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re bleaching and pulse duration, the rationale for nanosecond pulses
with similar interval between pulses was to avoid second photon hits
during the time the fluorophore was in the excited state - which means
that 10kHz is way off the mark.
Quoting David Knecht <[log in to unmask]>:
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> To join, leave or search the confocal microscopy listserv, go to:
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> *****
>
> It is not only possible, but Thorlabs sells an LED controller that
> modulates LED intensity in this way up to 10kHz which I think should
> be adequate for the effect discussed in the Hell paper. I have not
> had a chance to try it yet and see if it works to reduce bleaching.
> Dave
> http://www.thorlabs.com/NewGroupPage9.cfm?ObjectGroup_ID=4003
>
> On Nov 30, 2010, at 10:26 AM, Jeremy Adler wrote:
>
>> *****
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>>
>> It might also be useful to have pulsatile LEDs operating in the
>> nanosecond range - there was a Stefan Hell paper that showed very
>> significant reductions in photobleaching when the gap between pulses
>> was longer than the fluorescent lifetime. Is this possible with LEDs ?
>>
>>
>> Quoting Gordon Scott <[log in to unmask]>:
>>
>>> *****
>>> To join, leave or search the confocal microscopy listserv, go to:
>>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>>> *****
>>>
>>> Hi Guy,
>>>
>>> No the copy strategy didn't work. Of course one only finds that out
>>> after the event. Ho Hum. :-(
>>>
>>> My original thoughts on video were as yours, however I _think_ most
>>> cameras take a full image and then the lines are downloaded
>>> sequentially, but I'm also very aware that that may not be the whole, or
>>> even the correct story. If the line rate _is_ important, and I guess if
>>> the data is downloaded from the camera `live' line-by-line, rather than
>>> as a complete image, then 100kHz will definitely show artefacts and the
>>> case is closed .. I need to keep the linear control.
>>>
>>> The main aim is actually to avoid unnecessary waste heat, though saving
>>> money is always nice. I'm not sure it would make a particularly
>>> significant effect on price, but it may. Any waste heat I have I then
>>> have to get rid of. LEDs must not get as hot as bulbs and indeed we
>>> actively cool them to get the best out of them, so getting the excess
>>> heat out of the boxes needs heat sinks and fans or similar. At present
>>> that waste heat puts a frustrating limit on what's feasible with the
>>> units, and I'd like to remove that frustration.
>>>
>>> Thanks for your comments.
>>>
>>> Kind regards,
>>> Gordon.
>>> --
>>> Gordon Scott Design Engineering
>>> Custom Interconnect Ltd. http://www.cil-uk.co.uk
>>> CoolLED http://www.coolled.com
>>> CoolLED is a division of Custom Interconnect Ltd.
>>> Phone +44-1264-321321
>>> CIL House, Charlton Road, Andover SP10 3JL, UK
>>>
>>>
>>>> -----Original Message-----
>>>> From: Guy Cox [mailto:[log in to unmask]]
>>>> Sent: 29 November 2010 23:15
>>>> To: Confocal Microscopy List
>>>> Cc: Gordon Scott
>>>> Subject: RE: Guidance wanted on illumination stability
>>>>
>>>> Gordon,
>>>>
>>>> Your strategy of copying to the list didn't seem to work.
>>>> Anyway, I think that lots of list members, having seen the
>>>> question, like to see the answers.
>>>>
>>>> If someone is taking conventional images with a 1
>>>> second exposure 100kHz ripple will not be noticeable. But if
>>>> you are taking video at 25 fps 525 line (international video)
>>>> your line rate is about
>>>> 13 kHz (US video about 14kHz) so I'd imagine there would be
>>>> rather unwelcome diagonal stripes on the image and you'd be
>>>> getting angry phone calls from your customers.
>>>>
>>>> You haven't told us the other side of the
>>>> trade-off. Do you want to eliminate the linear stage to save
>>>> money - if so, how much cheaper would it make a CooLED
>>>> illuminator? Or is it to save power?
