option to display the data on a log scale.
C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of John Oreopoulos Sent: Friday, February 24, 2012 8:40 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** It would be nice to see such spectra on a logarithmic scale so that they could be compared to the optical density (OD) blocking performance of various filters. John Oreopoulos Research Assistant Spectral Applied Research Richmond Hill, Ontario Canada www.spectral.ca On 2012-02-24, at 8:35 AM, Guy Cox wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? > > Guy > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) > Sent: Friday, 24 February 2012 1:49 PM > To: [hidden email] > Subject: Re: LEDs > > "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." > > It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. > C > > > Carl A. Boswell > 520-742-6131 > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross > Sent: Thursday, February 23, 2012 6:14 PM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). > From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. > If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. > > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Julio Vazquez > Sent: Friday, 24 February 2012 4:14 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hello all, > > Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. > > Julio. > > == > > > On Feb 23, 2012, at 2:06 AM, jacques wrote: > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> Thanks for your help and comments ! >> >> Jacques |
In reply to this post by Guy Cox-2
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** ***COMMERCIAL RESPONSE*** Dear All, X-Cite has a fully configurable LED system, the X-Cite XLED1. Our spectra and module options are freely available here: http://www.ldgi-xcite.com/products-led-modules.php. A word of caution to those located in North America: When looking at LED systems, you may want to ensure that your supplier is paying the legally required licensing fee associated with the LED patent. Some light source manufacturers avoid use of the word LED even though the source of light is still very much an Light Emitting Diode. As always, please contact me offline if there is any other information I can help you with. Best, Kavita Dr. Kavita Aswani, PhD Senior Applications Scientist - Life Sciences LUMEN DYNAMICS GROUP INC.(formerly EXFO) 2260 Argentia Road, Mississauga, Ontario, Canada, L5N 6H7 Direct Tel: 905-812-3342 | Cell: 647 290-3506 | Toll Free: (USA and Canada): 1 800 668-8752 Email: [hidden email] | Website: www.LDGI.com Cheers, Kavita -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: February-24-12 8:36 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques Think Green Before You Print! This email and any attachments thereto may contain private, confidential, and privileged material for the sole use of the intended recipient. Any review, copying, or distribution of this email (or any attachments thereto) by others is strictly prohibited. If you are not the intended recipient, please contact the sender immediately, and permanently delete the original and any copies of this email and any attachments thereto. |
In reply to this post by Dirk
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Dear Jacques, make sure you try before you buy. We have tested systems from various manufacturers, and I found there are major differences in... # obviously brightness # switching speed: principally, LEDs switch on within microseconds, but it's the electronics that slows it down: some allow two-colour imaging at >25fps, others only make it to ~5-6fps #stability, especially with fast switching: when switching too fast, intensity might still increase during power ramp-up; if pushed too far, you end up with flickering light # number of LEDs per box (in the facility, the more the better, so you don't have to change them twice a day) # there is a huge difference how easy it is to swap LEDs, some really are plug&play, others need to take the whole thing apart Good luck, Martin On Wed, 22 Feb 2012 02:07:47 -0800, jacques <[hidden email]> wrote: >***** >To join, leave or search the confocal microscopy listserv, go to: >http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >***** > >Hi all, > >two years later, and a quite similar question. > >I use a Zeiss epifluorescence microscope, and I'm thinking to replace my HBO >housing by a LED multicolor source. >Of course, Zeiss sells the colibri setup, but the choice on the market is >growing day after day, and for example, Thorlabs also sells something with 4 >colors and a control unit, with the Zeiss mounting system. >I'm sure many other manufacturers do the same. > >Do anybody compared the HBO to a LED system for classical epifluorescence ? >What should be the LED power to have something similar to a 100W at common >wavelenghts (dapi, gfp, rhodamine, etc) ? Even after checking this page : >http://www.olympusfluoview.com/theory/noncoherentsources.html >It is still difficult to know which valeus and units should be considered... > >Thanks for the help > >J. > > >----- > >Jacques FATTACCIOLI > >Département de Chimie >Ecole Normale Supérieure >24 rue Lhomond 75231 Paris Cedex 05 >Email : [hidden email] >Web : http://jacquesfattaccioli.wordpress.com >-- >View this message in context: >Sent from the Confocal Microscopy List mailing list archive at Nabble.com. |
In reply to this post by James Pawley
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi Jim, Thank you for your comments and questions… all very good points. The numbers I gave were rather crude, and were primarily meant to give a rough idea of the typical illumination intensities needed for fluorescence excitation on widefield microscopes, and therefore of the output one would need from a light source, assuming a loss of maybe 50% through the scope. The power levels are measured with a 10x objective, primarily because it's easy for us to get accurate and repeatable measurements with 10x or lower power lenses and our power meter. The objectives are not comparable between different scopes, so the cross-comparisons are approximative. We generally use power values to track how any individual microscope is performing from day to day, not so much to do microscope-to-microscope comparisons, so no special care was taken to keep all things as equal as possible. For any given scope, we use the same settings each time though. Actually I