http://confocal-microscopy-list.275.s1.nabble.com/Glicerin-labeling-tp7265984p7268559.html
light image while collecting the fluorescence. This will record the
need to have zero intensity set correctly. Then divide the fluorescent
intensity. Not perfect and assumes a linear relationship between power
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>
> Dear list,
>
> Please advice what is the most efficient/cheap way to deal
> with confocal laser instability. Our LSM 510 is 7 years old
> but Ar and Hene 543 lasers have been replaced two years
> ago and have been used no more that 2000-2500 hrs.
> Users started complaining about 488nm and 543nm laser
> line stability, and I measured it using Chroma fluorescent
> plastic slides and 10x objective. The frame- averaged fluorescent
> signal is reduced by 40-50% in 1hr after laser start. The decline
> curve is quite steady after first 3-5 min with some fluctuations.
>
> The most disturbing are intermittent large scale signal
> fluctuations (2-4 fold)
> occurring on about a minute timescale. They kill dynamic (e.g. FRET) or
> long-term imaging. They happen irregularly, on average once in 4-5 days,
> and always after 2-3 hrs after the lasers are started.
>
> In a related post several years ago laser cooling, laser
> polarization, AOTF cooling,
> and AOTF driver stability were listed as the key factors. I am quite
> confident that cooling
> is not an issue as the fans within the equipment boxes are working
> well, and whole room
> has a powerful local conditioning system which maintains ambient
> temperature very stable.
> Zeiss engineer checked the fibers recently, and suggested that maybe
> AOTF drivers are
> not stable. As we are not on service contract and have to pay for
> every replacement I
> would be interested to do more testing myself to identify the
> failing component.
>
> Any suggestions/advices are very welcome. I can provide the
> measurement curves if
> somebody is intersted (contact me off list).
>
> Thanks,
> Arvydas
>
>
>
>
>
>
> Arvydas Matiukas, Ph.D.
> Director of Confocal&Two-Photon Core
> Department of Pharmacology
> SUNY Upstate Medical University
> 766 Irving Ave., WH 3167
> Syracuse, NY 13210
> tel.: 315-464-7997
> fax: 315-464-8014
> email:
[hidden email]
>