Hello Arnold,

I can just share a similar obervation on our system. We were using
multiphoton excitation in the range of 800 and 960 nm when we were
trying to calibrate the system for the FCS experiments. What we
observed was that there was a signal in case of no sample above the
objective, (inverted) there was a well - expressed fluorescence there.
However, whenever just a glass slide was put, the signal was
diminished almost to the background level. We still don't know what
was the exact source of this phenomena.

With regards,
Vladimir

On Fri, Mar 19, 2010 at 1:36 AM, Arnold Estrada <[log in to unmask]> wrote:
> It scales approximately linearly.
>
> Thanks for a response
> -Arnold E.
> On Mar 18, 2010, at 11:32 PM, Guy Cox wrote:
>
> Interesting.  Does the luminescence intensity scale linearly or
> quadratically with excitation?
>
>                                                 Guy
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[log in to unmask]]
> On Behalf Of Arnold Estrada
> Sent: Friday, 19 March 2010 9:01 AM
> To: [log in to unmask]
> Subject: Near IR luminescence observed from Microscope Objectives.
>
> In the process of trying to make phosphorescent lifetime measurements
> from a very week signal, I have observed that all of my microscope
> objectives seem to luminesce in the near-infrared. My research involves
> using two-photon excitation of phosphorescent probes with very small
> two-photon action cross-sections.  Consequently my excitation source is
> a gated Ti:Sapphire (@ 800 nm) such that the excitation reaches the
> sample (or objective in this case) for 20 microseconds out of every two
> milliseconds.  The laser power reaching the objective is about 400 mW
> while the gating is on.  Using a spectrometer, I have confirmed that
> this luminescence extends from ~ 700nm to greater than 850 nm.  The
> presence of the anti-stokes shifted light implies some sort of
> multi-photon absorption mechanism.  However, I observe this luminescence
> whether the laser is mode locked or not.  I have also observed that the
> luminescence process appears to have a lifetime of ~ 60-80 uS.  Has
> anyone else ever observed this?  I have observed this with other lenses
> (not microscope objectives) as well.  Is it possible that the high laser
> power used is causing a multi-step absorption process followed by
> luminescence from the metal dopants in the high index glass in the
> objective?
>
> Any help would be greatly appreciated.
>
> ---
> Arnold D. Estrada.
>
> Doctoral Candidate
> Department of Biomedical Engineering
> The University of Texas at Austin
> Biomedical Engineering Bldg, 1.324
> 107 W. Dean Keaton Street
> Austin, TX 78712
>
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