re-checked my numbers, and the fraction of laser light we get at the focal plane on our laser scanning confocal is around 20-25% of the nominal laser power. Our confocal does not have provisions to adjust the diameter of the laser beam to match the back aperture, as far as I know, so there is probably some loss there. Our values are close to the values we had after the instrument was installed years ago, and I think they represent the transmission efficiency of our system. Typically, this is still more power than we need. However, we have other laser-based systems were I was quite dismayed by the amount of light loss from laser source to sample. In fact, I do wonder why many systems these days come with lasers rated 50-100 mW, when we can collect perfectly good images with our 1 mW HeNe, even at 20% transmission efficiency... Incidentally, I originally performed the comparison between microscopes because some of our users think lasers are used on microscopes because they have more power than mercury lamp (or similar) illumination, and also that a laser based microscope will bleach the sample more than one with a lamp. I was trying to show that this is not true. In fact, we could easily get 10-20 mW at the focal plane with lamps and filters for most excitation bands on most of our microscopes, which is more than the nominal power of any laser line on our confocal (some of which are no stronger than a laser pointer), and definitely much more than the actual power we would use for imaging. Obviously, many forget that with widefield illumination, the power is distributed over the entire field of view, while in point scanning, it's all concentrated on one spot, so all this was aimed at illustrating that the illumination mode matters more than the light source, and that if lasers are used, it's typically for other reasons. I guess the reason why many think a laser bleaches samples more is primarily because many microscope users associate lasers with point scanning confocals, and there is a tendency to (mis-)use laser power as the primary means to adjust image brightness, and go way beyond saturation levels, something which can't be achieved as easily in wide field. Yet, it's all in the Handbook! best, Julio. == On Feb 23, 2012, at 10:24 AM, James Pawley wrote: > > Dear Julio, > > Thank you so much for putting some actual measurements into this important discussion. > > Could I ask you for a few more details? > > I assume that you did these tests with the same objective, or at least with an objective with the same NA and mag (and hence the same size and location of the BFP. Light that hits the metal will not pass through the objective). Assuming that you measured light coming out of the objective with some sort of photometer, the other big variable is how you set the size of the field diaphragm (referred to the image plane: For instance, you might set it to be the same as the field of view of a particular objective, or maybe to some known and repeatable fraction thereof.) > > Also, with respect to your comment on confocal performance, I am assuming that the 10% efficiency refers to the fraction of the light leaving the laser that arrives at the focus plane. Ten percent seems low and seems to suggest to me that either the optics used to launch the laser light into the fiber are not well aligned or that the optics are set up to over-fill the BFP of the particular objective used for the test. Otherwise it is hard to see where so much power would be lost: objective transmissions are now usually quoted in the 80-90% range, and a properly chosen dichroic should be about the same while beam-steering mirrors and galvo mirrors should be about 99%. > > In any case, thank you for the numbers. > > Regards, > > Jim Pawley |
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I vividly remember an incident from a lab I worked at years ago: A student managed to completely melt a Kodak wrattan filter with a mercury lamp. The lamp was barely focused, yet managed to reduce the filter to a lump of slag in a matter of seconds! Power is power... doesn't matter what the source is. Craig On Fri, Feb 24, 2012 at 4:11 PM, Julio Vazquez <[hidden email]> wrote: > Incidentally, I originally performed the comparison between microscopes > because some of our users think lasers are used on microscopes because they > have more power than mercury lamp (or similar) illumination, and also that > a laser based microscope will bleach the sample more than one with a lamp. > I was trying to show that this is not true. In fact, we could easily get > 10-20 mW at the focal plane with lamps and filters for most excitation > bands on most of our microscopes, which is more than the nominal power of > any laser line on our confocal (some of which are no stronger than a laser > pointer), and definitely much more than the actual power we would use for > imaging. Obviously, many forget that with widefield illumination, the power > is distributed over the entire field of view, while in point scanning, it's > all concentrated on one spot, so all this was aimed at illustrating that > the illumination mode matters more than the light source, and that if > lasers are used, it's typically for other reasons. I guess the reason why > many think a laser bleaches samples more is primarily because many > microscope users associate lasers with point scanning confocals, and there > is a tendency to (mis-)use laser power as the primary means to adjust image > brightness, and go way beyond saturation levels, something which can't be > achieved as easily in wide field. Yet, it's all in the Handbook! > > > best, > > Julio. > > == > On Feb 23, 2012, at 10:24 AM, James Pawley wrote: > > > > > Dear Julio, > > > > Thank you so much for putting some actual measurements into this > important discussion. > > > > Could I ask you for a few more details? > > > > I assume that you did these tests with the same objective, or at least > with an objective with the same NA and mag (and hence the same size and > location of the BFP. Light that hits the metal will not pass through the > objective). Assuming that you measured light coming out of the objective > with some sort of photometer, the other big variable is how you set the > size of the field diaphragm (referred to the image plane: For instance, you > might set it to be the same as the field of view of a particular objective, > or maybe to some known and repeatable fraction thereof.) > > > > Also, with respect to your comment on confocal performance, I am > assuming that the 10% efficiency refers to the fraction of the light > leaving the laser that arrives at the focus plane. Ten percent seems low > and seems to suggest to me that either the optics used to launch the laser > light into the fiber are not well aligned or that the optics are set up to > over-fill the BFP of the particular objective used for the test. Otherwise > it is hard to see where so much power would be lost: objective > transmissions are now usually quoted in the 80-90% range, and a properly > chosen dichroic should be about the same while beam-steering mirrors and > galvo mirrors should be about 99%. > > > > In any case, thank you for the numbers. > > > > Regards, > > > > Jim Pawley > |
