pH varience in RPMI media

Bernard P. Murray, PhD bpmurray*STUFFER* at
Mon Nov 23 20:57:14 EST 1998

In article <73d05t$kji$1 at>, darren at wrote:

> We are growing NS-1 cells (for making monoclonal antibodies) in complete
> media (10% FCS + commercial liquid RPMI 1640). All seems well, but i've
> noticed that the pH of the RPMI seems to shift around even before any cells
> are grown in it(usually gets pinker, ie pH increases). Can anyone give me
> some guidelines on the "acceptable" range of pH change? What pH parameters do
> NS-1 cells take? Should I take this up with our supplier?Thanks
> Darren

Is the RPMI "complete" as it comes or are you adding bicarbonate
and/or Hepes?
     My preference when developing monoclonals is to use Hepes
buffering as you end up using a lot of dishes that are frequently
taken out of the incubator for inspection which messes up the
CO2 buffering.  The colour should then remain stable.  If you
only use bicarbonate buffering you'll see colour changes in your
stock bottles of medium that are related to the dissolved CO2 content
so the colour will change depending on how often the bottle is
opened and you'll see some fluctuation with temperature (medium
stored at 4degC will get "pinker").  What matters is how stable
the pH is when the medium is in the incubator.  Put a flask
or dish containing medium alone in the incubator and make sure
that this remains at the appropriate pH.
     Also, I would recommend you use another myeloma line
(eg. NS0) for hybridomas as NS1 can end up using their
own light chain for your antibodies so that you end up with
two populations.  It doesn't always happen but when it does
it is a *real* pain.
     Good luck,
Bernard P. Murray, PhD
Dept. Cell. Mol. Pharmacol., UCSF, San Francisco, USA

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