spotty blots

RYBICKI, ED ED at micro.uct.ac.za
Fri Jan 21 04:05:49 EST 1994


> We are having trouble with small dark spots which appear on all of 
our blots.
> Our lab uses MSI's Magnagraph nylon and the following buffer for 
DNA 

I note you don't say whether you are using 32P or not - are you?  
And if so, small dark spots are nearly always probe sticking 
non-specifically to globs of stuff stuck to the membrane.

Why not try the Boehringer labelling and hyb kit, seeing as you are 
using the spin-column?  They have what is probably the safest 
blocking reagent around (protein derived from milk), and the kit is 
good too.  And if you use the non-radioactive labelling 
9digoxigenin-11-dUTP), you don't HAVE to clean up the probe...!  And 
you get sensitivities - especially if you use the chemiluminescent 
substrates - as good as 32P and quite often quicker.

And I don't work for Boehringer, but heartily recommend their kit as 
we have been using it since it came out without a backward look (and 
all the people who have gone off and had babies from my lab in that 
time are most appreciative).

  ____________________________________________________________________
 | Ed Rybicki, PhD             |                                      |
 | (ed at micro.uct.ac.za)        |      "Lord, won't you buy me         |
 | Dept Microbiology           |          A Mer-ce-des Benz           |
 | University of Cape Town     |     My friends all have Porsches     |
 |                             |          I must make amends..."      |                      |
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 | 7700, South Africa          |           - Janis Joplin             |
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