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Hello Ann,
there may be different opinions on this, but here is my take:
With a confocal microscope, you can add magnification by zooming in and matching the image size (pixel dimensions) for optimal resolution. This means you can get 100 x magnification with a 40x at zoom 2.5, or a 63x at zoom 1.6.
What the 100x/1.46 gives you in theory is greater resolution (20-30% more), since resolution increases with the square of the numerical aperture. In practice, you may not always achieve this extra resolution, unless you have quite bright samples that are relatively thin and properly mounted, in order to minimize spherical aberrations.
You also need to consider the type of objective and the application. PlanApos are corrected for more colors, and therefore are best for precise colocalization of up to four dyes. Fluars or similar objectives are corrected for fewer colors, but tend to be brighter than PlanApos, because they are simpler lenses with less glass... these are often better for live-cell imaging, or for samples with fewer colors... for a more precise description, you may want to check the tutorial on lenses and chromatic aberrations here:
We typically use objectives in the 40x-60x range for most of our confocal applications (in addition to a 10x or 20x for low-mag work, and possibly a long-working distance 40x for thick/unusual samples). We rarely use a 100x, except maybe to look at yeast cells, or some very fine cellular detail. A 100x/1.46 lens will often be less bright than a 40x/1.3, which means that your gain in resolution may be offset by increased bleaching. In addition, a 40x is more versatile since you can image a wider field of view at zoom 1, and then get a close-up at zoom 2-3 if you need more mag, without having to switch lenses.
A 100x/1.46 lens, on the other hand, would be quite nice on a widefield microscope... also, this type of lens with high NA is what you need for applications such as TIRF (total internal Reflection microscopy)...
--
Julio Vazquez
Fred Hutchinson Cancer Research Center
Seattle, WA 98109-1024
Tel: Office: 206-667-1215/ Lab: 206-667-4205
FAX: 206-667-6845
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On Oct 26, 2007, at 1:09 PM, Yang, Ann-Fook wrote:
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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
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