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). 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. Tom Leonard
This archive was generated by hypermail 2b29 : Sun Jan 05 2003 - 19:26:16 EST