Posted by
Arne Seitz on
URL: http://confocal-microscopy-list.275.s1.nabble.com/magnification-tp7580873p7580874.html
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Hi,
magnification can be very dangerous parameter. You obtain a magnification of 1 per definition only if you look at an object at the conventional viewing distance of 25 cm. Thus either changing the viewing distance or changing the size of an electronic image will change the magnification of the image. Therefore it is much better to include a scale bar in the image. This automatically also changes if you change the viewing distance and or the size of the image.
In order to create a scale for an image of an wide-field microscope you need to know the total magnification of you microscope (=magnification of the objective fold intermediate magnification; latter on can also be 1) and the pixel size of your camera (if you use binning on the camera you also have to account for this). If you are unsure of the total magnification and/or the pixel size you can also use a calibration slide to measure it (see following link:
http://www.swarthmore.edu/NatSci/nkaplin1/scalebar.htm).
For confocal microscopy it is a little more complicated as you have various other parameters which influence the pixel size in an image (e.g. zoom factor, image size). Therefore all of the images of commercially available confocal microscopes save this information with the image (sometimes maybe not for all of the image formats). So you do not have to care about it. If you open/imort the images with ImageJ (with the LOCI Bioformats plugin you can read even the proprietary company formats
http://loci.wisc.edu/software/bio-formats) you can directly add a scale bar via: Analyze-->Tools-->Scale Bar...
Best regards
Arne
> -----Original Message-----
> From: Confocal Microscopy List
> [mailto:
[hidden email]] On Behalf Of Nicola
> Green
> Sent: vendredi 16 août 2013 12:57
> To:
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> Subject: magnification
>
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> Hi
> This is probably a very foolish and basic question, but I am going to ask it any
> way and just apologise for my ignorance as a biological specialist using a
> confocal microscope.
>
> In microscopy I would generally multiply the objective lens magnification by
> the ocular lens (often 10x) to get the total magnification of an image.
> I want to know if this also applies for the confocal microscope or is the light
> path such that the objective magnification is the only one relevant?
> If I do need to include additional magnification what would these
> magnifications be, are they dependent upon the system being used (LSM
> 510
> META) or is there a standard magnification?
>
> Thanks for your help in clarifying this for me.
>
> Regards
> Nicola