Flow Cytometry (Principles and Main Applications)
Hossein Asgarian-Omran Ph.D., Immunology Dep. of Immunology, School of Medicine Mazandaran University of Medical Sciences
[email protected]
Workshopp objectives j Getting familiar with flow cytometry system How to interpret flow cytometry cytometry-related related results and
graphs in scientific documents How to use this system in your own projects
History The first fluorescence‐based flow cytometry device (ICP 11): by Wolfgang Göhde (University of Münster, Germany. 1968) The first commercialized by German developer and manufacturer Partec in Göttingen g ((1968‐69)) The first FACS instrument from Becton Dickinson (1974) Original name of flow cytometry technology was: "pulse cytophotometry cytophotometry" 8 years later in 1976, at the Conference in Florida, the name was changed to "flow cytometry", a term that quickly became popular.
Definition Flow Cytometry Flow Cytometry is the process whereby such measurements are made upon cells/particles as they pass through a measuring apparatus suspended in a fluid stream.
FACS Fluorescence Activated Cell Sorter
Basics of Flow Cytometry Fluidics
Cells in suspension flow in single-file single file through an illuminated volume where they
Optics
scatter light and emit fluorescence that is collected collected, filtered and
Electronics
converted to digital values that are stored on a computer
Fluidics Cells are presented t the to th laser l using i principles of hydrodynamic focusing
Sample
Sheath
Flow chamber
Laser optics
L Laser B Beam
Optical Design PMT 6
PMT 5
Sample PMT 4
Fl Flow cell ll
Dichroic Filters
Scatter
PMT 2
PMT 1 Sensor
PMT 3
Laser Bandpass Filters
Optical Filters Dichroic Filter/Mirror at 45 degg Light Source
Transmitted Light
Reflected light
Types of Optical Filters Wavelengths LONG (700nm)
Short Pass
Long Pass
Band Pass
SHORT (500nm)
650 Short Pass (600SP)
550 Long Pass (650LP)
600/100 Band Pass (600/100)
Transmitted
550 nm
550 - 650 nm (600±50)
Blocked
>650 nm
550 nm
