Flow Cytometry Core

Services

Consultation

All investigators are encouraged to meet with the Flow Cytometry Core technologists to discuss the experiment. This meeting is often useful in that the technical limitations of an experiment are realized and many potential problems can be averted before critical resources are used. The Core can also provide written protocols for staining, fixation and data analysis methods. The Core technologists are available for assistance with data retrieval, analysis and presentation.
Materials available for reference in the Flow Core lab include several textbooks on the basics of flow cytometry and the Current Protocols in Cytometry. In addition, the Core frequently organizes training seminars at different levels of expertise.

FACS Applications

  • Immunophenotyping
  • Intracellular staining
  • Molecular and cellular probes (Ca flux, GFP, DNA cell cycle
    Fixing-StainingCellularDNA.pdf )
  • Analysis of cells undergoing cell death or apoptosis (TUNEL assay
    TunelStaining.pdf, Annexin V)
  • Detection of soluble proteins via bead capture (Cytometric Bead Array, Flowcytomix)
  • Sterile isolation of selected cell populations on the basis of specific protein expression or cell size
    (AriaIISortingProcedure.pdf )
  • Use of the Countess automated cell counter
    (Countess.pdf )

Data is acquired as FCS 3.1 using Histogram Software on the Apogee A50 Micro. Additionally, BD FACSDiva Software is used to acquire data on the LSR II, Fortessa and Aria II. Data can be imported/exported as FCS 2.0 or 3.0 when using FACSDiva. List mode acquisition and storage of data allows for analysis and presentation many different formats according to the needs of the investigator. Data files are permanently stored in personal folders on the LRI servers, readily available to all LRI employees with proper logon accounts. (ServerAccessForm.pdf)
CellQuest, FlowJo (FlowJo License Requests.pdf ) and ModFit software are provided for analysis. Investigators are encouraged to schedule a training session to learn analysis of flow data utilizing one of the available software packages.

Training

The LRI Flow Cytometry Core offers training programs for Beginning and Advanced Flow Cytometer Operators. These courses cover the hardware and software of our analytical instrumentation. It is the belief of the Flow Core that greater understanding of the instruments and physics employed in experiments will lead to dramatically improved data and experimental design. The Flow Core is dedicated to assisting investigators to attain whatever their desired level of expertise.

The Basic course introduces users to the hardware and software of the LSR II, Fortessa or Apogee. It is not meant to immediately provide complete self-sufficiency but rather to embrace the major concepts needed to begin. It is expected that after this training the user will plan and perform their next experiment with the assistance of Core personnel.
The Advanced course provides methods and strategies to optimize high-level multicolor flow cytometry. Material covered is derived from the top thought-leaders in the field of flow cytometry tempered with personal experience and practicality. Building multicolor assays is a process that requires time, dedication and tenacity to overcome hurdles that can appear when attempting to measure many parameters simultaneously. When the investigator can patiently build an experiment with a series of small steps and optimally characterize the instrument settings for their particular research, truly spectacular results.

What to Bring

Bring something to take notes on. Samples and supplies will vary greatly based on specific research. Core personnel will help define what is needed on a person-to-person basis.

Please schedule appointments for analysis and sorting at 216-445-3798. The Flow Cytometry Core is located in NBb-40. The FACSAria II is located in NE3-224.