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35S levels in gel buffer

John A. Newitt newitt at nih.gov
Thu Nov 30 18:53:53 EST 1995

In article <49c4b2$4dl at mserv1.dl.ac.uk>, Neil Harris
<nh3 at st-andrews.ac.uk> wrote:

> After recently having an isotope safety inspection within our department 
> we have been asked to provide the inspector with details of the amount of 
> 35S dATP that ends up in the gel buffer after a sequencing run. Obviously, 
> not only will this be unincorporated isotope, but will also depend on the 
> length of your sequencing run i.e. how much DNA (containing incoporated 
> radionucleotide) is allowed to run of the bottom of the gel.I wondered if 
> anyone out there has had to do such an evaluation or can point me to a 
> good source of imformation.

Just take an aliquot of the buffer, place in a vial with fluor, and count
in a scintillation spectrometer.  Multiply the cpm times the efficiency
(usually around (1/0.80) for 35S) and times the (total volume/aliquot
volume) to get the total amount of dpm in your buffer.  Multiply by 2.2 x
10^6 uCi/dpm to express in microCuries.  This is more accurate than
"guessing" based on some theoretical calculation and isn't that difficult
to do.


John A. Newitt, Ph.D.           |   <newitt at nih.gov>
National Institutes of Health   |   FAX: 301-402-0387
Bethesda, Maryland  USA         |   

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