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Well, this is a excellent opportunity to use once more the cover-all
answer: It depends...
I did a pretty extensive comparison of the the objectives Apo 63x/1.4 Oil
and Apo 100x/1.46. All tests were done with Leica objectives, results may
differ for other brands. Test sample were beads 170nm in Glycergel close to
the coverslip or homogeneously stained cells for the "Sensitivity"
comparison.

Results: Resolution was visibly and measurably better with the 100x. But:
it is absolutely mandatory to adjust the correction collar with great care,
otherwise you will do more harm than good. Not difficult in itself,  but
you simply have to do it. You will also gain in "Sensitivity": you will
need less laser energy (measured in focal plane, not at AOTF) to achieve
the same apparent image brightness. Under these conditions also a lower
bleach rate was observed. So in principle I do think this is an excellent
objective improving the performance of the system. Before this objective I
had the opinion that a 100x would be allways less "bright" than a 63x. For
a 63x/1.2 Water all this would be valid to an even larger degree, just
consider the BUT...

BUT: keep in mind that if you are working with thicker samples (meaning for
me more than approx. 10-15um) the match of the RI (refractive index)
becomes much more important for your imaging than pure NA. So if you work
with living samples usually water immersion is the best, with fixed and
embedded samples  I would usually recommend  Glycerol objectives since most
commercial embedding media are very close in RI to this immersion medium.
So it is very important to decide what is most important for your
application: max. brightness/resolution in flat samples or maximum
penetration/resolution/brightness in thick samples.

Hope this comment is a bit helpfull. For background information go to
Pawley's Handbook of confocal microscopy, read about RI match and spherical
aberration. These factors have a huge impact on your image quality and can
hardly be overestimated.

Good luck

Boris

P.S. I am working for Leica Microsystems, but I think this makes no
difference for this case...
__________________________________________________

Boris Zarda
Dr. rer. nat.
Sales Manager Research Switzerland

Leica Microsystems (Schweiz) AG
Verkaufsgesellschaft
Max Schmidheiny-Strasse 201
CH-9435 Heerbrugg
Tel  +41 44 768 36 30
Fax +41 71 726 34 44




                                                                           
             "Yang, Ann-Fook"                                              
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Hi All,

I am new in confocal microscopy.
We have a 40x/1.3 oil and a 63X/1.2 water objectives. Is there any
advantage to add a 100X/1.46 Oil objective?

Ann Fook Yang,
Agriculture and Agri-Food Canada/Agriculture et Agroalimentaire Canada
EM Unit/ Unite EM
Edifice K.W. Neatby Building,
960 Carling Av /960 Boulevard Carling,
Ottawa,Ontario
K1A 0C6

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Telephone/Téléphone: 613-759-1638
 Facsimile/Télécopieur: 613-759-1701


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