>>>> How much would that save? Given that an LED source already
>>>> uses hugely less power that an HBO 100 mercury lamp, would
>>>> anyone care?
>>>>
>>>> Guy
>>>>
>>>> Optical Imaging Techniques in Cell Biology
>>>> by Guy Cox CRC Press / Taylor & Francis
>>>> http://www.guycox.com/optical.htm
>>>> ______________________________________________
>>>> Associate Professor Guy Cox, MA, DPhil(Oxon) Australian
>>>> Centre for Microscopy & Microanalysis, Madsen Building F09,
>>>> University of Sydney, NSW 2006
>>>>
>>>> Phone +61 2 9351 3176 Fax +61 2 9351 7682
>>>> Mobile 0413 281 861
>>>> ______________________________________________
>>>> http://www.guycox.net
>>>>
>>>>
>>>> -----Original Message-----
>>>> From: Confocal Microscopy List
>>>> [mailto:[log in to unmask]]
>>>> On Behalf Of Gordon Scott
>>>> Sent: Tuesday, 30 November 2010 2:34 AM
>>>> To: [log in to unmask]
>>>> Subject: Guidance wanted on illumination stability
>>>>
>>>> *****
>>>> To join, leave or search the confocal microscopy listserv, go to:
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>>>>
>>>> Hi Guys,
>>>>
>>>> I'm looking at ways to further improve the performance and
>>>> efficiency of our light sources.
>>>>
>>>> There are always tradeoffs when doing this and I'd like to
>>>> better understand what tradeoffs are acceptable for real
>>>> microscopy users.
>>>>
>>>> Our present illumination sources all use a switched-mode
>>>> pre-regulation and a linear final regulation for the LED
>>>> power, so ripple is very low, but at a cost for us of some
>>>> power wasted in the linear stages.
>>>>
>>>> I can improve that efficiency and reduce the waste by
>>>> foregoing the linear stage and regulating directly with the
>>>> switching mode, but the tradeoffs are a longer On/Off
>>>> switching time than is feasible with linear, and a
>>>> high-frequency ripple.
>>>>
>>>> My simulations suggest switch-on and switch-off times of
>>>> around 50us and a ripple of around 25% at 100kHz, which would
>>>> be reasonable from an electrical/energy point of view.
>>>>
>>>>
>>>>
>>>> The question is, of course, would any of the people likely to
>>>> use it find that performance difficult or unacceptable?
>>>>
>>>>
>>>>
>>>> I've copied to the list rather than posting direct, so
>>>> hopefully the replies will come to me rather than cluttering the list.
>>>>
>>>> Thanks for considering the question, even if you need not, or
>>>> choose not, to answer.
>>>>
>>>> Kind regards,
>>>> Gordon.
>>>> --
>>>> Gordon Scott Design Engineering
>>>> Custom Interconnect Ltd. http://www.cil-uk.co.uk
>>>> CoolLED http://www.coolled.com
>>>> CoolLED is a division of Custom Interconnect Ltd.
>>>> Phone +44-1264-321321
>>>> CIL House, Charlton Road, Andover SP10 3JL, UK
>>>>
>>>>
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>>>
>>
>>
>>
>> Jeremy Adler
>> Genetics & Pathology
>> Rudbeckslaboratoriet
>> Daghammersköljdsväg 20
>> 751 85 Uppsala
>> Sweden
>>
>> 0046 (0)18 471 4607
>
> Dr. David Knecht
> Department of Molecular and Cell Biology
> Co-head Flow Cytometry and Confocal Microscopy Facility
> U-3125
> 91 N. Eagleville Rd.
> University of Connecticut
> Storrs, CT 06269
> 860-486-2200
> 860-486-4331 (fax)
>
Jeremy Adler
Genetics & Pathology
Rudbeckslaboratoriet
Daghammersköljdsväg 20
751 85 Uppsala
Sweden
0046 (0)18 471 4607
|
