Nigericin contamination? -Reply

David Hedley (
Fri, 11 Jul 1997 07:22:27 -0500

Wayne Green asks if there is any evidence that nigericin, used for
intracellular pH calibration, could contaminate sample lines.
We have a great deal of experience with pHi measurement by flow
cytometry using nigericin for the calibration procedure. I have not
encountered sample line contamination as a problem, or seen any
reference to this. Presumably if it was a major problem you should see a
difference in the pHi value for the test sample if this was run before and
then after the calibration run using nigericin. This could easilly be tested if
you were concerned.
As an alternative to nigericin, we recently developed a calibration
procedure based on the partitioning of weak acids and bases across the
cell membrane. These shift pHi to an extent predicted by the
Henderson-Hasselbalch equation, and do not require the use of
ionophores such as nigericin. Sue Chow published this in Cytometry last
year (1996;24:360-367). Intellectually it is the most satisfying flow method
ever to come out of my lab.

David Hedley
Ontario Cancer Institute/Princess Margaret Hospital

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