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We recently moved solid tumor DNA cell-cycle analysis from an EPICS-
753 to our new ELITE. However, using TruPloid drift analysis
software, we detected a large upward shift in fluorescence intensity
during the first few minutes (2-4) of each new sample acquisition on
the ELITE. The fluorescence eventually would stabilize.
If others are noticing similar instabilities, we found a solution.
We were able to correct the problem by running our PI staining buffer
through the sample line for a few seconds after each sample flush but
before beginning aspiration of the next sample. We do NOT flush the
buffer from the line before beginning aspiration of the next sample.
Evidently, the sample line material has an affinity for PI and
extracts it from the sample until the sample-line material becomes
saturated. As the small volume of buffer around the cells in the
sample-line becomes depleted of PI, the concentration is lowered
sufficiently so that PI starts to leave the DNA and the cells become
dimmer. As the line becomes saturated, less PI leaves the buffer and
the fluorescence stabilizes. Unfortunately, the sheath flush between
samples seems to wash the PI from the line material and the next
sample run must go through the same cycle.
This kind of fluorescence drift was never noted on the old 753. I am
not sure if Coulter has gone to a new sample-line material or the
increase in the length of the sample-line on the ELITE is enough to
explain the differences. I know some institutions like to run PI in
their sheath to help with instabilities. Of course that would be
another approach to the same problem. In that case, the sample line
would remain saturated with PI between samples and would not have the
same oppotunity to scavange PI from each new sample.
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