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About

Microbiology & Immunology Shared Resources

 

The Department of Microbiology & Immunology Flow Cytometry Core assists researchers in flow cytometry based studies. Cell samples can be quickly analyzed based on phenotypic markers and functional assays. Characterization of distinct cell populations based on these techniques has become increasingly important in biomedical research. Flow cytometry is especially useful for physically separating specific sub-populations defined by specific parameters using a fluorescence activated cell sorter (FACS). The microscope core provides researchers the ability to obtain high quality imaging data using state-of-the-art microscopy instruments. 

 

Services include:

  • Analytical flow cytometry for immunophenotyping
  • Flow cytometry for apoptosis and cell cycle
  • Fluorescence-activated cell sorting (FACS)
  • Fluorochrome panel design
  • Flow Analyzer & Cell Sorter Training
  • Confocal Assistance & Training

The shared resource operates three flow cytometers. The LSRFortessa and LSR II are designed for multi-fluorescence analysis while the custom configured FACS Aria II SORP high-speed cell sorter is capable of rapid sorting of cells, bacteria, yeast, and other small particles based on multiple parameter characteristics into highly pure populations. Our two flow cytometers and cell sorter have five lasers capable of simultaneous 18-color detection (i.e. 488nm - 2 PMTs, 405nm - 7 PMTs, 355nm - 3 PMTs, 640nm - 3 PMTs, and 561nm - 3 PMTs) of apoptosis, DNA/cell cycle analysis, detection of novel living color fluorescent proteins (DsRed, mCherry, tdTomato, mOrange, mPlum, et al.), immunophenotyping and calcium influx experiments.

NOTE: Our Aria cell sorter contains only four lasers (405nm, 488nm, 561nm, 640nm) and is no longer equipped with a UV laser (355nm).

 

M&I Flow Core Instrument Configurations

2017 Upgrade

LSR II and LSR Fortessa full 5-laser configuration

Aria configuration lacks 355nm UV laser

Updated LSR II, Fortessa, and Aria II Configurations

      
**Optional Configuration**           (Filter/Configuration Change Required)#
Laser Channel Name PMT Position Dichroic Bandpass  Similar Parameter
(no filter change required)#		 Parameter/Stain Dichroic Bandpass
UV
355 nm
(Absent Aria)		 BUV395 C - 379/28   DAPI, Indo-1 (violet) - 450/50
BUV496 B 450 LP 515/30 Fixable Live/Dead Aqua Indo-1 (blue), Qdot 525 505 LP 530/30
BUV737 A 690 LP 735/35        
VIOLET
405 nm		 BV421 G - 450/50 Pacific Blue, eFluor450, violetFluor450, V450, Alexa405      
BV510 F 505 LP 525/50 AmCyan, Pacific Orange$, V500, Alexa430      
BV570 E 550 LP 575/25 PacificOrange$
*this channel can be difficult
to compensate* 		      
BV605 D 595 LP 610/20 Qdot 605      
BV650 C 635 LP 670/30 Qdot 655      
BV711 B 690 LP 710/50 Qdot 705      
BV785 A 750 LP 780/60 Qdot 800      
BLUE
488 nm		 SSC C - 488/10        
FITC B 505 LP 525/50 GFP, Alexa488, BrilliantBlue515      
PerCP-Cy5.5 A 685 LP 710/50 PE-Cy5.5, PerCP-eFluor710      

YELLOW/ GREEN
561 nm

 PE C - 582/15 DsRED, dTomato      
PE-TexasRed B 600 LP 610/20 PE-eFluor610, PE-CF594, mCherry, 7-AAD, Alexa568      
PE-Cy7 A 750 LP 780/40        
RED
640 nm		 APC C - 670/30 Alexa647      
AF700 B 685 LP 730/45 redFluor 710      
APC-Cy7 A 750 LP 780/60 APC-eFluor780, APC-H7      
#compensations must be made with the same conjugate/stain used within each experiment
 **For DAPI/Indo-1 please see Flow Core Manager about filter/configuration change**
$due to the broad emmision spectrum of Pacific Orange, both channels will detect this fluor

 

The flow cytometric separation of cells from unfixed animal tissue is possible on the BD FACSAria cell sorter, which is housed in a Baker BioProtect IV biohazard cabinet. For unfixed human specimens, please contact Ryan Wantroba. Before the initial sort is performed, each user is required to meet with the core manager to discuss your cell type, origin of sample, and special requirements.

 

In additional to these systems, the facility also has a state-of-the-art Zeiss LSM710 ConfoCorr confocal imaging microscope system, a Zeiss AxioVert microscope, and a Typhoon Trio imaging system. 

 

Leadership

 Ryan Wantroba
Flow Core Manager
Phone: 203-233-8665
Email: rmw2178@cumc.columbia.edu

 Nicholas Arpaia, Ph.D.
Director
Phone: 212-305-1379
Email: na2697@cumc.columbia.edu

  James Lapin
Dept. Administrator
Phone: 212-305-4046
Email: jl2787@cumc.columbia.edu

  Carol Duigou
Accounts Manager
Phone: 212-305-3647
Email: ced4@cumc.columbia.edu

 

Location and hours of operation

Hours Location

Staffed 
Monday through Friday 9 AM to 5 PM

Trained Users
24/7 access through a swipe card system.

Department of Microbiology & Immunology
Flow Cytometry Core
Columbia University
701 W. 168 St., HHSC 1211A & 1201
New York, NY 10032

Links and Resources

htttp://www.microbiology.columbia.edu/fcc

Contacts

Name Role Phone Email Location
Microbiology & Immunology Shared Resources
Managers
 
212-342-4089
 
microimmuno-facs@cumc.columbia.edu
 
HHSC 1211-A