What we do

ISAC Recognized Shared Resource Laboratory

The Flow and Image Cytometry Shared Resource (FICSR) at Roswell Park Comprehensive Cancer Center is a state-of-the-art facility providing advanced flow cytometric and morphology service at cellular and subcellular levels of resolution. The facility is one of 8 world-wide shared resource laboratories (SRL) recognized by the International Society for Advancement of Cytometry (ISAC) for operational excellence and adherence to best practices

The services offered by the Flow and Image Cytometry Shared Resources include:

  • Investigator access to cytometry equipment 24/7
  • Education and consultation services
  • Comprehensive sample processing, data acquisition, data analysis and reporting
  • Morphological services at the cellular and subcellular levels of resolution
  • Luminex multiplex bead array-based cytokine, chemokine and growth factor quantification
  • Core flow cytometry services for investigator and biotech sponsored clinical trials

Flow cytometry instrumentation and services

Our Flow and Image Cytometry Shared Resource (FICSR) facility houses five Becton Dickinson (BD) flow cytometry analyzers ( 2 LSRIIs, 3 Fortessas), a Beckman Coulter Cytoflex LX, two Amnis/Luminex ImageStreamX-MKII imaging flow cytometers, 3 flow cytometry sorters (BD Aria-I, Aria-II and a Sony MA900), a full spectrum flow cytometry analyzer (Cytek Aurora), a Luminex 200 bead array analyzer, a Zetaview small particle analyzer and a confocal microscope. These instruments are accessible 24/7 to all trained investigators.

Researcher working with equipment

During weekdays, our staff performs start-up and daily QC on the analyzers. During weekends and on holidays, users are responsible for these procedures. Investigators are expected to work in the facility independently, but FICSR staff is available during regular business hours for consultation. Our facility, including the user-accessible instrument room, is a designated BSL2+ area, therefore applicable biosafety precautions should be adhered to including wearing personal protection devices such as lab coats and gloves.

Users can reserve time on the equipment and monitor availability of the equipment using the online FOM calendar. Individual access to this system is granted only after completion of the free Introduction to Flow Cytometry class (sign up). The class outlines the guidelines and requirements for use of the facility and gives a basic overview of the available technology. Further one-on-one training to become proficient on the instruments is available by appointment with our staff.

Instrument use is charged by actual usage time, or 80% of calendar reservation time, whichever is greater. Reservations can be altered up to two hours before the start time. Reservations that do not begin within 30 minutes of the reservation start time will be released and charged at 80% of reserved time. A reduced rate is offered for evening or overnight hours (8 p.m. to 8 a.m.), weekends and holidays.

Custom flow cytometers

The Flow Cytometry Electronics Shop utilizes an optical bread board, supporting digital acquisition electronics, fluidics and cuvette flow cell for custom flow cytometric experiments.

Custom technology available for studies include:

  • Contiguous multispectral detection using diffraction grating optics
  • Infra-red flow cytometry
  • Fluorescence detection beyond 1100 nm
  • Optical breadboard flow cytometer open to diverse configurations

There are 20 detection parameters each with 262,144 channel (18 bit) resolution and 5-1/2 decade dynamic range electronic processing. Various laser lines also available for excitation.

Researchers interested in using this technology should contact _________.

Flow cytometry analyzers

Technical specifications

  • Five lasers, 18 color detection + 2 scatter parameters (from 488nm)
  • PC Windows 7, DiVa version 8.0.1
Laser Bandpass Filter Common Fluors (list is not exhaustive)
355 nm UV laser U379/28 BUV395, Indo-1
U460/14 DAPI, Hoechst 33342, LD Blue, Zombie UV
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V530/30 BV510, V500, AmCyan, LD Aqua
V610/20 BV605
V660/20 BV650
V710/50 BV711
V780/60 BV750, BV786, BV785
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B695/40 PerCPCy5.5, BB700, PerCP, 7AAD
561 nm yellow/green laser Y582/15 PE, mOrange, AF568, dTomato, PI
Y610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, RFP, dsRed, mCherry, PI
Y670/30 PECy5, PECy5.5, 7AAD, PI
Y710/50 PEeFluor710
Y780/60 PECy7, PEAF750
640 nm red laser R660/20 APC, AF647
R730/45 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical specifications

  • Four laser, 16 color detection + 2 scatter parameters (from 488nm)
  • PC Windows 7, FACSDiva version 8.0.1
Laser Bandpass Filter Common Fluors (list is not exhaustive)
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V525/50 BV510, V500, AmCyan, LD Aqua
V610/20 BV605
V660/20 BV650
V710/50 BV711
V780/60 BV750, BV786, BV785
488 nm blue laser B525/50 FITC, BB515, GFP, AF488, CFSE
B695/40 PerCPCy5.5, BB700, PerCP, 7AAD
561 nm yellow/green laser Y585/15 PE, mOrange, AF568, dTomato, PI
Y610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, RFP, dsRed, mCherry, PI
Y670/30 PECy5, PECy5.5, 7AAD, PI
Y710/50 PEeFluor710
Y780/60 PECy7, PEAF750
640 nm red laser R670/30 APC, AF647
R730/45 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical specifications

