A National Cancer Institute-designated Comprehensive Cancer Center

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Analytical Cytometry

Analytical Cytometry Core (ACC)
City of Hope’s Analytical Cytometry Core (ACC) Facility provides researchers with high-quality flow cytometry instrumentation, as well as expertise in analyzing and/or sorting sample populations of interest via interpretation of their physical, fluorescent and/or light-scattering properties. The facilities and their services are available to both City of Hope and non-City of Hope researchers.
 
The ACC Facility has two components: the Flow Cytometry Facility (FCF) and the Molecular Imaging Facility (MIF).
 
As an added convenience, the ACC supports several off-line workstations throughout the campus where investigators can analyze their ACC-derived data. FCF software includes Flowjo (only on some workstations), Modfit (only on some workstations)and Summit. MIF software includes ImageQuant, PDQuest (Bio-Rad) or Odyssey.
 
Flow Cytometry Facility
The Flow Cytometry Facility has three Cell sorters (MoFlo, Aria III, and Aria SORP) and 5 analyzers (CyAn, Gallios, Fortessa, C6, and Guava). All the analyzers are available for unassisted end user runs.
 
Molecular Imaging Facility
This facility contains instruments utilizing lasers to scan gels, membranes (e.g., nylon, nitrocellulose), microtiter plates or thin-layer chromatography plates. The molecules of interest (DNA, RNA or proteins) are labeled with radioactive or fluorescent tags. There are two types of instruments available:
 
  • The Typhoon 9410s (General Electric) has four excitation sources (457 nm, 488 nm, 532 nm and 632.8 nm). The Typhoon can also be used to acquire radioactive labeled data with a special phosphor screen, which is used to capture quantitative information that is then scanned on the Typhoon. This facility contains two Typhoon scanners.
  • Odyssey Infrared Imager (Li-Cor) has two excitation sources (680 nm and 780 nm).
 
Research reported in this publication included work performed in the Analytical Cytometry Core supported by the National Cancer Institute of the National Institutes of Health under award number P30CA33572. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Services

Flow Cytometry Facility
The Flow Cytometry facility uses flow cytometric principles to analyze and/or sort samples (cells, bacteria, chromosomes, or particles) based on fluorescence or physical properties.
 
  • Sorting: MoFlo™, Aria III, and Aria SORP
  • Analysis: Operator assisted and unassisted: CyAn™, Gallios, Fortessa, C6 and Guava
 
Molecular Imaging Facility
This facility provides instrumentation for scanning gels, microtiterplates, and membranes for a variety of radioactive, fluorescence, or chemiluminescence molecules.
 
  • Typhoon™ 9410
  • Odyssey®
 

Typhoon™ 9410

The Typhoon™ 9410 equipment is available to scan phosphor screens, which have been exposed to radioactive sources on gels/blots/dishes/trays/glass slides.  The Analytical Cytometry Core has a large format (35x40 cm) phosphor screen and several small (20x25 cm) format screens available for proof of methodology.  Screen erasers are located at the core workstations or nearby.  This allows linear quantification of data (over five orders of magnitude) and produces digital images, which are directly manipulatable for analysis and publication.  Exposure times are usually 20-25% of those required on film.  Radioisotope sources have included P32, S35, C14, and I125. There are also special screens available from Amersham Biosciences for tritium detection.
 
The Typhoon™ 9410 also has the capability to detect fluorescent labels on gels, blots, dishes, trays, and glass slides. This instrument has four excitation sources: 457nm, 488nm, 532nm, and 633nm that allows for use of a wide range of fluorescent dyes.  There are also selectable emission filters (520BP40, 555BP20, 580BP30, 610BP30, 670BP30, 526SP, and 560LP), which allow for collection of specific and multiple signals.  This methodology allows for linear quantification of the data (over three to four decades of magnitude) and produces digital images, which are directly manipulatable for analysis and publication.  The matrix of interest can be scanned at various resolutions depending on experimental design.  For radioactivity, the range in resolution is from 25u to 1000u and for fluorescence, the range is from 10u-1000u.  At 10u the scanner is capable of handling microarray slides.
 
Quality Control
Users are generally responsible for running their own controls.
 
Supplies
Cleaning solution and Kimwipes® are provided for cleaning the platen on the scanner.  All other reagents, solutions, buffers, and supplies must be provided by the user.  Several phosphor screens are provided by the core for radioisotope scans, but users are encouraged to buy their own screens.

Analytical Cytometry Equipment

We offer the following equipment in the Analytical Cytometry Core.
 
