FACS Facility 

 

Division of Biological Sciences

 

Indian Institute of Science 

        About

FACS (Fluorescence Activated Cell Sorter) facility was established in 1997 with   support from DBT, Govt. of India. There are six flow cytometers for advanced scientific research. BD FACScan, BD FACS Calibur TM, BD FACS CantoTM II, BD FACSVerse, Beckman Coulter MoFlo XDP High Speed Cell Sorter & Analyzer, and BD FACS AriaTMII.  BD FACScan, BD FACS CaliburTM and BD FACS CantoTM II are analyzers whereas MoFlo XDP and BD FACS AriaTMII are cell sorters with advanced features. All the flow cytometers are operator assisted. Moreover, FACS facility has offline analysis systems for data analysis by flow cytometer users.

    People  
    FACS committee and members  

       1.  Prof.  Ganesh Nagaraju Convener, FACS Facility

            Phone: +91-80-22933055, Email: nganesh@biochem.iisc.ernet.in

 

      2.  Prof. Patrick D' Silva, Department of Biochemistry

      3.  Prof.  Annapoorni Rangarajan, Department of Molecular Reproduction, Development and Genetics

      4.  Dr. Amit Singh, Department of Microbiology and Cell Biology 

     

    Faculty  
        Dr. William R. Surin, Senior Scientific Officer  

        Department of Microbiology and Cell Biology 

        Phone: +91-80-22933451/2883, Email: wrsurin@mcbl.iisc.ernet.in

    Scientific Staff  

        Dr. Uttara Chakraborty, 

        DBT-GIA Senior Research Scientist(DBT-IISc Partnership Program)  

        Phone: +91-80-22933451, Email: uttara@biochem.iisc.ernet.in

    Staff  

        Ms. Urvashi Chauhan, Project Assistant

        Ms. Leepika Baid, Project Assistant

        Ms. Ranjitha R, Project Assistant

        Ms. Sampratha Bhagyashekar, Project Assistant

 

           Flow Cytometers: Cell Sorters

MoFLo XDP Cell Sorter and Analyser: It is having three lasers (488nm, 355nm and 640) with 10-color parameters. Windows XP operating system with SummitTM Software (5.1) for acquisition, analysis and sorting.

Laser Specifications: 488nm iCyte 30-200mW air cooled solid state laser (5-color), 355nm JDSU 100mW UV solid state laser (2 color) and 640nm Coherent 100mW solid state laser (3 color)

Five wavelengths detected from the 488nm laser: (FL1).  510-550nm (FL2). 565-595 nm  (FL3).  603-623nm (FL4).  655-685nm and (FL5).  740LP

Two wavelengths detected from the 355nm laser: (FL6). 418-483 nm and  (FL7). 615-645 nm 

Three wavelengths detected from the 640nm laser: (FL8). 645-685nm (FL9). 713-727nm and (FL10). 755-815nm

Sort Performance

  • It is validated for 70,000 sorts per second with >99% purity at all data rates

  • 4 - Way Sort for multiple population sorting

  • Independent Sort Mode capability in each tube

  • Cyclone custom plate and slide sorting

Touch Screen Control Panel

  • Easy-to-use intuitive cytometer setup and control

  • Independent control of each system

  • Simple coarse and fine alignment

  • Cyclone calibration upto 1536-well plate

  • Real-time sort statistics

Summit Software

  • Established software with powerful data handling

  • 18x18 Auto-compensation matrix

  • Determine regions with full parameter resolutions

  • > 1 Billion event listmode files

  • Workspace concept for flexible experiment design

Features : 

This is a high speed sorter compatible with array of laser options. High viability, yield, and purity 

Biosafety

Aerosol Evacuation System (AES) removes aerosols from sort chamber. Replaceable probe and sample line. Smart Sampler sample input protects the sample and user.

Electronics

  • Zero dead time

  • >100,000 analysis events per second

  • Digital triggering on any parameter

  • Digital pulse processing

  • True dynamic range of 5 decades

  • Unmatched linearity

BD FACSAriaTMII: It is having three lasers with 9-color parameters. Windows XP operating system with FACS DivaTM Software for acquisition, analysis and sorting.

Laser Specifications: 488-nm Coherent SapphireTM solid state (13mW-20mW), 633-nm JDS UniphaseTM HeNe air-cooled(10mW-20mW), 375-nm near UV, air launched laser (7-10 mW). All lasers are Class IIIb.

Fluorescence Detectors: 9 PMTs in 5+2+2 configuration

Five wavelengths detected from the 488-nm laser (standard filters): (1).  515-545 nm FITC, (2). 564-606 nm or 563-589 nm PE,  (3). 600-620 nm PE-Texas Red (4).  675-715 nm or 665-685 nm PE-Cy 5 or PerCP or PI, (5).  750-810 nm PE-Cy7

Two wavelengths detected from the 633-nm laser (standard filters): (1). 650-670 nm APC (2). 750-810 nm APC-Cy7

Two wavelengths detected from the 375-nm laser (standard filters): (1). 430-470 nm Hoechst blue emission (2). 675 nm Long pass Hoechst red emission

Features: 

Sheath pressure is adjustable from 5 to 75 psi. Filters and mirrors are user changeable. Maximum acquisition rate (events per second) with 12 compensation pairs and 8 parameters: 70,000 events/sec.

