Methods Section of the Paper
Please use the charts below to create the configuration specific to your application. Following is an exampleof a typical description. Please contact the lab personnel if you wish assistance with the wording:
Two-color flow cyotmetric analysis was performed using a FACScan flow cytometer (BDBiosciences, San Jose, CA) equipped with an air-cooled 15mW argon ion laser tuned to 488nm. The emission fluorescence of (yada, yada conjugated to IgG…blah blah..) was detected and recorded through a 530/30 bandpass filter in the FL1 channel. (Something red) was detected in the FL2 channel through a 585/42 bandpass filter. List mode data files consisting of 10,000 events gated on FSC (forward scatter) vs SSC (side scatter) were acquired and analyzed using CellQuest PRO software (BD Biosciences, San Jose, CA) at a rate of 200-400 events per second. Appropriate electronic compensation was adjusted by acquiring cell populations stained with each dye/fluorophore individually, as well as an unstained control.
BD Biosciences FACSAria Configuration
Excitation Laser |
Emission Detection Filter |
Detector |
Example (there are many others) |
488 nm (blue) |
1.5 ND Filter |
FSC Diode |
|
780/60 (750-810 nm) |
PMT A |
PE-Cy7 |
|
695/40 (675-715 nm) |
PMT B |
PerCP, PE- Cy5.5 |
|
610/20 (600-620 nm) |
PMT C |
PE-Texas Red |
|
576/26 (563-589 nm) |
PMT D |
PE |
|
530/30 (515-545 nm) |
PMT E |
FITC |
|
488/10 (483-493 nm) |
PMT F |
SSC only |
|
633 nm (red) |
780/60 (750-810 nm) |
PMT A |
APC-Cy7 |
660/20 (650-670 nm) |
PMT B |
APC |
|
407 nm (violet) |
530/30 (515-545 nm) |
PMT A |
Alexa-430 |
450/40 (430-470 nm) |
PMT B |
DAPI |
BD Biosciences FACScan Configuration
Excitation Laser |
Emmision Detection Filter |
Detector |
Example |
488 nm (blue) |
1.0 ND Filter |
FSC Diode |
|
|
488/10 (483-493 nm) |
SSC Detector |
|
530/30 (515-545 nm) |
FL1 Detector |
FITC |
|
585/42 (564-606 nm) |
FL2 Detector |
PE |
|
650 LP (650-800 nm) |
FL3 Detector |
PerCP |