  • Three laser, 16 color detection + two scatter parameters (from 488nm)
  • Hydrodynamically focused sample interrogated within a quartz cuvette flow cell
  • (HTS ready, has internal components allowing a HTS to be connected, HTS software option enabled)
Laser Bandpass Filter Common Fluors (list is not exhaustive)
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V525/50 BV510, V500, AmCyan, LD Aqua
V560/20 Qdot565, BV570
V585/15 BV570
V610/20 BV605
V660/20 BV650
V710/50 BV711
V780/60 BV750, BV786, BV785
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B575/26 PE, mOrange, AF568
B610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, PI
B695/40 PerCPCy5.5, BB700, PerCP, PECy5, PECy5.5 7AAD, PEeFluor710
B780/60 PECy5, PEAF750
640 nm red laser R670/14 APC, AF647
R730/45 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical specifications

  • Four laser, 18 color detection + two scatter parameters (from 488nm)
  • PC Windows 7, DiVa version 8.0.1
Laser Bandpass Filter Common Fluors (list is not exhaustive)
355 nm UV laser U379/28 BUV395, Indo-1
U460/14 DAPI, Hoechst 33342, LD Blue, Zombie UV
U740/35 BUV737
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V525/50 BV510, V500, AmCyan, LD Aqua
V585/15 BV570
V610/20 BV605
V660/20 BV650
V710/50 BV711
V780/60 BV750, BV786, BV785
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B575/26 PE, mOrange, AF568
B610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, PI
B695/40 PerCPCy5.5, BB700, PerCP, 7AAD
B780/60 PECy7, PEAF750
640 nm red laser R670/14 APC, AF647
R730/45 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical specifications

  • Four laser, 18 color detection + two scatter parameters (from 488nm)
  • PC Windows 7, DiVa version 8.0.1
Laser Bandpass Filter Common Fluors (list is not exhaustive)
355 nm UV laser U379/28 BUV395, Indo-1
U460/14 DAPI, Hoechst 33342, LD Blue, Zombie UV
U740/35 BUV737
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V525/50 BV510, V500, AmCyan, LD Aqua
V585/15 BV570
V610/20 BV605
V670/30 BV650
V710/50 BV711
V780/60 BV750, BV786, BV785
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B575/26 PE, mOrange, AF568
B610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, PI
B710/50 PerCPCy5.5, BB700, PerCP, PECy5, PECy5.5, 7AAD, PEeFluor710
B780/60 PECy7, PEAF750
640 nm red laser R670/30 APC, AF647
R710/50 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical Specifications

  • 6 laser, 21 color detection + two scatter parameters (from 488nm)
  • PC Windows 10, CytExpert version 2.4
Laser Bandpass Filter Common Fluors (list is not exhaustive)
375 nm near UV laser NUV450/45 BUV395
NUV525/40 BUV496, Indo-1
NUV675/30 BUV661
405 nm violet laser V450/45 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V525/40 BV510, V480, V500, AmCyan, LD Aqua
V610/20 BV605
V660/10 BV650
V763/43 BV750, BV786
488 nm blue laser B525/40 FITC, BB515, GFP, AF488, CFSE
B610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD
B690/50 PerCPCy5.5, BB700, PerCP
561 nm yellow/green laser Y585/42 PE, mOrange, AF568, dTomato
Y610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, RFP, dsRed, mCherry, PI
Y675/30 PECy5, PECy5.5, 7AAD, PI
Y710/50 PEeFluor710
Y763/43 PECy7, PEAF750
638 nm red laser R660/10 APC, AF647
R712/25 AF700, APC-R700, DRAQ5
R763/43 APC-Cy7, APC-AF750, APCH7
808 nm near IR laser IR840/20 AF790, LD Near-IR
IR885/40 LD Near-IR, ViaKrome 808, PromoFluor 840, iFluor 840

