Tom Leonard wrote- >I'm on a mission to determine bacterial viability as well as quantify >bacteria using Molecular Probes' BacLight viability kit. Using an EPICS >Elite, I have routinely found bacterial counts to be 10% of those >determined with fluorescence microscopy. The accuracy of the microscopy >counts have been verified by other labs. I recognize that the organism has >mechanisms to pump out fluorochrome, but to the extent that 90% of bacteria >go undetected? We're analyzing on the log scale. Our operators are >accustomed to analyzing mammalian cells, but claim they have done all the >"tweaking" possible to optimize bacterial analysis. > >Why such a disparity in counts? My conclusion is that the instrument simply >can't detect the smaller organisms in the population (of E. coli). Possibly; what signals do you trigger on? >Also, we can see distinct populations when analyzing live controls and dead >controls independently, but when we mix the live and dead populations we >are often unable to distinguish between live and dead populations. >Sometimes we can, other times we can't. It is not only strange but >frustrating. > >Has anyone had similar experiences? Any thoughts on what might be >contributing to these unexpected results? Thanks in advance. If you can't get the bugs out of the noise, all bets are off, but I would expect you'd be able to do that in an Elite. -Howard
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