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Dear Hanna,

There may be several reasons to the problem depending on the pattern. It
may be "ringing" or reflections. While the first one can be fixed by
adjusting the threshold of the signal (increasing the baseline and getting
rid of some noise in the microscope environment), the second problem will
be more difficult to address in a turn-key system. This may be coming from
two reflecting surfaces (mirrors or filters). It is most probably in the
laser coupling part of the setup. This pattern will have to be addressed
anyway before you can do any measurements.

Best wishes,
Vladimir


On Fri, Nov 20, 2015 at 3:14 AM, Hanna Sas Nowosielska <
[log in to unmask]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
> Thank you for all suggestions, unfortunately we cannot use scatter light to
> get IRF. There is some problem either with our WLL laser or with our
> microscope system (Leica SP8) and when using scatter light we don’t obtain
> specific IRF. Instead we get multiple picks of different height and size.
> We talked to service but they’re still trying to find the source of the
> problem.
>
>
> Hanna
>
> 2015-11-20 3:25 GMT+01:00 yuansheng.sun <[log in to unmask]>:
>
> > you can try to use the cream coffee mate solution to record the laser
> > scatter directly.
> >
> > Sheng
> >
> >
> > Sent from my T-Mobile 4G LTE Device
> >
> >
> > -------- Original message --------
> > From: Hanna Sas Nowosielska <[log in to unmask]>
> > Date:11/19/2015  6:49 AM  (GMT-06:00)
> > To: [log in to unmask]
> > Cc:
> > Subject: Measuring IRF in red channel
> >
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Dear Colleagues,
> >
> >
> >
> > Could someone suggest me a fluorophore with lifetime short enough to
> > measure IRF in red channel (excitation: 520nm/ Detection : 565 – 605)? I
> > would be very grateful for an advice.
> >
> >
> >
> > Best regards,
> >
> > Hanna
> >
> >
> >
> > --
> > Hanna Sas-Nowosielska
> >
> > Laboratory of Imaging Tissue Structure and Function
> > Nencki Institute of Experimental Biology
> > Warsaw, Poland
> >
>
>
>
> --
> Hanna Sas-Nowosielska
>
> Laboratory of Imaging Tissue Structure and Function
> Nencki Institute of Experimental Biology
> Warsaw, Poland
>