George McNamara |
In reply to this post by Julio Vazquez
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi Julio, If you are able to get your Hg arc lamp to provide any 2-photon excitation fluorescence, you will deserve a Nobel prize. Until then, your users are correct. George On 2/24/2012 6:11 PM, Julio Vazquez wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Incidentally, I originally performed the comparison between microscopes because some of our users think lasers are used on microscopes because they have more power than mercury lamp (or similar) illumination, and also that a laser based microscope will bleach the sample more than one with a lamp. I was trying to show that this is not true. In fact, we could easily get 10-20 mW at the focal plane with lamps and filters for most excitation bands on most of our microscopes, which is more than the nominal power of any laser line on our confocal (some of which are no stronger than a laser pointer), and definitely much more than the actual power we would use for imaging. Obviously, many forget that with widefield illumination, the power is distributed over the entire field of view, while in point scanning, it's all concentrated on one spot, so all this was aimed at illustrating that the illumination mode matters more than the light source, and that if lasers are used, it's typically for other reasons. I guess the reason why many think a laser bleaches samples more is primarily because many microscope users associate lasers with point scanning confocals, and there is a tendency to (mis-)use laser power as the primary means to adjust image brightness, and go way beyond saturation levels, something which can't be achieved as easily in wide field. Yet, it's all in the Handbook! > > > best, > > Julio. |
In reply to this post by Martin Spitaler
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I just spent two days at Applie Precision - I am writing here because I was also imprssed by the (non-LED) Insight SSI. with Leanna's (applkication engineer) was I able to acquire interference reflection contrast microscopy (IRM, aka RICM) with the cyan excitation and DAPI emission filter. Their custom specified (PCO) 15-bit sCMOS was impressive. We mostly used the lasers and 3D-SIM (now have 642 nm light path also working for SIM). On 2/24/2012 1:24 PM, Martin Spitaler wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Dear Jacques, > > make sure you try before you buy. We have tested systems from various > manufacturers, and I found there are major differences in... > > # obviously brightness > > # switching speed: principally, LEDs switch on within microseconds, but it's > the electronics that slows it down: some allow two-colour imaging at>25fps, > others only make it to ~5-6fps > > #stability, especially with fast switching: when switching too fast, > intensity might still increase during power ramp-up; if pushed too far, you > end up with flickering light > > # number of LEDs per box (in the facility, the more the better, so you don't > have to change them twice a day) > > # there is a huge difference how easy it is to swap LEDs, some really are > plug&play, others need to take the whole thing apart > > Good luck, > > Martin > > > On Wed, 22 Feb 2012 02:07:47 -0800, jacques<[hidden email]> wrote: > > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> Hi all, >> >> two years later, and a quite similar question. >> >> I use a Zeiss epifluorescence microscope, and I'm thinking to replace my HBO >> housing by a LED multicolor source. >> Of course, Zeiss sells the colibri setup, but the choice on the market is >> growing day after day, and for example, Thorlabs also sells something with 4 >> colors and a control unit, with the Zeiss mounting system. >> I'm sure many other manufacturers do the same. >> >> Do anybody compared the HBO to a LED system for classical epifluorescence ? >> What should be the LED power to have something similar to a 100W at common >> wavelenghts (dapi, gfp, rhodamine, etc) ? Even after checking this page : >> http://www.olympusfluoview.com/theory/noncoherentsources.html >> It is still difficult to know which valeus and units should be considered... >> >> Thanks for the help >> >> J. >> >> >> ----- >> >> Jacques FATTACCIOLI >> >> Département de Chimie >> Ecole Normale Supérieure >> 24 rue Lhomond 75231 Paris Cedex 05 >> Email : [hidden email] >> Web : http://jacquesfattaccioli.wordpress.com >> -- >> View this message in context: >> > http://confocal-microscopy-list.588098.n2.nabble.com/LEDs-tp4283200p7307846.html > >> Sent from the Confocal Microscopy List mailing list archive at Nabble.com. >> > -- George McNamara, Ph.D. Image Core Manager Analytical Imaging Core Facility (AICF) University of Miami, Miller School of Medicine http://www.sylvester.org/AICF (AICF home page) PubSpectra data (XLSX file inside) http://www.sylvester.org/documents/PubSpectra.zip (download 2000+ spectra) http://works.bepress.com/gmcnamara/ PubSpectra / UA Graphing Site http://www.mcb.arizona.edu/ipc/fret/index.html (Carl Boswell, now retired) New UA Spectra Database Site http://www.spectra.arizona.edu/ (Urs Utzinger) UMiami Scholarly Repository "selected works" http://works.bepress.com/gmcnamara Care to link? http://www.linkedin.com/in/georgemcnamara Ready for imaging in 2012? Check out: Miami 2012 Winter Symposium: Nanotechnology in Biomedicine February 26-29, 2012, Miami, FL Nature Publishing Group / University of Miami / Scripps Florida http://www.nature.com/natureconferences/miami/mws2012/speakers.html Association of Biomolecular Resource Facilities (ABRF) International Symposium March 17-20, 2012, Orlando, FL http://conf.abrf.org/index.cfm