Technical specifications

  • 5 laser, 40+ color detection + three scatter parameters (FSC from 488nm and SSC from 488 or 405nm)
  • PC Windows 10, SpectroFlo version 3
Laser Channel Bandpass Filter Common Fluors (list is not exhaustive)
355 nm UV laser UV1 NUV373/15
UV2 NUV388/15 BUV395
UV3 NUV428/15
UV4 NUV443/15
UV5 NUV458/15
UV6 NUV473/15
UV7 NUV542/28 BUV496
UV8 NUV542/28
UV9 NUV582/31 BUV563
UV10 NUV613/31
UV11 NUV664/27 BUV661
UV12 NUV692/28
UV13 NUV720/29 BUV737
UV14 NUV750/30
UV15 NUV780/30
UV16 NUV812/34 BUV805
405 nm violet laser V1 V428/15 BV421
V2 V443/15 AF405, SB436
V3 V458/15 PacBlue, eFluor450
V4 V473/15 BV480
V5 V508/20 eFluor506
V6 V525/17
V7 V542/17 BV510
V8 V581/19 BV570, PacOrange
V9 V598/20
V10 V615/20 BV605, SB600
V11 V664/27 BV650, SB645
V12 V692/28
V13 V720/29 BV711, SB702
V14 V750/30 BV750
V15 V780/30 BV785, BV786
V16 V812/34
488 nm blue laser B1 B508/20 BB515
B2 B525/17 FITC, AF488
B3 B542/17 AF532
B4 B581/19
B5 B598/20
B6 B615/20
B7 B661/17
B8 B679/18 PerCP
B9 B697/19 BB700, PerCPCy5.5
B10 B717/20 PerCP-eFluor710
B11 B738/21
B12 B760/23
B13 B783/23
B14 B812/34
561 nm yellow/green laser YG1 YG577/20 PE
YG2 YG598/20
YG3 YG615/20 PEDazzle594, PECF594, PE-AF610, PE-eFluor610
YG4 YG661/17
YG5 YG679/18 PECy5
YG6 YG697/19
YG7 YG720/29 PECy5.5, PE-AF700
YG8 YG750/30
YG9 YG780/30 PECy7
YG10 YG812/34
638 nm red laser R1 R661/17 APC
R2 R679/18 AF647, eFluor660
R3 R697/19
R4 R717/20 AF700, APC-R700
R5 R738/21
R6 R760/23
R7 R783/23 APCCy7, APCH7, APC-AF750, APCFire750
R8 R812/34

The ZetaView small particle analyzer employs Nanoparticle Tracking Analysis (NTA) which relates the rate of Brownian motion of the particles in liquid suspension to particle size. The rate of movement is related to the viscosity and temperature of the liquid; it is not influenced by particle density or refractive index. NTA allows the determination of a size distribution profile of small particles with a diameter of approximately 10-1000 nanometers (nm) in liquid suspension. Additionally, the ZetaView is also capable of measuring the so-called zeta-potential of the particles which is a measure of the electric charge on the nanoparticle surface.

Our facility has two ZetaView analyzers which share a single flow cell module interchangeable between the two analyzers. One analyzer is equipped with a 488nm laser while the other is equipped with 405nm laser. For particle size, concentration, and zeta potential measurements, both analyzers can be used interchangeably. When detection of fluorescent particles is desired, the analyzer of choice needs to match the excitation spectrum of the intended fluorochrome to be detected.

The primary application of the NTA analysis at Roswell Park is in the field of extracellular vesicles (EV) research where NTA is the accepted method for determining size distribution and concentration of EVs. Exosomes are cell-derived vesicles with a diameter between 30 and 100 nm. Evidence is accumulating that exosomes have specialized functions and play a key role in intercellular communication and can potentially be used for prognosis, for therapy, and as biomarkers for health and disease. Amongst the many detailed studies of exosomes that can include studying their phenotype and cargo, measurement of their size and concentration are additional key parameters in the context of their biological function.

The ZetaView instruments are QC’d prior to reservations and are available 24/7 to trained users by reserving time on the FOM calendar. 

Fluorochrome Filter available
BUV395 379/28
BUV737 740/35
BV605 610/20
BV650 660/20
BV711 710/50
Calcium Flux 405/20
Side Population 645LP
YFP 546/20
PerCP 670/LP

Technical Specifications

  • 12 channel, 2 camera instrument
  • Five lasers, up to 10 fluorescent parameters with two brightfield images
  • Side scatter available in either channel 6 or channel 12 (from dedicated 785nm scatter laser)
  • 20x, 40x, 60x magnification
  • Extended Depth of Field (EDF, 16 µ) and autosampler option (96-well plate format)
Camera 1 Bandpass Filter Common Fluors (list is not exhaustive)
488 nm blue
561 nm yellow/green
592 nm orange
Ch1: 420-480 nm Brightfield
Ch2: 480-560 nm FITC, BB515, GFP, AF488, CFSE
Ch3: 560-595 nm PE, AF568
Ch4: 595-642 nm PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, AF594
Ch5: 642-745 nm PerCPCy5.5, BB700, PerCP, PECy5, PECy5.5, 7AAD, PEeFluor710
Ch6: 745-800 nm PECy7, PEAF750
Camera 2 Bandpass Filter
405 nm violet
640 nm red
Ch7: 420-505 nm BV421, BV480, DAPI, Pacific blue, V450, LD Violet
Ch8: 505-570 nm BV510, V500, AmCyan, LD Aqua
Ch9: 570-595 nm Brightfield
Ch10: 595-642 nm BV605
Ch11: 642-745 nm AF647, APC
Ch12: 745-800 nm APC-Cy7, APC-AF750, APCH7

