Just take a guess and do it. Your controls are always going to be flaming
hot compared to the samples and the dilutions are never going to be right
the first time but it's amazing how off you can be and the gel can still go
into a journal with a little manipulation. The first gel will tell you all
you need to know to calibrate the label from then on. Keep records of
everything including age of label. And even when the slippery gel falls to
the floor and breaks into a million pieces just jigsaw puzzle it back
together on the blot and squeeze. I haven't done a Southern since before
they invented phosphorimagers but I here you can do wonders with enhancing
the data on those things. (Don't cut and paste though, that's called
"faking your data" and is frowned upon).
Not sure what you mean by "internal to the samples" but hope this helps, Paul
Paul Morrison D830
Molecular Biology Core Facilities
Dana-Farber Cancer Institute
44 Binney Street
Boston, MA 02115
p_morrison@dfci.harvard.edu
http://mbcf.dfci.harvard.edu
phone 617-632-3082
fax 617-632-4814