From our experiences with the 0.5mW green and 5mW red HeNe of our Zeiss
LSM410 I agree with all comments so far that the power of the HeNe
lasers is more than sufficient - at least for monolayer cells, I don't
know about tissue sections.
Particularly I agree with Paul Rigby that correct alignment of the
lasers is critical. However, as far as I understand, one should have the
same problem with a multiline laser, as the different wavelenghts have
to go on separate paths, at least if one wants to attenuate the
wavelengths independently. Could anybody please correct me, if I am
wrong? Is it possible (e.g. with AODs) to attenuate the lines
independently without splitting them?
Bob, what do you mean by saying the argon-krypton laser has more noise?
Is the laser power fluctuating? Is this a common feature of argon /
argon-krypton lasers?

Regards, Joachim


Robert Zucker wrote:
>
> We have compared a sample stained with Lysotracker red ( optimally excited at
> 568)   on a Zeiss 510 with a HENE 543 (.5mw laser) and an  Argon Krypton laser
> in a Leica TSC-SP1. The power measured on a stage with the equivalent 5x lens
> was approximately  .3mw with a Zeiss unit and 1.5mw with a Leica unit. There was
> sufficient excitation with the Zeiss 510 unit to allow our sample to be
> visualized . In fact the power had to be reduced as it saturated the PMT's. The
> HENE has the advantage of having less noise than the argon -kypton laser. This
> data has made me change my opinion on the necessity of a argon-kypton laser and
> it seems that if the system has good optical transmission.  I would definitely
> use a HENE laser in preference to a noisy argon-krypton laser.
> Bob
>
> Robert M. Zucker, PhD
> U.S. Environmental Protection Agency
> MD 72
> National Health and Environmental Effects Research Laboratory
> Research Triangle Park, North Carolina, 27711
> Tel: 919-541-1585; fax 919-541-4017
> e-mail: [log in to unmask]
>
> |--------+----------------------->
> |        |          [log in to unmask]
> |        |          ensu.ca      |
> |        |                       |
> |        |          08/28/00     |
> |        |          02:11 PM     |
> |        |          Please       |
> |        |          respond to   |
> |        |          CONFOCAL     |
> |        |                       |
> |--------+----------------------->
>   >----------------------------------------------------|
>   |                                                    |
>   |       To:     [log in to unmask]   |
>   |       cc:                                          |
>   |       Subject:     Green HeNe laser power          |
>   >----------------------------------------------------|
>
> We are contemplating a Zeiss LSM510 and are trying to think through the
> laser configuration.  They can only demo Option 1.
>
> Option 1: Argon, Green HeNe, Red HeNe (457, 488, 512, 543, 633)
> Option 2: Argon/Krypton, Red HeNe (or Red Diode) (488, 568, 633)
> Option 3: Argon, Nd:YAG, Red HeNe (or Red Diode) (488, 532, 633)
>
> Is a 1mW Green HeNe of any use in "typical" sample scenarios ie; not bright
> actin or tonnes of GFP?  Is the 568 line of an Argon Krypton more useful
> (power vs wavelength)?  What about a diode pumped Nd:YAG 20mW @ 532nm?
>
> Any comments welcome.
>
> - Derek Schulze
>
> Flow Cytometry and Confocal Microscopy Core Facility
> Cancer Research Labs
> 353 Botterell Hall
> Queen's University
> Kingston, Ontario
> K7L 3N6
> Canada
>
> http://meds.queensu.ca/medicine/crl/

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Joachim Walter, Dipl. Phys.
Institut für Anthropologie und Humangenetik der LMU München
AG Cremer
Richard-Wagner-Straße 10/I
D-80333 München               Tel. +49 - 89-2180-6713
Germany                       Fax. +49 - 89-2180-6719
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