Technical Specifications

  • 12 channel, 2 camera instrument
  • Four lasers, up to 10 fluorescent parameters with two brightfield images
  • Side scatter available in either channel 6 or channel 12 (from dedicated 785nm scatter laser)
  • 20x, 40x, 60x magnification
Camera 1 Bandpass Filter Common Fluors (list is not exhaustive)
488 nm blue
561 nm yellow/green
Ch1: 420-480 nm Brightfield
Ch2: 480-560 nm FITC, BB515, GFP, AF488, CFSE
Ch3: 560-595 nm PE, AF568
Ch4: 595-642 nm PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, AF594
Ch5: 642-745 nm PERCPCy5.5, BB700, PerCP, PECy5, PECy5.5, 7AAD, PEeFluor710
Ch6: 745-800 nm PECy7, PEAF750
Camera 2 Bandpass Filter
405 nm violet
640 nm red
Ch7: 420-505 nm BV421, BV480, DAPI, Pacific blue, V450, LD Violet
Ch8: 505-570 nm BV510, V500, AmCyan, LD Aqua
Ch9: 570-595 nm Brightfield
Ch10: 595-642 nm BV605
Ch11: 642-745 nm AF647, APC
Ch12: 745-800 nm APC-Cy7, APC-AF750, APCH7

New accounts with the Flow and Image Cytometry Facility

Our facility uses FOM (Facility Online Manager) as an integrated calendar, instrument sign-on, and billing system.

Every new user will need to set up an individual account in FOM. Instructions for Roswell Park users can be found in the FOM Internal New User Instruction Document, and users not affiliated with Roswell Park will need to follow the instructions in the FOM New User Instruction Document.

Each lab (or company) will also need a group account for logging in to the computer software on our analyzer instruments. Only one account will be created per PI. In order to create this account for a lab new to our facility, please fill out the New Cytometer Account Form.

Users new to flow cytometry or any specific instrumentation in our facility are requested to contact us to set up an appointment during regular business hours, so that we can properly assist you with setting up your experiments. Several free courses are available to help you gain proficiency on the instruments. For independent use of the flow cytometers, it is mandatory to attend our class Introduction to Flow Cytometry.

For general information on new accounts, contact Orla Maguire at 716-845-5890.

Analysis software

FACSDiVa is BD Bioscience's next generation data acquisition and analysis program and operates in the PC Windows environment on the FACSAria, FACSCanto, LSRII and Fortessa.

ModFit is a flow cytometry-based DNA analysis software that operates in the PC Windows or Mac environment. It is designed to read FCS files acquired on any model flow cytometer. DNA cell cycle and proliferation analysis can be performed using this software. 

Vendor Website

FCS Express is a flow cytometry analysis software that operates in the PC Windows or Mac environment. It is designed to read FCS files acquired on any model flow cytometer. Roswell Park has a network license for version 7.0.

Vendor Website

OMIQ is a cytometry analysis platform encompassing machine learning and analytical pipelines with conventional manual analysis. It allows researchers to complete their entire workflow from raw data to statistical significance in one platform.

Vendor Website

WinList is a flow cytometry analysis software that operates in the PC Windows environment. It is designed to read .fcs files acquired on any model flow cytometer. Roswell Park has network licenses for versions 7.1, 8.0, and 9.0. Downloading the most recent version (9.0) is recommended. 

Vendor Website

Imaging software

IDEAS is the software package used to analyze ImageStream data. Since efficient ImageStream data analysis demands ample computer storage space, powerful processing speed, and fast network connections, dedicated 64-bit, dual quad-core workstations with 24GB of RAM and gigabit network connections are set-up at the facility.

Currently, there are four workstations with IDEAS v6.2, access to them is free of charge and reservations are not required. The software has options to export calculated image features in FCS format, which can be read by conventional flow cytometry software and has options to export analysis statistics in TXT format for import into graphing programs. In addition to support provided by the facility, the vendor also provides support to registered users. Registration is free and is available here.

Leica AF is the software package that enables image capture and image analysis of files captured by the Leica SP8 confocal microscope and the Leica AF6000 live cell imager. Files can be saved in the proprietary Leica .lif and .lei formats or can be exported as TIFF or JPG files suitable for analysis with third-party image analysis software such as ImagePro or ImageJ.

Amnis AI works with the IDEAS 6.3 analysis software to help researchers classify objects using computer-aided tagging and AI algorithms. The software creates a model of all the cell classes in the base population without the need for masking or feature calculation. It can also help identify cell differences that may be difficult to measure using traditional image analysis approaches. More information can be found here.

Currently, there is one workstation with Amnis AI and IDEAS v6.3, access to it is free of charge. Inquiries about use of the software can be made to Hans Minderman at 716-845-1162.

Leica AF Lite is a version of the LAF software with limited functionality but is freely down loadable from the vendor website. It primary use is to view files that were saved in the proprietary LIF and LEI formats

Image Pro Plus 6.3 is a software package for advanced image analysis. It accepts a wide variety of image file formats. The software is installed on all imaging workstations in the facility but since we only have one license which is controlled by a dongle it will only work on one station at a time.

Vendor Website

ImageJ is a popular Java-based free-ware image analysis package developed at the National Institutes of Health. Custom acquisition, analysis and processing plugins can be developed using ImageJ's built-in editor and a Java compiler.

ImageJ can be run as an online applet, a downloadable application, or on any computer with a Java 5 or later virtual machine. Downloadable distributions are available for Microsoft Windows, Mac OS, Mac OS X, Linux, and the Sharp Zaurus PDA.

