Acid Precipitation

Bill Melchior, NCTR/FDA wmelchior at NTBTOX.NCTR.FDA.GOV
Thu Jul 23 08:12:41 EST 1992


>>When one filters the acid-precipitated material through a glass filter, does
>>the material actually _stick_ to the glass, or is this a simple particulate
>>filtration?  If the latter, Why do we use glass?  Is it just for acid
>>resistance?

It's probably just physically trapped.  I've not done it for many years, but
my recollection is that you want to use a weak vacuum, because a strong one
pulls the DNA through.

>>Can acid precipitated nucleic acids be pelleted and washed as a pellet?  How
>>hard do they have to be centrifuged for this?  Has anyone tried this before?  
>>I would like to try precipitating in 96-well dishes and spinning using a plate
>>carrier in my refrigerated centrifuge, but the maximum RCF is about 2500 x g. 
>>Does this sound sufficient?

>Yes, you certainly can precipitate nucleic acids with TCA in a centrifuge.
>Standard DNAse assays are based on the efficient removal of large DNA
>and the measuring of the acid soluble (small fragments) left in the sup.
>I am not sure if you have enough G force for the microtiter dishes or not,
>I guess it is worth a try. It may be quite dependent on the DNA concentration,
>in fact I would strongly guess that it is.

As MB says, centrifugation certainly works, though 2500 g sounds marginal.
Something else to check for is that TCA, an organic compound, doesn't
dissolve the plastic in your plates; it may be that your solution will be
dilute enough that it won't matter.
________________________________________________________________________________
The opinions stated are mine, not those of NCTR or its sponsoring organizations.

Bill Melchior                                ||   "You have lawyers the way
National Center for Toxicological Research   ||    other people have mice."
Jefferson, AR  72079                         ||
(501) 543-7206                               ||   -Kenneth Duncan, English
                                             ||    Health & Safety Executive,
WMELCHIOR at NTDOC.NCTR.FDA.GOV                 ||    to US regulators
________________________________________________________________________________
The opinions stated are mine, not those of NCTR or its sponsoring organizations.

Bill Melchior                                ||   "You have lawyers the way
National Center for Toxicological Research   ||    other people have mice."
Jefferson, AR  72079                         ||
(501) 543-7206                               ||   -Kenneth Duncan, English
                                             ||    Health & Safety Executive,
WMELCHIOR at NTDOC.NCTR.FDA.GOV                 ||    to US regulators



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