Phosphorimager Questions

David Glover D.J.Glover at bham.ac.uk
Thu Mar 2 15:14:15 EST 1995


                      I am using a Molecular Dynamics Phosphorimager to 
obtain quantitative data from northern blots. I have a couple of questions 
that other users around me have been unable to answer to my satisfaction.

                     One of the advantages of a phosphorimager over 
autoradiography and densitometry is that the response of a phosphor screen is 
linear over a much greater range than an autorad. film. Nevertheless, there 
is a limit to the linearity of a phosphor screen.Therefore, should we always 
calibrate our screen to check our signal is within the linear range of the 
screen. I don't know anyone here who does. My 32P signals are often quite weak 
so I am not worried about saturation but rather the lineararity of the screen 
at low range signals. 

                      My second question regards the production of a 
publication quality hardcopy of the image. I vaguely remember some discussion 
on this newsgroup about this a long time ago.If I remember rightly the best 
solution anybody offered was to expose the blot to an autorad. after exposure 
to the phosphor screen and photograph it. This has also been suggested to me 
by others here. I find this very disappointing for such an expensive piece of 
equipment. I do a lot of these so if I had to expose to an autorad for several 
days after quantitation on the phosphorimager I'd never get to finish my PhD! 
At the moment I expose to the phosphor screen for 3-4 days so that I get a 
good contrast between the signal and background and then print it out on an HP 
laserjet at 600 dpi. I'm not totally satisfied with this but I guess I'll have 
to make do unless anyone else has any suggestions.

                       BTW, thanks to all those who take part in the 
phosphorimager discussions. I find them most informative.

Thanks in advance,

David Glover
School of Biochemistry,
University of Birmingham, UK.



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