ImageJ can display, edit, analyze, process, save, and print 8-bit color and grayscale, 16-bit integer and 32-bit floating point images. It can read many image formats including TIFF, PNG, GIF, JPEG, BMP, DICOM, FITS, as well as raw formats. ImageJ can calculate area and pixel value statistics of user-defined selections and intensity thresholded objects.

Cell sorting

The Flow and Image Cytometery Shared Resource (FICSR) houses three cell sorter instruments in a BSL2+ facility. Two of these are available for operator-assisted sorting, each with a slightly different configuration, but otherwise similar capabilities.

The BD Aria I, upgraded in 2010, is equipped with 12 fluorescent PMTs and 405, 488, 561, and 640 nm lasers.

The Aria II is equipped with four lasers (355nm, 405nm, 488nm, and 633nm) and has 13 fluorescent detectors. Both instruments have the capacity to perform four-way sorts, as well as sorts into plates and onto slides. Both instruments are placed in a Bio-Bubble biocontainment system and have additional aerosol containment capabilities.

The Aria instruments are maintained and operated by dedicated facility staff members and are available during regular business hours by reservation. In a typical workflow, researchers will bring their stained single-cell suspensions to FICSR along with controls, buffers, and collection tubes. Then, together with the operator, the experiment is set up and sort logic is defined. After setup, researchers will not need to be present for the remainder of the sort duration. There are training options for individuals who would like to run these sorters unassisted.

A third, more user-friendly flow cytometry sorter, the SONY MA900, is available for unassisted use 24/7 following training by our staff. This instrument is equipped with 12 fluorescent PMTs and 405, 488, 561, and 638 nm lasers, and is placed in a biosafety cabinet. The instrument has the capacity to do four-way tube sorts as well as sorts into plates.

The sorter schedules can be found on our Facility Online Manager (FOM) calendar, and an active FOM account is required for access to the sorters. Reservations for the Aria sorters can only be scheduled by the FICSR sorter operators.

For sorts that have not been performed previously, please submit the online Sort Information Form to request time. The operators will contact you to confirm your appointment and to discuss further specifics. Investigators new to sorting or embarking on a new project are encouraged to discuss their goals with FICSR staff prior to scheduling time.

If you are requesting a repeat of a sort that has been done before in our facility, a short Sort Appointment Request form can be submitted. The FICSR staff will contact you to finalize the appointment details. Appointments are not guaranteed until confirmed and entered in FOM.

Reservations for the SONY sorter can be placed on FOM by approved users after submission of the Sort Information Form to relay safety concerns.

Detailed information pertaining to our operation routine, reservations, cancellation policies, sample requirements, and collection options can be found in our document “Guidelines to Sorting”.

In addition to flow sorting capabilities, our facility also hosts a Miltenyi Classic AutoMACS for automated magnetic cell sorting. Reservations for this instrument are available to all trained researchers through FOM.

Contact Information: 

Orla Maguire: 716-845-5890
Xiaojun Liu: 716-845-1745  
Roswell Park Campus Location: Cancer Cell Center, Room C315

Cell sorting instrumentation

Our BSL2+ room in the Cancer Cell Center, Room C315, houses the two BD Aria cell sorter instruments and the SONY MA900.

The BD Aria I was upgraded in 2010 and is equipped with 12 fluorescent PMTs over four lasers (405 nm violet, 488 nm blue, 561 nm yellow-green, and 640 nm red). This configuration is similar to the LSRII B analyzer, but with less options for the violet laser. The instrument operates with a 100-micron nozzle/20 PSI or 70 micron nozzle/35 PSI configuration. Detailed specifications can be found below. The Aria I is placed in a Bio-Bubble negative-pressure biosafety cabinet that provides the BSL2+ environment to protect the operator.

The BD Aria II is equipped with four lasers (355 nm UV, 405 nm violet, 488 nm blue, and 633 nm red) and has 13 fluorescent detectors. This configuration is similar to the Fortessa B analyzer, but with less options for the violet laser. Current options for configuration on the Aria II are 100/20; 85/45; 85/30; 70/35, and 130/10 (nozzle size/PSI). Detailed specifications can be found below. The Aria II is placed in a Bio-Bubble negative-pressure biosafety cabinet that provides the BSL2+ environment to protect the operator.

The SONY MA900 was installed in 2019 and is equipped with 2 coaxial laser combinations, 488/561 nm and 405/638 nm, for detection of up to 12 fluorescent colors. The instrument operates with sorting chips that come in 70, 100 and 130-micron nozzle sizes, and setup is automated so that the user requires minimal training. Further detailed specifications can be found below.

All instruments have the capacity to do one- to four-way sorts into a variety of tube styles, such as 12x75 mm tubes of any material, Eppendorfs, PCR tubes and 15-ml conical tubes (two-way only). They can also sort into plates (6, 12, 24, 48, 96, and 384 wells).

