Hello ! I just wanted to share with you the suggestions I received from the mailing list users for the bacterial sort on the Vantage SE DIVA. After trying some of the suggestions I finally had success. However is not very straightforward to do this. In my case, to get rid of the noise (on the oscilloscope) that was caused by turning on the drop drive, the solution was to increase the frequency and sheath pressure,(62.3 kHZ, 34 psi, 70 um nozzle) then adjust the FSC and SSC obscuration bars. To get rid of the left over noise on the digital (DIVA) display I used FSC in linear and trigger in both FSC and SSC. I had to play a lot with the threshold , specially on FSC to be able to get rid of the noise on FSC and still see the bacteria. The neutral density filter was taken out. After doing all that I had a successful sort. Thanks to all of you who gave me ideas and suggestions. Johana Johana Melendez, MS Research Associate Flow Cytometry Laboratory H. Lee Moffitt Cancer Center 12902 Magnolia Dr. Tampa, Florida 33612-9497 Phone (813) 745-6610 Lab. phone (813) 972-8400 x. 2005 <<cytometry mailing list summary.doc>> Fax (813) 903-7140 <<Melendez, Johana.vcf>> >This was my previous e-mail: I would like to know if anybody out there that has done bacterial cell sorting can be kind enough to share the sort settings used on their machines. We have a Vantage SE DIVA and we are trying to sort E.coli based on porphyrin expression. In terms of the optics configuration, it looked OK, like we can distintinguish the bacteria from the noise, no problem, but as soon as I turn the ddf on I can't get rid of the noise. (By the way the neurtal density filter was removed as suggested by Mark) (The first try was done with the 70 um nozzzle, at 49200Hz, 31 psi. , (Hey, I know I should have tried a smaller one, but I guess we wanted to test the limits ...:)) , and I wonder if using a smaller size nozzle or using other settings can help on this? So, If anybody can provide suggestions as to which size nozzle, frequencies and buffers to use we will greatly appreciate it. Thanks in advance ----------------------------------------- ########################################################################### ## This transmission may be confidential or protected from disclosure and is only for review and use by the intended recipient. Access by anyone else is unauthorized. Any unauthorized reader is hereby notified that any review, use, dissemination, disclosure or copying of this information, or any act or omission taken in reliance on it, is prohibited and may be unlawful. If you received this transmission in error, please notify the sender immediately. Thank you. ######################################### #################################### This attachment - 'cytometry mailing list summary.doc' - 55.81 KBytes - can be viewed at http://www.cyto.purdue.edu/MD-parts/6b7fd8db914bda1cda3db52910abcb042b050e48.doc
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