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Hi Dan,

Check out Ian Parker's gizmo in Fig1D of Demuro and Parker 2005

http://www.jgp.org/cgi/content/full/126/3/179/FIG1

Not that different from yours, but does not use 
the camera. The same authors have a similar 
diagram in their 2006 papers (pubmed: Parker I optical patch clamp).

best wishes,

George


At 09:36 AM 11/17/2006, Dan Focht wrote:
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>List
>
>I would like to hear some criticism on the following concepts
>relative to focus drift.
>This is more of a philosophical approach than what I have seen in the
>past but this is how I see and approach it.
>
>I think it is nearly unreasonable to expect the variant factors in
>the world that affect focus drift to remain constant or to be
>controlled enough to prevent the problem.
>Therefore, I think it is reasonable to approach the problem in
>acceptance and anticipation of the variants.
>
>So far all of the attempts to maintain constant focus that I have
>seen adjust the space between the objective and the specimen to
>compensate for the following:
>thermal expansion
>mechanical settling
>perfusion induced focus shift
>evaporation when the gravitational force on a coverslip is reduced
>when the fluid level changes.
>The problem is that you are always out of focus while you are trying
>to find focus.
>You are not in focus again until you arrive at the right point.
>During that time an automated image capture could occur or you may be
>unnecessarily photo-bleaching your specimen while finding focus.
>If you use the coverslip as a reference point and the specimen moves
>relative to the coverslip you are still at a disadvantage.
>
>So, you might ask. What else can be done?
>Here is where I am requesting criticism of a proposed system.
>I have prepared a model of it located at:
>http://66.39.20.117/autonomous/autonomous.gif
>
>The system achieves focus by splitting the image emerging from the
>objective lens into two pathways, (for you 
>photon hungry types a 100%/ 100% mirror can be 
>used)  a focus profiling pathway, diverted path,
>and an observation pathway, straight path.  Each pathway contains an
>electronically indexed lens module.  The focus profiling pathway lens
>module is continuously varied providing Z slices to the focus
>camera.  The numeric value of the lens position in the focus
>profiling pathway is correlated and evaluated with the  image in the
>focus profiling camera by computer.  The indexed value for the
>optimal image correction, obtained from the test path, is then
>compared to the index value of the observation path lens module where
>immediate correction is applied if and when necessary. Therefore,
>time and three-dimensional information which would otherwise be lost
>with conventional focusing methods is acquired, processed, and
>utilized without negative effects to the observation path.  In this
>system all focusing is done in the back focal plane of the
>objective.  The distance between the specimen and the objective
>remains fixed. Variations in focus can be made over a limited but
>useful range + and - with respect to the objectives principle working
>distance with simultaneous compensation for spherical aberration.
>
>Advantages:
>Ambient temperature regulation for the scope will not be necessary.
>Dynamic compensation is independent of real world variants. (listed
>above)
>Specimens can be tracked in 3D independent of the coverslip surface.
>The quantitative or observation path always knows where focus is and
>can go directly to it without "delay"
>Focus profiling can occur in the IR where there is no photo damage
>and the numerical value of the focus for the observation path can be
>offset for visual wavelengths.
>Focus corrections in the observation path are only applied precisely
>and only when necessary so there are no unnecessary interruptions in
>the observation path.
>A further extension of this technology can be applied to serpentine
>scan applications where optical information profiled in the
>peripheral region of an image can be combined with the intended
>vector of travel so that the system can predetermine focus correction
>before the specimen is brought into position.  This saves time in
>applications where repetitive focusing is required along a
>predetermined path.
>
>There is far more I could write about this system but don't want to
>be more wordy than I already have.
>I would appreciate if you would look at it and send me your comments
>off line.
>I submitted a grant to continue development of this but the reviewers
>commented that there were already enough commercially available focus
>correcting systems and it was not necessary.  What do you think?
>Does this have merit or am I barking up the wrong tree?
>
>
>Dan
>
>
>
>
>On Nov 17, 2006, at 1:15 AM, Stephen Cody wrote:
>
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>G'day Olaf,
>
>I did some rough experiments to get an idea of this using a Nikon 60x
>Oil lens, 1.4na.
>
>These experiments were conducted as a teaching exercise and I show
>the results at Jim Pawley's 3D Microscopy of 
>Living Cells course www. 3dcourse.ubc.ca/index.htm   (Free plug Jim :) ).
>
>In a nutshell I found that with a temperature fluctuation of about
>1.5Deg C the focus drift was approximately 1.5um.
>
>This was a simple "hand wavy" experiment, rather than designed to be
>empirically correct.
>
>This experiment has been discussed on the list before....see subject:
>"time-lapse trouble on SP2"
>
>For those who may be interested I've posted details of this
>experiment on my web site www.ludwig.edu.au/confocal/drift .
>
>I recommend anyone setting up live cell imaging for the first time
>use this technique to monitor temperature fluctuations and z-drift.
>
>Cheers
>
>Stephen H. Cody
>
>Microscopy Manager
>Central Resource for Advanced Microscopy
>Ludwig Institute For Cancer Research
>PO Box 2008 Royal Melbourne Hospital
>Parkville  Victoria    3050
>Australia
>Tel: 61 3 9341 3155    Fax: 61 3 9341 3104
>email: [log in to unmask]
>www.ludwig.edu.au/labs/confocal.html
>www.ludwig.edu.au/confocal
>
>-----Original Message-----
>From: Confocal Microscopy List [mailto:[log in to unmask]]
>On Behalf Of Dr. Olaf Selchow
>Sent: Wednesday, 15 November 2006 7:12 AM
>To: [log in to unmask]
>Subject: Thermal Focus Drift - Olympus BX 51 WI -
>
>Search the CONFOCAL archive at
>http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal
>
>Hello List,
>
>anybody out there who has an idea about how much focus shift
>(approximately) I can expect from a 5 (10) deg celsius temperature
>change in the lab on an Olympus BX 51 WI?
>With aproximately I mean: is it 1, 5 or 10 µm?? Or even more?
>
>I know I should keep the temperature constant (the goal in our case is
>plus minus 1 deg C) but we need to know how much the focus shifts to
>have good arguments to ask for better air conditioning the lab...  A
>measurement would take time - therefore it would help a lot if someone
>has done the test already and can help out with his experience.
>
>Thanks very much!
>
>Olaf
>
>
>This communication is intended only for the named recipient and may
>contain information that is confidential, legally privileged or
>subject to copyright; the Ludwig Institute for Cancer Research does
>not waiver any rights if you have received this communication in error.
>The views expressed in this communication are those of the sender and
>do not necessarily reflect the views of the Ludwig Institute for
>Cancer Research.
>
>
>Dan Focht
>Bioptechs Inc.
>V (724)282-7145





George McNamara, Ph.D.
Glendale, CA
818-547-6909
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