Sample Prep and Information for BD LSR Fortessa
The BD LSRFortessa has 16 detectors plus forward- and side-scatter detectors and an additional FSC PMT. The Fortessa is used for analysis only. The Fortessa can analyze up to 16 colors at one time. The Fortessa is also the instrument used for cell cycle, cell proliferation, cell membrane potential, and other analysis. To determine if your fluorochromes can be analyzed on the LSRFortessa, see the table below to determine which fluorochromes may be used based on your fluorochromes excitation and emission.
The BD Biosciences Fluorescence Spectrum Viewer will help you determine whether the fluorochromes you want to use will have a significant overlap.
|Excitation Laser||Detection Filter||Detector||Example|
|488 nm (blue)||488/10 (483-493 nm)||SSC||SSC ONLY|
|695/40 (675-715 nm)||PMT A||PERCP, PERCP5.5, 7AAD, EF710|
|515/20 (505-525 nm)||PMT B||AF488, GFP, FITC|
|561 nm (green)||780/60 (750-810 nm)||PMT A||PE-CY7|
|710/50 (685-735 nm)||PMT B||PC CY5|
|670/30 (655-685 nm)||PMT C||PE CY5.5|
|610/20 (600-630 nm)||PMT D||PE TXRED, PI, MCHERRY, PE CF594|
|582/15 (574-590 nm)||PMT E||PE|
|633 nm (red)||780/60 (750-810 nm)||PMT A||APC-Cy7, EF780|
|720/40 (700-740 nm)||PMT B||AF 700|
|660/20 (650-670 nm)||PMT C||APC, AF 647, EF660|
|407 nm (violet)||780/60 (750-810 nm)||PMT A||BV785, QD800|
|710/50 (685-735 nm)||PMT B||BV 711|
|660/20 (650-670 nm)||PMT C||BV650, QD655|
|610/20 (600-630 nm)||PMT D||BV605, QD605|
|525/50 (500-550 nm)||PMT E||BV510, AMCYAN|
|450/50 (425-475 nm)||PMT F||DAPI, BV421, EF450, PACBLUE|
The Fortessa computer uses FACS Diva software to collect data. These files will be uploaded onto the Flow server for the user to then analyze in FlowJo or similar software. You are able to download FlowJo from their website and the core has a USB dongle you can check out free of charge.
All data files are uploaded to a folder on a UAMS server under the Flow Core.