densitometry on western blots

Jose de las Heras via methods%40net.bio.net (by josenet At tiscali.co.uk)
Thu Nov 23 15:56:54 EST 2006


<martinhoehne At gmx.net> wrote in message 
news:1164299514.092168.323980 At f16g2000cwb.googlegroups.com...
> Hello,
>
> I have some basic questions on densitometry and would be glad if anyone
> can point me to a good source to learn more or even answers some of the
> questions.
>
> 1) As I understand it the bands have to be non-saturated. Does this
> really mean that as soon as I have pixel values of 255 (on a 8-bit
> image) I can forget about doing a densitometric analysis?

on any digital image system, once you reach the limit of the dynamic range 
(256 for 8-bit, 65536 for 16-bit, etc) it doesn't make sense to use it.
You can get away with *a few* saturated pixels within the area you are 
measuring. Usually you would use the median (or mean) of the pixels in the 
area, or some other measurement, and having a few saturated pixels won't be 
too bad. But you should aim to avoid any saturation if you want accurate 
results.

It's one little bug of mine, when you see figures with a loading control 
that's clearly saturated... and sure, they all look the same then! :-)

 Jose 




More information about the Methods mailing list