Our cell sorters undergo a stringent routine maintenance and cleaning protocol to warrant aseptic sorting conditions at all times. We use sterile-filtered HEPES-buffered saline with EDTA as sheath fluid.

For magnetic cell sorting, our facility houses a Miltenyi Classic autoMACS. This instrument is located in a biosafety cabinet in the Cancer Cell Center, Room 322, and does not require operator assistance. Columns, clean bottles, and cleaning solution are provided. The user is responsible for running buffer and beads. Daily usage charges apply.

Technical specifications

  • Sort fractions collected jet-air
  • Four laser - separate interrogation of sample for each laser, 14 parameters: 12 fluorescent and two scatter parameters + time
  • Sheath pressure is adjustable from 2 to 75 psi
  • At 70 psi and 90 kHz with an average threshold rate of 25,000 events per second for a four-way sort results in a purity of > 98% and a yield > 80% of Poisson's expected yield
  • Sample Collection Cooling 4°, 20°, 37°, and 42°C (Water recirculator for refrigeration/heating)
  • Aerosol Management Option (AMO) with Bio-Bubble negative pressure. Bio Safety Level 2+ environment
Laser Bandpass Filter Common Fluors (list is not exhaustive)
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V530/30 BV510, V500, AmCyan, LD Aqua
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B710/50 PerCPCy5.5, BB700, PerCP, 7AAD
561 nm yellow/green laser Y585/15 PE, mOrange, AF568, dTomato, PI
Y610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, RFP, dsRed, mCherry, PI
Y670/30 PECy5, PECy5.5, 7AAD, PI
Y710/50 PEeFluor710
Y780/60 PECy7, PEAF750
640 nm red laser R660/20 APC, AF647
R730/30 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

Technical specifications

  • Sorting chip-based fluidics
  • Available nozzle sizes: 70, 100, and 130 micron
  • Automatic calibrations for chip alignment, laser delay, droplet formation, sort delay, and side stream using setup beads
  • Four lasers – two laser intercepts: coaxial interrogation for 488/561 laser lines and 405/638 laser lines
  • 14 parameters: 12 fluorescent and two scatter parameters + time
  • Fixed sheath pressure for each nozzle size
  • Sample collection cooling (5°C) by Peltier coupling
  • Sample chamber cooling (5°C) or heating (37°C)
  • Aerosol Management System (AMS) of flow chamber plus instrument containment in full
Laser Bandpass Filter Common Fluors (list is not exhaustive)
488 nm blue laser
561 nm yellow/green laser
525/50 FITC, BB515, GFP, AF488, CFSE
585/30 PE, mOrange, AF568
617/30 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, PI
695/50 PerCPCy5.5, BB700, PerCP, PECy5, PECy5.5, 7AAD, PEeFluor710
785/60 PECy7, PEAF750
405 nm violet laser
640 nm red laser
450/50 BV421, BV480, DAPI, Pacific blue, V450, EF450, SB436, AF405, LD Violet
525/30 BV510, V500, AmCyan, LD Aqua
585/30 BV570
617/30 BV605
665/30 APC, AF647, BV650
720/60 AF700, APC-R700, DRAQ5
785/60 APC-Cy7, APC-AF750, APCH7, BV785, BV786

Technical specifications

  • Sort fractions collected jet-air
  • Four laser, 15 parameters: 13 fluorescent and two scatter parameters + Time
  • Sheath pressure is adjustable from 2 to 75 psi.
  • At 70 psi and 90 kHz with an average threshold rate of 25,000 events per second for a four-way sort results in a purity of > 98% and a yield > 80% of Poisson's expected yield
  • Aerosol Management Option (AMO)
Laser Bandpass Filter Common Fluors (list is not exhaustive)
355 nm UV laser U379/28 BUV395, Indo-1
U460/14 DAPI, Hoechst 33342, LD Blue, Zombie UV
405 nm violet laser V450/50 BV421, BV480, DAPI, Pacific Blue, V450, EF450, SB436, AF405, LD Violet
V530/30 BV510, V500, AmCyan, LD Aqua
488 nm blue laser B530/30 FITC, BB515, GFP, AF488, CFSE
B575/25 PE, mOrange, AF568
B610/20 PECF594, PEAF610, PEDazzle594, PEeFluor610, ECD, PI
B710/50 PerCPCy5.5, BB700, PerCP, PECy5, PECy5.5, 7AAD, PEeFluor710
B780/60 PECy7, PEAF750
640 nm red laser R660/20 APC, AF647
R730/30 AF700, APC-R700, DRAQ5
R780/60 APC-Cy7, APC-AF750, APCH7

The autoMACS Separator is a benchtop instrument for high-speed automated cell sorting. Employing the MACS® Technology, the autoMACS Separator is designed for positive selection as well as depletion of magnetically labeled cells. The autoMACS Separator is operated with pre-set separation programs, thus, allowing optimization of cell sorting approaches according to cell abundance and the intensity of marker expression. The separated cells are immediately ready for experiments, cell analysis, or further subset sorting.

