Core Flow Cytometry Facility

The Saint Louis University, School of Medicine Core Flow Cytometry Facility provides comprehensive flow cytometry and cell sorting services. As part of the Department of Molecular Microbiology & Immunology, the Flow Cytometry Core Facility serves investigators of the Saint Louis University by providing access to routine flow cytometry assays such as immunophenotyping and DNA analysis. However, research users encompass individuals with a broad spectrum of training in the basic and clinical sciences, and thus, a major goal of the facility is to provide the guidance, technical assistance, and equipment for investigators to utilize more complex multi-parametric, multi-laser measurements as well as cell sorting in their research regardless of their level of cytometry expertise. Services provided by the facility personnel extend from consultation on experimental design, sample preparation and data analysis to instrument operation and set-up for cell-sorting and multi-laser operation. In addition, the Core Facility has an active program of assay development guided by the interests of the investigators. As such, the Flow Cytometry Core Facility serves as a focus for individuals interested in cellular based measurements and cellular heterogeneity in disease providing critical support for cancer related research within the institution. The equipment and expertise housed within the Core Facility permits implementation of most flow cytometry assays that have been published. The facility includes a bench top single two laser instrument (Beckton Dickinson, FACScaliber), with a 96 and 384 well plate adapter as well as off-line data analysis hardware and software.

Joy Eslick serves as Manager of the Core Facility. Joy Eslick comes to SLU with 5 1/2 years of full-time flow cytometry experience and served as senior technologist for 2 of those years. Her experience includes development of 2, 3, and 4-color acquisition and analysis protocols using CellQuest software, antibody QC and validation, instrument maintenance and troubleshooting, analysis of diagnostic leukemia/lymphoma samples including peripheral blood, bone marrow, and tissue, peripheral blood lymphocyte subset profiles for monitoring of immune system function, DNA ploidy analysis of leukemic cells using ModFit software, peripheral blood progenitor cells monitoring for collection timing, and analysis of peripheral blood, bone marrow, cord blood, and transduced cellular clinical transplant products in a cGMP facility. Ms. Eslick is an active participant in the national and international flow cytometry society meetings as well as assisting in educational activities at the national course level.

Performance of flow cytometric assays:
The assays offered by the facility range from routine cell cycle analyses and immunophenotyping to use of the multi-laser capabilities and cell sorting. Most applications are combined in multiparametric assays. Examples of the assays available include, but are not limited to the following:

• DNA content/cell cycle measurement, including the simultaneous measurement of multiple fluorescent probes if required.
• Cytokine bead arrays for mouse and human samples
• Immunofluorescence analyses (three and four color analyses are routinely performed) including characterization of plasma membrane antigens/receptors, cytoplasmic proteins, nuclear antigens, and intracellular cytokines.
• Characterization of cell populations based on scattered light intensity measurements, and autofluorescence.
• Cell sorting including viable, sterile cell sorting and spot blot analyses. This includes isolation of monocytes/macrophages, dendritic cells, lymphocytes form single cells suspensions
• Intracellular calcium flux
• A range of apoptosis assays including Annexin V/PI, TUNEL, mitochondrial membrane potential, and measurement of proteins involved in the apoptosis regulation such as BCL-2 family proteins, CD95 and caspase protein levels and activity
• Receptor-ligand interactions
• Cell proliferation studies including BrdU incorporation
• Viability assays (membrane exclusion and metabolic viability)
• Various function assays including oxidative metabolism, neutrophil function (oxidative burst, phagocytosis), cytoplasmic pH, membrane potential.
• Kinetic analyses

Full service consultation and training
An hourly rate of $80.00 will be assessed to all investigators. Investigators seeking to utilize flow cytometry in their research may request from the Core Facility staff consultation regarding method choice or development, sample preparation and data analysis. Training and follow up consulting is provided by the Core Facility staff. Investigators are not charged for these services which in many instances are extensive. Joy may be reached at 314-577-8420 or eslickjb@slu.edu.

Examples of data obtained by flow cytometry:

Unfixed human mononuclear cells directly stained for Fas and FasL ligand expression.

Human fibroblasts examined for mitochondrial mediated apoptosis. The top panel shows unfixed cells stained with Rh123 to measure mitochondrial function and PI to measure membrane permeability. The bottom panel shows cells fixed in 70% ethanol and stained with PI to determine DNA content.