Flow Cytometry Facility (FCF):
  • MoFlo™ MLS
  • Aria III
  • Aria SORP
  • CyAn™ ADP 9 Color
  • Gallios
  • Fortessa
  • C6
  • Guava
 
Molecular Imaging Facility (MIF):
 
  • Typhoon™ 9410
  • Odyssey® Infrared Imager
 

Odyssey® Infrared Imager

The Odyssey® Infrared Imager scanner can be used to analyze two-color western blots, two-color in-gel westerns, two-color northern blots, and any other primary antibody tagged samples, which can be stained with near-infrared secondary antibodies (IRDye 700 and IRDye 800) or Alexa Fluor 680.  It can also visualize Coomasie stained protein gels, which are fluorescent in the infrared.  Along with scanning gels and membranes, it can also scan microplate cell-based assays.  Various resolutions can be used for scanning, which range from 21u to 337u.
 
Quality Control
Users are generally responsible for running their own controls.
 
Supplies
Cleaning solution and Kimwipes® are provided for cleaning the platen on the scanner.  All other reagents, solutions, buffers, and supplies must be provided by the user.

MoFlo MLS

The MoFlo™ MLS has a four-laser optical bench configuration. The four lasers (one HeNe, two argon gas, and one krypton gas) provide fluorescent excitations from 351-752.5 nm. The instrument is built around dedicated electronics with control of acquisition, sorting and analysis directly from a computer with dual CPU and dual monitors. This instrument has 10 separate photomultiplier tubes (PMTs) plus a forward scatter detector. Peak height, integrated and logarithmic data can be collected from each of the PMTs. A digital signal processor (DSP) computed-parameter board allows for up to 32 parameters (including pulse width on the trigger event and time) to be stored in listmode for a sample. DSP also allows for 8x8 inter-laser compensation of fluorescence parameters for sorting. The facility normally sorts with a 100 micron nozzle at 30 PSI. Up to four sort decisions can be performed simultaneously. Along with bulk sorting, cell deposition into wells as single cells or multiple cells is also a possibility. The instrument can accommodate plates that contain from 6 to 384 wells.
 
MoFlo: 4-way sorting streams MoFlo: Cytomation sorting unit
MoFlo MLS (Multi-Laser System)  
 
Quality Control
Bead fluorescent standards are used to align the solid state HeNe, two argon gas lasers, and one krypton gas laser. Once a month, mean equivalent fluorescence standards (FITC, PE) are used to check the linearity and sensitivity of the instrument.
 
Supplies Needed
Investigators must supply their own reagents. We have an extensive library of vendors to facilitate the search and purchase of reagents. We do have a variety of fluorochrome labeled secondary anti-mouse IgG antibodies, which we can supply on a trial basis if the investigator is interested in trying multi-color staining. We provide the tubes necessary to sort user samples on the MoFlo™. The investigator should supply the facility with the plates or tubes with appropriate media or collection solution to collect the sorted cells.
 

CyAn

The CyAn™ ADP 9 Color has three excitation sources: 405nm, 488nm & 635nm. This instrument has 11 parameter capabilities (two scatter and nine fluorescent detectors).All parameters can be collected in linear peak height, linear area, and/or log scales.It is capable of acquisition rates up to 50,000 events per second.This cytometer is available for trained users to operate, or user samples may be analyzed by the core at an additional cost.
 
CyAn™ (Flow Cell) CyAn™ ADP
(Advanced Digital Processing)
Quality Control
Standard fluorescent beads are run once a week to ensure proper laser alignment for stability and reproducibility.
 
Supplies
Investigators must supply their own reagents. We have an extensive library of vendors to facilitate the search and purchase of reagents. We do have a variety of fluorochrome labeled secondary anti-mouse IgG antibodies, which we can supply on a trial basis if the investigator would be interested in trying multi-color staining. Users that are trained to run their own samples on the analyzers (CyAn™ or FACScalibur) will need to provide their own tubes.
 

Using the Facility

Scheduling Appointments
 
Flow Cytometry Facility
Lucy Brown, Claudio Spalla, or Alex Spalla, 626-256-HOPE (4673) ext. 63306, 65832, or 64727

To schedule appointments it is best email Lucy Brown (lbrown@coh.org).
 
Molecular Imaging Facility
Claudio Spalla
626-256-HOPE (4673), ext. 63306

Turn-around Time
 
Flow Cytometry Facility
Simple analysis schemes are accomplished as samples are run.If more detailed analysis is needed, it is usually done within a 24-hour period.
 
Molecular Imaging Facility
Scanning time for the Typhoon™ 9410 and the Odyssey® is dependent on format size and desired resolution.The range is fromfive to 45 minutes.
 