  • Fast — up to 4×109;magnetically labeled cells are sorted automatically within a few minutes.
  • Efficient — rare cells can be enriched up to 10,000-fold or unwanted cells may be depleted up to 99%.
  • MACS® Cell Separation Technology is available for human, mouse, rat and non-human primate cell isolation. For any other cell type MicroBeads for indirect magnetic labeling can be used.
  • The separated cells remain viable and their functionality is not compromised. 
  • For a full repertoire of available analytes please see the Miltenyi Biotec website: www.miltenyibiotec.com.
  • AutoMACS applications contacts:
    • Orla Maguire: ext 8590
    • Hans Minderman: ext. 1162

Cell imaging services

Confocal microscopy

Our Leica TCS SP8 Laser Scanning Spectral Confocal Microscope was put in operation in early 2018. The confocal has four laser lines (405nm, 488nm, 552nm, and 638nm) and is equipped with 10X dry, 20X dry, and 63X oil immersion objectives. Further specifications are detailed below.

First-time users of the Leica TCS SP8 confocal microscope must contact the principal operator Meghana Bapardekar at 716-845-3470 to schedule a new user training session (~45 minutes) before working independently on the microscope. All users should reserve time on the microscope using the FOM calendar system. Operator assistance is available by scheduling an appointment during regular business hours.

Live cell imager

The Leica AF6000LX live cell imaging system enables time-lapse videography of cell cultures in a temperature- and CO2-controlled environment. The microscope stage is adaptable to receiving multi-well plate formats as well as slide-based formats (including multi-well chamber slides). The software enables multiple user-defined imaging positions, as well as user-defined imaging time intervals.

In addition to polarized bright field imaging, the instrument has B/G/R fluorescent imaging capabilities using a mercury short-arc lamp excitation source. For multiple position imaging, the microscope is equipped with long-working distance 10x, 20x and 40x objectives.

A 63x objective is also available but due to the requirement of immersion-oil, this magnification is not suitable for multiple positions if the positions require relatively large stage movement or for long time periods of imaging. See details below.

The Live Cell Imager is available by appointment only. To make an appointment please contact Hans Minderman at 716-845-1162.

Technical specifications
  • Laser Scanning Spectral Confocal Microscope
  • Real-time xy scanning, z-stack and λ scan
  • Simultaneous multi-spectral scanning
  • 3 Acousto-Optical spectral detectors
  • 10X dry, 20X dry, and 63X oil immersion objectives
Illumination and detection
  • Four laser lines
  • 405nm and 638nm diode; 488nm and 552nm OPSL
  • 1 PMT, and 2 hybrid (HyD) spectral detectors
  • Transmitted Light PMT Detector (TLD)
Advantages of Confocal Microscopy on the Leica TCS SP8
  • Simultaneous, tunable multi-channel detection
  • Visible, far-red and infrared emission detection
  • Minimize laser exposure with ultra-sensitive HyD detectors
  • Superior background suppression and cross-talk correction
  • HyVolution high-definition 3D reconstruction
  • Fluorescence Resonance Energy Transfer (FRET) capability
Technical specifications
  • Time lapse, live cell imagery at 37°C for multidimensional 4D records (X,Y, Z time)
  • All components, camera, shutter and monochromator are fully integrated and optimized with regard to light efficiency and acquisition speed through a synchronized sequencing board.
  • Control of temperature, CO2 concentration and humidity within a climate chamber for optimum experiment conditions.
  • Computer, XP-Pro connected to Roswell Network
Excitation sources
  • Mercury Sort-Arc reflector lamp
  • Monochromatizing light source, Wavelength range 320nm-694nm
Filter sets available
  • RGB
  • YFP/ CFP FRET
  • DIC
  • Bright Field

Luminex services

The Luminex 200 is a flow-based multiplex analyzer that can perform up to 100 different tests in a single reaction volume, typically using magnetic beads to capture soluble analytes from plasma, serum and cell lysates.

Interested investigators should contact our principal operator Nitika Sharma at 716-845-5782 or Hans Minderman at 716-845-1162 for consultation on the Luminex multiplex assay, to reserve instrument usage time, or to request complete assay services.

Luminex 200 users may either perform the assay themselves using our Luminex facility (located in CCC 324), or simply request the kit(s) to be purchased and provide the experimental samples for complete assay service by our staff. If complete service is requested, the desired Luminex assay kit(s) will be ordered and processed with the provided experimental samples. The data will then be analyzed by our staff using

Upstate BeadView software and will be provided in a report detailing the analyte concentrations for each sample and the reliable limits of detection for each analyte.

Assay service fees are dependent on the kit(s) used, the number of samples, and the number of analytes, and will be determined before service is rendered by consultation with the principal operator.