Biomedical Optics 2012 (OSA BIOMED) - Optical Society of America April 29-May 2, 2012, Miami, FL http://www.osa.org/meetings/topical_meetings/BIOMED/default.aspx "Old soldiers never die, they just fade away." - Douglas Macarthur. "Old antibodies die, please throw them away." - GM. “Well of course you can’t understand your data, you have too many controls” - Anna M. Wu, quoted in Andreas Markus Loening, Ph.D. dissertation, UCLA, 2006. "If you do all the controls, you'll never publish." - GM. "If you don't do the controls, you shouldn't publish." ... alternative: "If you don't do the controls, don't waste everyone's time in lab meeting." - GM. |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi John, One of the recent posts in this thread indicated that API is using a Lumencor light source. An API brochure http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf lists the 7 color combined set (4 color standard and 4 color fluorescent protein sets, share 461-489nm). The API pdf includes the following power chart: Power Chart nm mW average power 381-399 55 426-450 85 461-489 54 505-515 22 529-556 85 563-588 89 621-643 48 I don't know the Lumencor line well enough to address this, looks similar to http://www.lumencor.com/spectra_light_engine.html SPECTRA 7 light engines (see table on web page) On 2/25/2012 1:03 PM, John Oreopoulos wrote: > Hi George, > > Just curious, if they're not LEDs, what are the light sources in the Insight? > > John Oreopoulos > > On 2012-02-25, at 12:00 PM, George McNamara<[hidden email]> wrote: > > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> I just spent two days at Applie Precision - I am writing here because I was also imprssed by the (non-LED) Insight SSI. with Leanna's (applkication engineer) was I able to acquire interference reflection contrast microscopy (IRM, aka RICM) with the cyan excitation and DAPI emission filter. Their custom specified (PCO) 15-bit sCMOS was impressive. We mostly used the lasers and 3D-SIM (now have 642 nm light path also working for SIM). >> >> On 2/24/2012 1:24 PM, Martin Spitaler wrote: >> >>> ***** >>> To join, leave or search the confocal microscopy listserv, go to: >>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >>> ***** >>> >>> Dear Jacques, >>> >>> make sure you try before you buy. We have tested systems from various >>> manufacturers, and I found there are major differences in... >>> >>> # obviously brightness >>> >>> # switching speed: principally, LEDs switch on within microseconds, but it's >>> the electronics that slows it down: some allow two-colour imaging at>25fps, >>> others only make it to ~5-6fps >>> >>> #stability, especially with fast switching: when switching too fast, >>> intensity might still increase during power ramp-up; if pushed too far, you >>> end up with flickering light >>> >>> # number of LEDs per box (in the facility, the more the better, so you don't >>> have to change them twice a day) >>> >>> # there is a huge difference how easy it is to swap LEDs, some really are >>> plug&play, others need to take the whole thing apart >>> >>> Good luck, >>> >>> Martin >>> >>> >>> On Wed, 22 Feb 2012 02:07:47 -0800, jacques<[hidden email]> wrote: >>> >>> >>> >>>> ***** >>>> To join, leave or search the confocal microscopy listserv, go to: >>>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >>>> ***** >>>> >>>> Hi all, >>>> >>>> two years later, and a quite similar question. >>>> >>>> I use a Zeiss epifluorescence microscope, and I'm thinking to replace my HBO >>>> housing by a LED multicolor source. >>>> Of course, Zeiss sells the colibri setup, but the choice on the market is >>>> growing day after day, and for example, Thorlabs also sells something with 4 >>>> colors and a control unit, with the Zeiss mounting system. >>>> I'm sure many other manufacturers do the same. >>>> >>>> Do anybody compared the HBO to a LED system for classical epifluorescence ? >>>> What should be the LED power to have something similar to a 100W at common >>>> wavelenghts (dapi, gfp, rhodamine, etc) ? Even after checking this page : >>>> http://www.olympusfluoview.com/theory/noncoherentsources.html >>>> It is still difficult to know which valeus and units should be considered... >>>> >>>> Thanks for the help >>>> >>>> J. >>>> >>>> >>>> ----- >>>> >>>> Jacques FATTACCIOLI >>>> >>>> Département de Chimie >>>> Ecole Normale Supérieure >>>> 24 rue Lhomond 75231 Paris Cedex 05 >>>> Email : [hidden email] >>>> Web : http://jacquesfattaccioli.wordpress.com >>>> -- >>>> View this message in context: >>>> >>>> >>> http://confocal-microscopy-list.588098.n2.nabble.com/LEDs-tp4283200p7307846.html >>> >>> >>>> Sent from the Confocal Microscopy List mailing list archive at Nabble.com. >>>> >>>> >>> >>> >> >> -- >> >> >> George McNamara, Ph.D. >> Image Core Manager >> Analytical Imaging Core Facility (AICF) >> University of Miami, Miller School of Medicine >> >> http://www.sylvester.org/AICF (AICF home page) >> PubSpectra data (XLSX file inside) >> http://www.sylvester.org/documents/PubSpectra.zip (download 2000+ spectra) >> http://works.bepress.com/gmcnamara/ >> PubSpectra / UA Graphing Site >> http://www.mcb.arizona.edu/ipc/fret/index.html (Carl Boswell, now retired) >> New UA Spectra Database Site >> http://www.spectra.arizona.edu/ (Urs Utzinger) >> UMiami Scholarly Repository "selected works" >> http://works.bepress.com/gmcnamara >> Care to link? >> http://www.linkedin.com/in/georgemcnamara >> >> >> Ready for imaging in 2012? Check out: >> >> >> Miami 2012 Winter Symposium: Nanotechnology in Biomedicine >> February 26-29, 2012, Miami, FL >> Nature Publishing Group / University of Miami / Scripps Florida >> http://www.nature.com/natureconferences/miami/mws2012/speakers.html >> >> Association of Biomolecular Resource Facilities (ABRF) >> International Symposium >> March 17-20, 2012, Orlando, FL >> http://conf.abrf.org/index.cfm >> >> Biomedical Optics 2012 (OSA BIOMED) - Optical Society of America >> April 29-May 2, 2012, Miami, FL >> http://www.osa.org/meetings/topical_meetings/BIOMED/default.aspx >> >> >> >> "Old soldiers never die, they just fade away." - Douglas Macarthur. >> "Old antibodies die, please throw them away." - GM. >> >> “Well of course you can’t understand your data, you have too many controls” - Anna M. Wu, quoted in Andreas Markus Loening, Ph.D. dissertation, UCLA, 2006. >> "If you do all the controls, you'll never publish." - GM. >> "If you don't do the controls, you