6 replies to this topic

[rocio]

Hi
I’m starting to work whit Azacytidina. To prepare a mother solution I dissolve 24mg on 20uL of wather: acetic acid (1:1) as SIGMA says I but the aza not dissolve. Then I’m going to add PBS to obtain a 10mM mother solution.
I would appreciate if somebody could help me to dissolve the Aza!
Thanks in advance
rocio

[kawaka]

Do you mean azacytindine stock solution? check this discussion
http://www.protocol-...?showtopic=4772

[methylnick]

Hi again Kawaka,

I think your mother solution is too concentrated, 25mg in 20uL? You can dissolve in 1:1 H2O/acetic acid to a final conc of 10mM, but once you add it to your culture media, the pH significantly decreases thus affecting your cells, I would take the suggestion from the link mentioned above and dissolve in PBS. it works!

Nick

[eagertolearn]

i am pretty sure azaC is unstable in solution. Read the Sigma product sheet. It is available online. It would be wise, even if painful to weigh out a little quantity and make a fresh stock everytime.

Any comments.

[methylnick]

i am pretty sure azaC is unstable in solution. Read the Sigma product sheet. It is available online. It would be wise, even if painful to weigh out a little quantity and make a fresh stock everytime.

Any comments.

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you are absolutely correct eagertolearn. azaC has a very short half life and if possible you should use freshly made up stocks each time.

Nick

[epibet]

Well i dissolved Azacytidine (Sigma) 5mg/ml in DMSO it was stable up to almost one year at -80.
Any comments?

[pcrman]

Read the instruction from Sigma, it is very useful.

Azacitidine is very unstable in aqueous solutions, with
a 10% loss of the product in 2 to 3 hours at room
temperature in lactated Ringer's solution.13
Degradation is complex, with an initial reversible
formation of an intermediate formyl product, which
then undergoes slower irreversible change to produce
further degradation products. Loss of azacytidine is
rapid initially, but as the intermediate accumulates,
reversal of the first reaction slows down the apparent
loss of azacytidine. Misunderstanding of the kinetics
has led to erroneously long (90%) stability times being
quoted in the literature.14,15 The half-life of the
compound in phosphate buffered saline, pH 7.4, when
heated to 50 °C is 90 minutes. The stock solution,
when kept at this temperature for 20 hours, lost the
ability to cause cytotoxicity and multinucleation. Stock
solutions should be prepared fresh for each
experiment, sterilized by filtration, and kept at 0 °C
until needed