Illumination
  • 532 nm, 10-15mW
  • 635 nm, 9mW
Fluorescence wavelengths
  • Reporter 1 (RP1): 575nm, Bandwidth 20nm
Optical design elements
  • Avalanche photodiodes in red and infrared
  • PMT for best sensitivity in RP1 wavelengths
  • Square quartz cuvette
  • Outside - 2mm x 2mm
  • Inside - 200μm x 200μm
  • DSP based waveform analysis for maximum sensitivity
Fluidics
  • 20μl to 200μl sample uptake volume via sample loop
  • Variable sample injection rate on syringe pump: 60 μl/min, 45 μl/min and 30 μl/min
  • Sample core: 15-20 μm core @ 60 μL/min default
  • Sheath flow rate: 90 μl/sec
  • Totally closed system for biohazard containment
  • On-board sheath fluid pressure pump
  • External sheath/waste containers

Education

The Flow and Image Cytometry Shared Resource (FICSR) teaches various free courses to educate the Roswell Park and Buffalo community on the techniques employed in our facility. Classes typically take about three hours and have a lecture presentation as well as a lab demonstration. For the dates and location of our classes, see the listing of Upcoming Courses below.

For beginners in Flow Cytometry and those new to our facility, we request that you take the Introduction to Flow Cytometry course before using the instruments outside regular business hours when no operator assistance is available. One-on-one training on the equipment is freely available to everyone upon prearrangement with FICSR staff.

It is highly recommended that after taking the Introductory Course you schedule a one-on-one session with a member of our staff when you are ready to acquire and analyze your data.

Offered on a monthly basis, this course is mandatory for those who want to use the flow cytometers on their own. The course covers facility logistics, flow cytometry basics, data analysis basics, and the use of proper controls. A simple instrument set-up with compensation is demonstrated on a FACSDiVa-based instrument (LSRII or Fortessa).

Next Class

Date: TBD
Microsoft Teams meeting

Sign Up

For those who have learned flow cytometry basics, this course gives intermediate-level users the opportunity to learn the BD DiVa software application more in-depth. The focus is primarily on experiment set-up and acquisition with compensation.

Next Class

Date: TBD
Two attendees limit
Date/time by arrangement
Please enter today’s date in the sign-up form

Sign Up

Designed for newcomers to the software, this class will teach all basic operations for data analysis and presentation through hands-on instruction.

Next Class

Date: TBD
Microsoft Teams online meeting

Sign Up

The introduction to ImageStream cytometry course is a three-hour course that is divided into two parts. During the first part, the basics on how the ImageStream works and its similarity and differences with conventional flow cytometry will be presented. Various examples of the most common ImageStream applications will be shown and tips and tricks and potential pitfalls for experimental setup will be discussed.

The second part is a hands-on/interactive instruction (in the RSC computer classroom) on using the IDEAS analysis software. In this portion, the basic features of the IDEAS software are explained by analyzing actual data sets. As a follow-up to this introductory course, one-on-one sessions should be scheduled with our operators for instruction on instrument setup and data acquisition.

Next Class

Date: TBD
Microsoft Teams online meeting

Sign Up

This course outlines various methods for DNA ploidy and cell cycle analysis and their applications. The practical part focuses on cell preparation for DNA staining with propidium iodide, followed by flow cytometry acquisition on the FACS Calibur and ModFit analysis.

Next Class

Date: TBD – after return to normal operations
Grossberg Classroom CCC314

Sign Up

This course introduces the basic concepts of fluorescent microscopy, as well as the understanding of confocal microscopy and its applications. The specific focus is on how to use our Leica SP2 confocal microscope.

Next Class

Date: TBD
Microsoft Teams meeting

Sign Up

This advanced course details the features of fluorescent probes typically used for flow cytometry.

The recommended participants include anyone embarking on multicolor flow cytometry experiments.

This course will teach the user how to select the right combination of these fluorochromes for your studies and how to perform proper compensation. The course concludes with a demo of compensation adjustments on a DiVa-based instrument (LSRII or Fortessa) and on several analysis programs.

Next Class

Date: TBD
Microsoft Teams online meeting

Sign Up

Services & fees

Frequently asked questions

Image
Roswell Park Comprehensive Cancer Center

Location & hours

Roswell Park Comprehensive Cancer Center
Department of Flow and Image Cytometry
Elm and Carlton Streets
Buffalo, New York 14263

Clinical lab phone: 716-845-3528

Research lab phone: 716-845-3470

Fax: 716-845-8806

Meet our team

Orla Maguire, PhD
Supervising Flow Cytometrist
Phone: 716-845-5890
Email: Orla.Maguire@RoswellPark.org

Please acknowledge the Flow and Image Cytometry Shared Resource in your publication as follows: "Cytometry services were provided by the Flow and Image Cytometry Shared Resource at the Roswell Park Comprehensive Cancer Center which is supported in part by the NCI Cancer Center Support Grant 5P30 CA016056 and NCI R50 R50CA211108."

If you are using the ImageStream imaging flow cytometer in your publications, please acknowledge the Shared Instrument grant that funded the acquisition of this instrument as follows: "The ImagestreamX-MKII instrument was funded through NIH Shared Instrument Grant 1S10OD018048."

If you are using the SONY MA900 in your publications, please acknowledge the Shared Instrument grant that funded the acquisition of this instrument as follows: "The SONY MA900 sorter was funded through NIH Shared Instrument Grant 1S10OD025183."

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Publications