Analyzer Guidelines
The FACScan is a Becton Dickinson Immunocytometry Systems (BDIS) benchtop flow cytometer equipped with a flow cell and an air-cooled Argon gas laser that produces a blue light for excitation at 488 nm. Simultaneous detection of two non-fluorescent parameters , P1 for forward scatter and P2 for side scatter, provide information about relative cell size verses surface morphology and cellular complexity.
Up to three additional parameters enable fluorescent detection of common fluorochromes such as FITC, PE, PerCP or Cychrome for immunophenotyping, GFP or DsRed for tranfection efficiency profiling, and cellular dyes such as PI or 7-AAD for cell viability or cell cycle determinations. Additional studies can be done using other dyes that excite at 488nm and emit in the ranges of 530+/-15nm, 585+/-21nm, or at wave lenghts > 670nm. The newer designed instrument, the FACScalibur, has a second laser that enables a 4th fluorescent range (See FACScalibur below). Cell Quest software is used to collect and analyze data on MacIntosh computers.
| Tips for Sample Preparation Cells should be counted and an appropriate amount of antibody or dye added to facilitate a consistent cell number per amount of antibody or dye per tube. Incubation should be performed in the dark at room temperature or 4 oC. Most samples can be fixed with 1% para-formaldehyde. |
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 The FACSCalibur is a newer BDIS benchtop flow cytometer similarly equipped to the FACScan but with an additional 633nm red diode laser, similar to a HeNe laser. This second laser detects a 4th fluorescent color that enables the use of APC conjugates emitting in the range of 660+/- 10 nm. Cell Quest is used for data acquisition and analysis in FCS 2.0 format on MacIntosh computers. | |
 Sample Port and Flow Cell | |
 Cell Cycle | |
 Cell proliferation | |
 4-Color Immunophenotyping | |