shouldn't publish." ... alternative: "If you don't do the controls, don't waste everyone's time in lab meeting." - GM. >> >> >> >> >> >> >> > -- George McNamara, Ph.D. Image Core Manager Analytical Imaging Core Facility (AICF) University of Miami, Miller School of Medicine http://www.sylvester.org/AICF (AICF home page) PubSpectra data (XLSX file inside) http://www.sylvester.org/documents/PubSpectra.zip (download 2000+ spectra) http://works.bepress.com/gmcnamara/ PubSpectra / UA Graphing Site http://www.mcb.arizona.edu/ipc/fret/index.html (Carl Boswell, now retired) New UA Spectra Database Site http://www.spectra.arizona.edu/ (Urs Utzinger) UMiami Scholarly Repository "selected works" http://works.bepress.com/gmcnamara Care to link? http://www.linkedin.com/in/georgemcnamara Ready for imaging in 2012? Check out: Miami 2012 Winter Symposium: Nanotechnology in Biomedicine February 26-29, 2012, Miami, FL Nature Publishing Group / University of Miami / Scripps Florida http://www.nature.com/natureconferences/miami/mws2012/speakers.html Association of Biomolecular Resource Facilities (ABRF) International Symposium March 17-20, 2012, Orlando, FL http://conf.abrf.org/index.cfm Biomedical Optics 2012 (OSA BIOMED) - Optical Society of America April 29-May 2, 2012, Miami, FL http://www.osa.org/meetings/topical_meetings/BIOMED/default.aspx "Old soldiers never die, they just fade away." - Douglas Macarthur. "Old antibodies die, please throw them away." - GM. “Well of course you can’t understand your data, you have too many controls” - Anna M. Wu, quoted in Andreas Markus Loening, Ph.D. dissertation, UCLA, 2006. "If you do all the controls, you'll never publish." - GM. "If you don't do the controls, you shouldn't publish." ... alternative: "If you don't do the controls, don't waste everyone's time in lab meeting." - GM. |
In reply to this post by Dirk
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, we are just about to go thru the same procedure. Some things became obvious while unravelling the marketing pdfs. 1. nobody tells you where and how they measure output or what the illuminated field of view is... 45mW on a 11mm f.o.v is different from 45 mW on a 25mm field of view (I assume the direct coupled will all fill the 25 while a 0.3NA fibre only fills F.oV of 16mm reasonably) 2. solid-state is just another word for "yes, for some we use a phosphor in front the led chip" and this will definatly not increase overall power since phosphorescence will never reach 100% efficiency.... My advice... get a device to your lab and test it. Best wishes from the cold north, Chris |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Some of these 'solid state' light sources claim to be plasma based? What are these things, and does anyone care to share their experiences with them? Here's a link to the kind of source I'm talking about: http://www.thorlabs.com/NewGroupPage9.cfm?ObjectGroup_ID=5551 I don't have any experience with these and was curious. Craig On Sun, Feb 26, 2012 at 10:38 AM, Christian Mohn <[hidden email] > wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > > we are just about to go thru the same procedure. Some things became obvious > while unravelling the marketing pdfs. > 1. nobody tells you where and how they measure output or what the > illuminated field of view is... 45mW on a 11mm f.o.v is different from 45 > mW > on a 25mm field of view (I assume the direct coupled will all fill the 25 > while a 0.3NA fibre only fills F.oV of 16mm reasonably) > 2. solid-state is just another word for "yes, for some we use a phosphor in > front the led chip" and this will definatly not increase overall power > since > phosphorescence will never reach 100% efficiency.... > > My advice... get a device to your lab and test it. > > Best wishes from the cold north, > > Chris > |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** If you follow the links from that Thorlab website you gave, you find: "the heart of LIFI(r) is the bulb sub-assembly where a sealed bulb is embedded in a dielectric material. This design is more reliable than conventional light sources that insert degradable electrodes into the bulb. The dielectric material serves two purposes: first as a waveguide for the RF energy transmitted by the RF Power Amplifier Circuit (PA) and second as an electric field concentrator that focuses energy in the bulb. The energy from the electric field rapidly heats the material in the bulb to a plasma state that emits light of high intensity and full spectrum." In other words, it's not solid-state at all. But it's a very neat idea. Note that they don't tell you what's in the bulb! Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Craig Brideau Sent: Monday, 27 February 2012 12:11 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Some of these 'solid state' light sources claim to be plasma based? What are these things, and does anyone care to share their experiences with them? Here's a link to the kind of source I'm talking about: http://www.thorlabs.com/NewGroupPage9.cfm?ObjectGroup_ID=5551 I don't have any experience with these and was curious. Craig On Sun, Feb 26, 2012 at 10:38 AM, Christian Mohn <[hidden email] > wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > > we are just about to go thru the same procedure. Some things became > obvious while unravelling the marketing pdfs. > 1. nobody tells you where and how they measure output or what the > illuminated field of view is... 45mW on a 11mm f.o.v is different from > 45 mW on a 25mm field of view (I assume the direct coupled will all > fill the 25 while a 0.3NA fibre only fills F.oV of 16mm reasonably) 2. > solid-state is just another word for "yes, for some we use a phosphor > in front the led chip" and this will definatly not increase overall > power since phosphorescence will never reach 100% efficiency.... > > My advice... get a device to your lab and test it. > > Best wishes from the cold north, > > Chris > |
In reply to this post by Boswell, Carl A - (cboswell)
