From: Hazel Davey (hlr@aber.ac.uk)
Date: Tue Aug 04 1998 - 04:28:54 EST
Rochelle A. Diamond wrote: > We now have two investigators that need to > analyze and sort E. Coli and other bacteria (waste water type). Here are several thoughts based on our analysis of bacteria including E. coli : 1) Make sure that all buffers etc are filtered. With our bacterial samples I use a 0.1um filter rather than a 0.22 to prepare sheath fluid / dilution buffers etc. 2) Is it possible to use a fluorescent stain for your E. coli that would allow you to discriminate them from the other particles? 3) If you have the gated amp electronics on your Elite use them even if your only using one laser - we find it really does improve detection. 4) I doubt that settings are directly transferrable between machines, but just to show that you don't need to push the voltage etc on the forward scatter detector to extreme levels, here are our starting "cytosettings" for bacteria using the Argon ion 488 nm laser (All detectors are set to logarithmic): FS 400 Volts, IGAIN=5, PGAIN=1, DELAY=UP PMT1 (<440nm) 630 Volts, IGAIN=3, DELAY=LOW PMT2 (Side Sc) 400 Volts, IGAIN=5, PGAIN=5 DELAY=UP PMT3 (525nm) 570 Volts, IGAIN=5, PGAIN=10 DELAY=UP PMT4 (575nm) 700 Volts, IGAIN=5, PGAIN=10 DELAY=UP PMT3 (>600nm) 500 Volts, IGAIN=3, DELAY=UP With this setup "noise" is all below channel 50, and roughly speaking, bacteria are in the 200-500 channel number region and yeast appear around channel 700. I hope some of this helps, Haze ---------------------------------------------------------- | Hazel Marie Davey hlr@aber.ac.uk | |Sefydliad y Gwyddorau Biolegol*Inst. Biological Sciences| |Prifysgol Cymru * University of Wales| | ABERYSTWYTH, Ceredigion, CYMRU / WALES SY23 3DD | | http://pcfcij.dbs.aber.ac.uk/index.htm |
This archive was generated by hypermail 2.1.6 : Thu Jan 01 2004 - 17:35:21 EST
