Cell Sorting Flow Cytometry
Instruments:
Becton and Dickinson FACSAria
Cell Sorter
The FACSAria flow cytometer is used for cell sorting only. For non-sort applications the FACSCaliber is available.
The FACSAria is a benchtop high-speed sorter.with two lasers at 488-nm and 633-nm wavelengths. Digital acquisition rates of up to 70,000 events/second. Multicolor analysis of up to 9 parameters (Forward Scatter, Side Scatter, and 7 Fluorescence). Two- and four-way bulk sorting devices for micotubes, 12X 75 mm and 15 mL tubes. Automated Cell Deposition Unit for sorting to multiwell plates (6,24,48,96, and 384) or microscope slides.
Most dyes designed for the FACSAria are specially manufactured
by BD.We recommend you used the following fluorescent labels (only one dye can be used in each category):
488 PMT A: (750-810 nm)
PE-CY7 (phycoerythrin-Cy7)- Excitation at 488 nm, emission at 767 nm.
488 PMT B: (675-715 nm)
PI (Propidium Iodide)-Excitation at 488 nm, emission at 617 nm
PE-Cy5.5 (phycoerythrin-Cy5.5)- Excitation at 488 nm, emission at 667 nm.
PerCP (peridinin chlorophyll protein)- Excitation at 488 nm, emission at 678
nm.
PerCP-Cy5.5 (peridinin chlorophyll protein)- Excitation at 488 nm, emission at 695
nm.
488 PMT C: (600-620 nm)
PE-TXRed (phycoerythrin-TexasRed) -Excitation at 488 nm, emission at 578 nm.
488 PMT D : (564-606 nm)
PE (phycoerythrin)- Excitation at 488 nm, emission at 578 nm.
488 PMT E: (515-545 nm)
FITC (Fluorescein) - Excitation at 488 nm, emission at 519 nm.
Alexa Fluor 488 - Excitation at 488 nm, emission at 519 nm.
GFP (Green Fluorescent Protein)- Excitation at 488 nm, emission at 508 nm.
633 PMT A : (750-810 nm)
APC-Cy7 (allophycocyanin- Cy7) Excitation at 633 nm, emission at 785 nm.
633 PMT B : (650-670 nm)
APC (allophycocyanin)- Excitation at 633 nm, emission at 660 nm.
AlexaFluor 647- Excitation at 633 nm, emission at 668 nm.
Assisted use is not available you must be a trained to access this instrument.
For information about training, stop by the Microscopy Facility Office (1.426
H) and pick up a training request form and sign up for training seminars. Each trainee must first attend training seminars (no charge). These seminars discuss the theory of flow cytometry and how to obtain high quality data. After you attend these seminars you may then schedule a “hands-on” training with either Angela Bardo. For the “hands-on” training there is a flat fee of $150. Once
training is complete the trained user will be given unassisted access to the
instrument.
Important note: This instrument is housed in a Biosafety Level II room. You may be required to take additional safety courses through UT EH&S before you are trained.
Location:
The FACSCalibur is located in NMS
2.114.
Instrument Supervisor:
Please contact Angela
Bardo for information.
Secondary contacts for this Instrument are
Cecil Harkey and Michelle
Gadush.
Fees:
ICMB charges all users $45.00/ hour
for use of the FACSAria.
Flow Cytometry Information and Links:
Becton and Dickinson:
FACSAria
Information
BD-Pharmingen-Protocols
BD-Fluorchrome Information
General Cytometry Information
Purdue
University Cytomery Labs (PUCL)
PUCL Lecture
Slides
cytomics.info
Clinical Cytometry Scoiety
ISAC Cytometry Page
Protocol
Online
Flow
Cytometry at UT
Cell Cycle Information
Cell Cycle Fundamentals -Written by Nate Regimbal Biogen Idec, Inc. Posted on Purdue Site
Cytometry FreewareWinMDI Version 2.8
This is a very good,
easy to use, software for general analysis. This freeware is provided by Joseph
Trotter of the Scripps Research Institute, La Jolla, CA. Check out Purdue's
WinMDI tutorial.
Cylchred
This is an automatic cell cycle analysis program provided by UWCM in Cardiff,
UK. You will need to use WinMDI to prepare files for analysis.
Cytometry Shareware
FCS Press and FCS Assistant
This software will you let you edit FCS files and create presentation
quality graphics. This software is provided by Ray Hicks and costs about $50,
plus $5 for each additional licence.