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, My experience has been similar to Carl's, I have almost never gotten an actual emission spectra for a solid state light source. Hence I always measured it myself. The manufacturer just quotes a centre wavelength and sometimes a bandwidth. Maybe I'm overly cautious but I do the same for optical filters too, especially older coloured glass ones. IMO it is very important to know the actual spectrum of the light source. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Saturday, 25 February 2012 3:12 AM To: [hidden email] Subject: Re: LEDs Hi Guy, My reference to lack of spectra came from the online documentation for API's InsightSSI http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no spectra) and X-Cite's XLED1's http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012. pdf separate LED spectra. Lumencor's SOLA light engine brochure http://www.lumencor.com/products/sola_product_sheet.html is the closest I've seen to a useable spectrum. In fact, thanks to Lumencor's openness the SOLA spectrum is on the database, www.spectra.arizona.edu. With the ability to juxtapose (and download) this spectrum with spectra from the listed filters, mirrors and labels, the value to the imaging community of making these light source spectra available becomes more obvious. Cheers, C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: Friday, February 24, 2012 6:36 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques |
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I have been using the Thorlabs LED's for several years with good success. If you look at http://www.thorlabs.com/NewGroupPage9.cfm?ObjectGroup_ID=2615 and click on the Spec tab, they show the spectrum of each LED. I measured one myself because I was concerned about spurious wavelengths causing damage to cells but did not find any and got similar results to the published spectrum. I have found that you need to use excitation filters in the cube. One of the things that has not been mentioned, but I really like about them, is that my system (home built) also uses a white LED to power the transmitted light path. With this addition, I don't need a shutter to control the light source. Unfortunately, I have not seen any of the commercial LED's include this option. Have I missed one? Dave On Feb 27, 2012, at 7:20 AM, Justin Ross wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > My experience has been similar to Carl's, I have almost never gotten an > actual emission spectra for a solid state light source. Hence I always > measured it myself. The manufacturer just quotes a centre wavelength and > sometimes a bandwidth. Maybe I'm overly cautious but I do the same for > optical filters too, especially older coloured glass ones. IMO it is > very important to know the actual spectrum of the light source. > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Boswell, Carl A - (cboswell) > Sent: Saturday, 25 February 2012 3:12 AM > To: [hidden email] > Subject: Re: LEDs > > Hi Guy, > > My reference to lack of spectra came from the online documentation for > API's InsightSSI > http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no > spectra) and X-Cite's XLED1's > http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012. > pdf separate LED spectra. Lumencor's SOLA light engine brochure > http://www.lumencor.com/products/sola_product_sheet.html is the closest > I've seen to a useable spectrum. > > In fact, thanks to Lumencor's openness the SOLA spectrum is on the > database, www.spectra.arizona.edu. With the ability to juxtapose (and > download) this spectrum with spectra from the listed filters, mirrors > and labels, the value to the imaging community of making these light > source spectra available becomes more obvious. > > Cheers, > C > > > Carl A. Boswell > 520-742-6131 > > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Guy Cox > Sent: Friday, February 24, 2012 6:36 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Well, I think they do. The second edition of my book 'Optical Imaging > Techniques in Cell Biology' has spectra from one of the major players in > the field. I do think that this data is freely available. Has anyone > actually had trouble getting these spectra? > > > Guy > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Boswell, Carl A - (cboswell) > Sent: Friday, 24 February 2012 1:49 PM > To: [hidden email] > Subject: Re: LEDs > > "If you have a spectrometer available I suggest scanning the emission to > get the real emission spectrum of the LED." > > It would be of more general use if the manufacturer supplied a > standardized, trustworthy spectral output of these devices. Then > everyone could determine for themselves what would work best for their > systems and samples without the need for additional instrumentation. > C > > > Carl A. Boswell > 520-742-6131 > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Justin Ross > Sent: Thursday, February 23, 2012 6:14 PM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > We previously had the Xe lamps and changed to the SSI illumination on > our Deltavision Core. The exposure times were reduced by a factor of 5 > to 10. The SSI is actually made by Lumencor (no commercial interest). > From my experience it works really well without the downsides of a lamp > of heat generation or having to align and change bulbs. > If you do decide to use LEDs, I strongly suggest still using bandpass > excitation filter (as suggested previously) because with some LEDs there > upto 5% light emission at wavelengths upto 50 nm higher than the peak > which will generally add a lot of background to your images. If you have > a spectrometer available I suggest scanning the emission to get the real > emission spectrum of the LED. > > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List [mailto:[hidden email]] > On Behalf Of Julio Vazquez > Sent: Friday, 24 February 2012 4:14 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hello all, > > Applied Precision just told me that the InsightSSI illuminator is > actually not LED based, so please ignore my previous posting, as far as > LEDs are concerned. > > Julio. > > == > > > On Feb 23, 2012, at 2:06 AM, jacques wrote: > >> ***** >> To join, leave or search the confocal microscopy listserv, go to: >> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy >> ***** >> >> Thanks for your help and comments ! >> >> Jacques Visiting Professor David Knecht Beatson Institute for Cancer Research University of Glasgow Switchback Road, Bearsden Glasgow Scotland G61 1BD UK Visiting Professor David Knecht Beatson Institute for Cancer Research University of Glasgow Switchback Road, Bearsden Glasgow Scotland G61 1BD UK |
Sylvie Le Guyader-2 |
In reply to this post by Justin Ross