Purity and Yield

At 70 psi and 90 kHz with an average threshold rate of 25,000 events per second, a four-way sort achieved a purity of >98% and a yield >80% of Poissonís expected yield. Higher threshold rates of up to 70,000 events per second can be achieved without affecting purity; however, yield will decrease based on Poisson statistics.

Viability

All sorts resulted in cells that proved viable and proliferated for several days post-sort.

Sort Collection Devices: Two-way sorting: microtubes, 12 x 75 mm, and 15 mL, Four-way sorting: microtubes and 12 x 75 mm, Automatic Cell Deposition Unit (ACDU) for slide and plate sorting: 6, 24, 48, 96, and 384-well plates

Sample Injection Chamber; Sample Input Sizes: Microtubes, 12 x 75 mm, and 15 mL Polystyrene or polypropylene tubes can be used.

Sample Flow Rates: Adjustable sample pressures from 1 to 11

Sample Input Agitation: Adjustable through the software to keep sample constantly suspended

Temperature Control :Sample input, software-adjustable: 4, 20, 37, and 42OC, Sample output for sort collection: water refrigerator or heater option is available to provide cooling or heating for sort collection into tube holders, multi-well plates, and slides

BD FACS Accudrop: Red diode laser provided for manual or fully automated drop-delay determination. Automated drop break-off monitoring. Automated clog detection system using Sweet Spot technology

             Flow Cytometers: Analyzers

BD FACSCantoTMII: It is configured with 3 lasers and 8 fluorescence parameters

Windows XP operating system with FACS DivaTM Software for acquisition & analysis

Laser Specifications: Air-cooled, 20 mW solid state 488 nm, 17 mW HeNe 633 nm laser and 10mW-17mW Point Source Violet solid state laser 405nm 

Optics configurations: 

FSC Detector Photodiode with 488/10BP, 

SSC Detector PMT with 488/10BP

Fluorescence Detectors: 8 PMTs in 4+2+2 configuration

Four wavelengths detected from the 488-nm laser (standard filters): (1).  515-545 nm FITC  (2). 564-606 nm PE  (3).  670LP nm PE-Cy5 or PerCP (4).  750-810 nm PE-Cy7

Two wavelengths detected from the 633-nm laser (standard filters): (1). 650-670 nm APC (2). 750-810 nm APC-Cy7

Two wavelengths detected from the 405-nm laser (standard filters): (1). 485-535 nm AmCyan (2).  425-475 nm Pacific Blue 

Sample Injection: Direct into flow cell

Max Particle Size: 50 μm

Flow Rate, min: 10 μL/min

Flow Rate, max: 120 μL/min

Sample Acquisition Rate: 10,000 events/sec,

10 channels with 8 compensated fluorescent parameters

Nominal Minimum Sample Volume: 30 μL

 

BD FACSVerseTM : It is configured with 3 lasers and 8 fluorescence parameters Windows 7  operating system with  BD FACSuite Software for acquisition & analysis:

 Laser Specifications:

  •  20 mW  488 nm  Solid state Blue laser with Beam spot size: 9 μm x 63 μm

  •  40 mW 640 nm Solid stateRed laser with Beam spot size: 9 μm x 63 μm

  •  40 mW 405 nm  Solid stateViolet laser with Beam spot size: 9 μm x 63 μm

Fluorescence Detectors: 8 PMTs in 4+2+2 configuration

Four wavelengths detected from the 488-nm laser (standard filters): (1).  511-543 nm FITC  (2). 565-607 nm PE  (3).  673-727 nm PE-Cy5 or PerCP (4).  755-811 nm PE-Cy7

Two wavelengths detected from the 633-nm laser (standard filters): (1). 650-670 nm APC (2). 755-811 nm APC-Cy7

Two wavelengths detected from the 405-nm laser (standard filters): (1). 506-550 nm AmCyan (2).  425-470 nm Pacific Blue

BD FACSCaliburTM:It is dual laser with 4-color parameters. Mac  operating system  with   CellQuestPro Software for acquisition & analysis.

Laser Specifications:15mW 488nm, air-cooled argon-ion laser; Red diode laser, 635nm

Fluorescence Detectors and Filters: High performance, high dynamic range photo- multipliers with bandpass filters : 530nm (FITC), 585nm (PE/PI), 661nm (APC) and >650nm (PerCP) with base unit, >670nm (PerCP) with FL4 option

Sample Flow rate: Three selectable flow rates of 60mL/min, 35mL/min and 12mL/min. Particle velocity in flow cell is approximately 6 meters/sec

Sample Concentration: Single-cell suspension of 105 to 2 x 107 particles/mL recommended range. Kindly use 12 x 75-mm polystyrene tubes (Falcon) (Part No. 54-10001-00) for samples for acquisition.

BD FACScan:

Single Laser with 3-color parameters. Mac operating system with CellQuestPro Software for acquisition & analysis.

Laser Specifications:15mW 488nm, air-cooled argon-ion laser

Parameters: Forward Scatter, Side Scatter, FL1, FL2 and FL3

 

 

Contact Us: @

FACS Facility

Ground Floor, GC-03 (GCCL-01), Biological Sciences Building,

Indian Institute of Science

Bangalore-560012, Tel: +91-80-22933451  

Some useful links:

 

The Cytometry Society, India

International Society for Advancement of Cytometry

by wrsurin@mcbl.iisc.ernet.in,    Last updated on 13th December, 2016