Abstract for Grants

The Analytical Cytometry Core (ACC) is comprised of two facilities: the Flow Cytometry Facility and the Molecular Imaging Facility. Both facilities are available for every investigator of the Beckman Research Institute of City of Hope.
 
The Flow Cytometry facility contains 3 cell sorters and 4 analytical cytometers.
 
The cell sorters include:
  • MoFlo legacy (Beckman Coulter) with 3lasers (405nm, 488nm & 640nm) and up to 9 parameters
  • Aria III (Becton Dickinson) with 4 lasers (405nm, 488nm, 561nm & 633nm) and up to 15 fluorescent parameters
  • Aria II SORP (Becton Dickinson) with 6 lasers (355nm, 405nm, 457nm, 488nm, 561nm & 640nm) and up to 18 simultaneous fluorescent parameters. This sorter is contained in a biosafety cabinet

 

The Analytical cytometers include:
  • Gallios (Beckman Coulter) with 10 fluorescent detectors and 3 lasers (488nm-5detectors, 405nm-2detectors, & 638nm-3detectors). It also has a 32 tube carousel for walk away data acquisition.
  • CyAn ADP (Beckman Coulter) with 9 fluorescent detectors and 3 lasers (488nm-5detectors, 405nm-2detectors, & 640nm-2detectors). We also have the ability to attach a Hypercyte for high throughput screening from 96/384 well platform.
  • Fortessa SORP (Becton Dickinson) with 5 lasers (355nm, 405nm, 488nm, 561nm, 640nm) and up to 17 simultaneous fluorescent parameters. HTS module for 96 well sample delivery allowing for automation is also available.
  • C6 (Becton Dickinson) with 4 fluorescent detectors and 2 lasers (488nm-3 detectors, 640nm-1 detector).

 

ACC also provides training for user operation of the analytical instruments, basic flow theory, software, and data analysis for result generation and presentation. Other functions of ACC include application assistance, experimental design, and site license management for Flowjo and FCSexpress.
 
The Molecular Imaging Facility is utilized for quantitation of radioactivity or fluorescence, chemiluminence, or chemifluorescence of gels, blots, membranes, glass slides & microtiter plates. The equipment includes 2-GE Typhoon 9410 scanners and a LiCor Odyssey Near InfraRed Scanner.
 

Pricing

Prices and availability vary. Please contact us for current information.
 
If you are a City of Hope employee, please visit this core's intranet site for pricing.
 

Contact Us

Email us for pricing or more information about the Analytical Cytometry Core or to schedule an appointment for services or equipment.
 
Address:
Beckman Research Institute of City of Hope
1500 East Duarte Road
Duarte, CA  91010-3000
Phone:  626-256-HOPE (4673), Ext. 63306
Fax:  626-358-7703
 
Lippman-Graff Building:
Room 101 - Office
Room 101B - MoFlo
Room 102 - Tissue Culture for Flow Cytometry Facility
Room 103 - Supply Room and Preparation Room for MIF
Room 114 - Molecular Imaging Facility Scanners

Gonda Building:
Room 1006B - BD Fortessa Cytometer
Familian Science Building:
Room 1215 – CyAn, Gallios, AriaIII, and Aria SORP cytometers
RoboSep magnetic bead separator

Beckman Research Center:
Room 3300– C6

Workstation Locations:
Lippman-Graff Building, Room 101, 118, 221
Fox North Building, Room 1001
Fox North Building, Room 2002C
Kaplan Black Building, Room 106D
Kaplan Medical Building, 2nd Floor
Hilton Building, Room 127
Flower Building, Room 150
Gonda Building, Room 2006
Shapiro Building, Room 1033
Familian Research Building, Room 1215
 
Research Shared Services

City of Hope embodies the spirit of scientific collaboration by sharing services and core facilities with colleagues here and around the world.
 

Recognized nationwide for its innovative biomedical research, City of Hope's Beckman Research Institute is home to some of the most tenacious and creative minds in science.
City of Hope is one of only 41 Comprehensive Cancer Centers in the country, the highest designation awarded by the National Cancer Institute to institutions that lead the way in cancer research, treatment, prevention and professional education.
Learn more about City of Hope's institutional distinctions, breakthrough innovations and collaborations.
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By giving to City of Hope, you support breakthrough discoveries in laboratory research that translate into lifesaving treatments for patients with cancer and other serious diseases.
 
 
 
 
Media Inquiries/Social Media

For media inquiries contact:

Dominique Grignetti
800-888-5323
dgrignetti@coh.org

 

For sponsorships inquiries please contact:

Stefanie Sprester
213-241-7160
ssprester@coh.org

Christine Nassr
213-241-7112
cnassr@coh.org

 
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