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To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi list! One quick question on the side of the LED topic: what is your favourite device to measure the excitation light power at the object plane? I saw that Xcite has one (http://www.ldgi-xcite.com/products-xr2100-xp750.php) as well as Gigahertz (http://www.gigahertz-optik.de/357-0-PT-9610+mit+PD-2T.html). Does anyone have some experience with them or others? Med vänlig hälsning / Best regards Sylvie @@@@@@@@@@@@@@@@@@@@@@@@ Sylvie Le Guyader Live Cell Imaging Unit Dept of Biosciences and Nutrition Karolinska Institutet Novum 14183 Huddinge Sweden office: +46 (0) 8 5248 1107 LCI room: +46 (0) 8 5248 1172 mobile: +46 (0) 73 733 5008 > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] On Behalf Of Justin Ross > Sent: 27 February 2012 08:21 > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > My experience has been similar to Carl's, I have almost never gotten an > actual emission spectra for a solid state light source. Hence I always > measured it myself. The manufacturer just quotes a centre wavelength and > sometimes a bandwidth. Maybe I'm overly cautious but I do the same for > optical filters too, especially older coloured glass ones. IMO it is > very important to know the actual spectrum of the light source. > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] > On Behalf Of Boswell, Carl A - (cboswell) > Sent: Saturday, 25 February 2012 3:12 AM > To: [hidden email] > Subject: Re: LEDs > > Hi Guy, > > My reference to lack of spectra came from the online documentation for > API's InsightSSI > http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no > spectra) and X-Cite's XLED1's > http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012. > pdf separate LED spectra. Lumencor's SOLA light engine brochure > http://www.lumencor.com/products/sola_product_sheet.html is the closest > I've seen to a useable spectrum. > > In fact, thanks to Lumencor's openness the SOLA spectrum is on the > database, www.spectra.arizona.edu. With the ability to juxtapose (and > download) this spectrum with spectra from the listed filters, mirrors > and labels, the value to the imaging community of making these light > source spectra available becomes more obvious. > > Cheers, > C > > > Carl A. Boswell > 520-742-6131 > > > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] > On Behalf Of Guy Cox > Sent: Friday, February 24, 2012 6:36 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Well, I think they do. The second edition of my book 'Optical Imaging > Techniques in Cell Biology' has spectra from one of the major players in > the field. I do think that this data is freely available. Has anyone > actually had trouble getting these spectra? > > > Guy > > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] > On Behalf Of Boswell, Carl A - (cboswell) > Sent: Friday, 24 February 2012 1:49 PM > To: [hidden email] > Subject: Re: LEDs > > "If you have a spectrometer available I suggest scanning the emission to > get the real emission spectrum of the LED." > > It would be of more general use if the manufacturer supplied a > standardized, trustworthy spectral output of these devices. Then > everyone could determine for themselves what would work best for their > systems and samples without the need for additional instrumentation. > C > > > Carl A. Boswell > 520-742-6131 > > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] > On Behalf Of Justin Ross > Sent: Thursday, February 23, 2012 6:14 PM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hi, > We previously had the Xe lamps and changed to the SSI illumination on > our Deltavision Core. The exposure times were reduced by a factor of 5 > to 10. The SSI is actually made by Lumencor (no commercial interest). > From my experience it works really well without the downsides of a lamp > of heat generation or having to align and change bulbs. > If you do decide to use LEDs, I strongly suggest still using bandpass > excitation filter (as suggested previously) because with some LEDs there > upto 5% light emission at wavelengths upto 50 nm higher than the peak > which will generally add a lot of background to your images. If you have > a spectrometer available I suggest scanning the emission to get the real > emission spectrum of the LED. > > > Regards, > Justin. > > -----Original Message----- > From: Confocal Microscopy List > [mailto:[hidden email]] > On Behalf Of Julio Vazquez > Sent: Friday, 24 February 2012 4:14 AM > To: [hidden email] > Subject: Re: LEDs > > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Hello all, > > Applied Precision just told me that the InsightSSI illuminator is > actually not LED based, so please ignore my previous posting, as far as > LEDs are concerned. > > Julio. > > == > > > On Feb 23, 2012, at 2:06 AM, jacques wrote: > > > ***** > > To join, leave or search the confocal microscopy listserv, go to: > > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > > ***** > > > > Thanks for your help and comments ! > > > > Jacques |
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I think you can use a powermeter. As it's possible use a model with lambda mode. Thorlabs send one and i use it on a two-photon microscope. |
In reply to this post by Justin Ross
*****
To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I find this very difficult to stomach. Has anyone actually known X-cite refuse to release actual spectra? What's on the web site is what some publicity person thinks should be there, the question is what the company will give you. If X-cite are happy to release full spectra to me, for publication in a textbook available to everyone, I find it totally beyond belief that they would not give the same information to a customer. The same applies to filters - I've never, ever, had a filter company refuse to provide a spectrum - how could they? And no request from me to publish a spectrum in a paper or book has ever been refused. Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Monday, 27 February 2012 6:21 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, My experience has been similar to Carl's, I have almost never gotten an actual emission spectra for a solid state light source. Hence I always measured it myself. The manufacturer just quotes a centre wavelength and sometimes a bandwidth. Maybe I'm overly cautious but I do the same for optical filters too, especially older coloured glass ones. IMO it is very important to know the actual spectrum of the light source. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Saturday, 25 February 2012 3:12 AM To: [hidden email] Subject: Re: LEDs Hi Guy, My reference to lack of spectra came from the online documentation for API's InsightSSI http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no spectra) and X-Cite's XLED1's http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012. pdf separate LED spectra. Lumencor's SOLA light engine brochure http://www.lumencor.com/products/sola_product_sheet.html is the closest I've seen to a useable spectrum. In fact, thanks to Lumencor's openness the SOLA spectrum is on the database, www.spectra.arizona.edu. With the ability to juxtapose (and download) this spectrum with spectra from the listed filters, mirrors and labels, the value to the imaging community of making these light source spectra available becomes more obvious. Cheers, C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: Friday, February 24, 2012 6:36 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques |
Hi Guy,
I usually don't interpret a lack of response from a commercial data source as unwillingness to release data, but more a matter of they can't be bothered. Your status as a recognized expert and book author puts you in a different category than the majority of us "toiling in the field". Indeed I hold the same view of the response (or lack thereof) to requests to labs for spectral data of their published findings. The data MUST be on hand, otherwise how are the published graphs generated, yet getting it can be problematic. We are dependent on the good graces of others and the implied openness and collegiality of modern science. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: Monday, February 27, 2012 3:26 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** I find this very difficult to stomach. Has anyone actually known X-cite refuse to release actual spectra? What's on the web site is what some publicity person thinks should be there, the question is what the company will give you. If X-cite are happy to release full spectra to me, for publication in a textbook available to everyone, I find it totally beyond belief that they would not give the same information to a customer. The same applies to filters - I've never, ever, had a filter company refuse to provide a spectrum - how could they? And no request from me to publish a spectrum in a paper or book has ever been refused. Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Monday, 27 February 2012 6:21 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, My experience has been similar to Carl's, I have almost never gotten an actual emission spectra for a solid state light source. Hence I always measured it myself. The manufacturer just quotes a centre wavelength and sometimes a bandwidth. Maybe I'm overly cautious but I do the same for optical filters too, especially older coloured glass ones. IMO it is very important to know the actual spectrum of the light source. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Saturday, 25 February 2012 3:12 AM To: [hidden email] Subject: Re: LEDs Hi Guy, My reference to lack of spectra came from the online documentation for API's InsightSSI http://www.api.com/downloads/pdfs/lifescience/InsightSSI.pdf (no spectra) and X-Cite's XLED1's http://www.ldgi-xcite.com/downloads/brochures/LDGI-XCite-XLED1-FAQ-2012. pdf separate LED spectra. Lumencor's SOLA light engine brochure http://www.lumencor.com/products/sola_product_sheet.html is the closest I've seen to a useable spectrum. In fact, thanks to Lumencor's openness the SOLA spectrum is on the database, www.spectra.arizona.edu. With the ability to juxtapose (and download) this spectrum with spectra from the listed filters, mirrors and labels, the value to the imaging community of making these light source spectra available becomes more obvious. Cheers, C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Guy Cox Sent: Friday, February 24, 2012 6:36 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Well, I think they do. The second edition of my book 'Optical Imaging Techniques in Cell Biology' has spectra from one of the major players in the field. I do think that this data is freely available. Has anyone actually had trouble getting these spectra? Guy -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Boswell, Carl A - (cboswell) Sent: Friday, 24 February 2012 1:49 PM To: [hidden email] Subject: Re: LEDs "If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED." It would be of more general use if the manufacturer supplied a standardized, trustworthy spectral output of these devices. Then everyone could determine for themselves what would work best for their systems and samples without the need for additional instrumentation. C Carl A. Boswell 520-742-6131 -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Justin Ross Sent: Thursday, February 23, 2012 6:14 PM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hi, We previously had the Xe lamps and changed to the SSI illumination on our Deltavision Core. The exposure times were reduced by a factor of 5 to 10. The SSI is actually made by Lumencor (no commercial interest). From my experience it works really well without the downsides of a lamp of heat generation or having to align and change bulbs. If you do decide to use LEDs, I strongly suggest still using bandpass excitation filter (as suggested previously) because with some LEDs there upto 5% light emission at wavelengths upto 50 nm higher than the peak which will generally add a lot of background to your images. If you have a spectrometer available I suggest scanning the emission to get the real emission spectrum of the LED. Regards, Justin. -----Original Message----- From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Julio Vazquez Sent: Friday, 24 February 2012 4:14 AM To: [hidden email] Subject: Re: LEDs ***** To join, leave or search the confocal microscopy listserv, go to: http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy ***** Hello all, Applied Precision just told me that the InsightSSI illuminator is actually not LED based, so please ignore my previous posting, as far as LEDs are concerned. Julio. == On Feb 23, 2012, at 2:06 AM, jacques wrote: > ***** > To join, leave or search the confocal microscopy listserv, go to: > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy > ***** > > Thanks for your help and comments ! > > Jacques |
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