S35 safety (re: high activity factor)

HARDIES at THORIN.UTHSCSA.EDU HARDIES at THORIN.UTHSCSA.EDU
Mon Jan 23 14:02:09 EST 1995


Peter Riss, Technical Services at Amersham wrote in response to my
post about volatile S35 in S35 labeled nucleotide preparations:

> One point requires clarification.  S-35 compounds are not prepared by
direct irradiation: only S-35 sulphate is prepared in this way.  The
sulphate is then used in the synthesis of the labelled compound.  This
is important because potentially volatile impurities are not present
when the compounds are prepared but grow in with time through the
processes of chemical and radiolytic decomposition.

> The fact that impurities increase with time suggests a few ways to
prevent contamination:  First, use fresh S-35  labelled compounds, so
that the time for the growth of impurities is minimized.  Second, use
stabilized formulations of labelled compounds.  Various stabilizers
(such as pyridine 3,4-dicarboxylic acid, which is used with methionine
and cysteine) are effective at minimizing radiolytic decompostion of
stock solutions. Third, monitor work surfaces, incubators and
temperature blocks scrupulously.  Finally, consider the  use of other
isotopes such as P-33 where experimental requirements permit.

First, thanks to Peter for this clarification and helpful advice.

Second, the numbers I posted were probably from a stock of S35
nucleotides that had aged past the point were my people were willing
to use it, so they would weigh in on the high side of the problem, 
given that radiolysis is the culpret.

Finally, note that the activity of the nucleotide will
play into the radiolysis problem.  When we stepped up from 500 Ci/mmol
to 1500 Ci/mmol, the radiolysis problem became much more severe.
For example, to maintain the same quality of sequence ladder, we
had to increase the amount of label in the reaction as if it
decays with a half life of 2-3 weeks instead of the physical
half life of 87 days.  Now I see why the companies want to sell
the nucleotide prediluted into preextension mix.  Both the dilution
and the cold nucleotides will stabilize against radiolysis.

Cheers,
Steve Hardies, Assoc. Prof. of Biochem., Univ. of Texas HSC at San Antonio
Hardies at uthscsa.edu




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