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>Dear All,
>
>We have a Zeiss LSM 510 setup, version 2.5. We recently acquired a 100x
>objective (plan-neofluar 100x/1.3 oil). We are unable to do Z sectioning
>using this objective, and always get the same error message
>"Range exceeds limit of 1um for plan-neofluar 100x/1.3 objective. Reduce
>slices and interval"
>If I reduce the stack size to less then 1 um then it will do the z section,
>but obviously this is not much use.
>What is going on?
>
>A second problem we have with both the 40x and 100x oil objectives, is that
>the transmission DIC images are full of the black spots and streaks, which
>are consistent every time we scan. The Zeiss technician has taken apart and
>cleaned everything he can in the beam path, but there is no improvement. Any
>suggestions will be gratefully received.
>
>Thanks,
>
> Rebecca Haffner, Ph.D
> Dept. of Veterinary Resources,
> Weizmann Institute,
> Rehovot, Israel.
> 972 8 9342719 (fax 972 8 9342823)
> [log in to unmask]

Hi,

The 1st problem is related to Zeiss's "smart" LSM system. the LSM software
will not allow you to obtain z-stack thicker than the working distance of the
lens you are using. In this case, the software "thinks" the lens has a working
distance of 1um. I think somewhere the database for objectives is messed up.
go to "maintenance", "objective" and I do not remember if you can edit the
database there. But you can certainly create a new objective with whatever the
working distance it should be there, you need to invent a objective number
though. then you will be fine. If you have trouble to get it done, email me
and I will give you the detailed procedute how to do it (I am not at work
now). Or you can ask your local Zeiss guy to edit the database and put the
correct value for the lens in it (use "dbtool" open your LSM database, under
available or potential objectives).

As to the DIC images, I am sure you thought of these already, set up the
Kolher illumination and set the iris on condensor  wide open. See if this
helps. Another thing to check if your immersion oil has crystals in it.

Good luck!

Xuejun