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Zhaojie,

I'm not familiar with the LSM 700, but based on what you wrote where the Zeiss representatives said this is normal because their are no mechanical shutters, they could be referring to the fact that the confocal laser engine is AOTF-based, and therefore what you might be seeing is AOTF "leakage" of the laser light onto the specimen plane. I believe this is a common issue with AOTF-based laser engines. That is to say, the AOTF always passes a small (non-zero) amount of light into the diffracted order beam. Perhaps you can see it by eye because it is focused to a small spot at the front of the objective lens and it scatters from that point. Should you be concerned about leakage light reaching your sample? I think this is a valid question, but I'm not quite sure what the best test for that would be. I can think of a few.

Maybe other LSM 700 users can comment and confirm/deny this experience with their systems as well.

John Oreopoulos
Staff Scientist
Spectral Applied Research
Richmond Hill, Ontario
Canada
www.spectral.ca


On 2013-08-12, at 1:11 PM, Z.J. Zhang wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
> 
> Hi everyone:
> 
> I am having a weird problem with my Zeiss LSM 700 confocal microscope and hope you could shed light on it -
> 
> When I tried to scan with the lasers off (turning the lasers off via the ZEN software), I can literally see a relatively bright light coming from the objective! The light looks white, which I assume is the combination of the blue, green and red lasers. Although the detected fluorescence signal is 'minimal', it still worries that 1). It might photobleach my sample; and 2). When I scan with one laser, the other lasers might be on as well, so I wouldn't know what exactly I am scanning with!
> 
> I was told by Zeiss that this is "normal" because there are no mechanical shutters in the scan head. I have a hard time to believe it and here are my questions:
> 
> 1). Does anyone experience the same/similar problem?
> 2). What could be the possible cause?
> 3). How am I supposed to fix it?
> 
> Thank you,
> 
> Zhaojie
> 
> Zhaojie Zhang, Ph. D.
> Director, Jenkins Microscopy Facility
> University of Wyoming
> Laramie, WY 82071
> PHONE: 307-766-3038
> FAX: 307-766-5625