Flow Cytometry Facility

The Wistar Institute Flow Cytometry Shared Resource provides flow cytometric services, training, advice, and support for the use of flow cytometric techniques by Wistar Cancer Center investigators, as well as the greater basic research community. 

Services and capabilities

Flow cytometric analysis

Instrument

Investigator Operated or Operator Assisted

Operator-Assisted Only

BD LSRII - 14 or 18 color

X

 

DakoCytomation CYAN-ADP - 9 color

X

 

BD FACSCalibur - 4 color

X

 

Beckman-Coulter XL - 4 color

X

Immunophenotyping/immunofluorescence

Immunophenotyping/immunofluorescence utilizing single or multiple laser analysis of up to 18 fluorochromes combined with forward- and right angle-light scatter. This includes various immunofluorescence analyses such as cell surface and intracellular phenotyping and making use of QuantumDot (QDot) technology.

Fluorescent protein expression

Fluorescent protein expression with GFP, YFP, CFP, and various red fluorescent proteins including monomeric red proteins such as Cherry.

Cell-cycle analysis

Cell-cycle analysis with propidium iodide or 7-AAD, anti-BrDU antibodies, tracking of cell division using CFSE or similar dyes, apoptosis in cell lines and primary cell populations using various techniques, including propidium iodide (PI) staining, terminal desoxytransferase (TdT) staining, and Annexin V labeling.

Functional assays

Functional assays, such as intercellular conjugation run on cells stained with intracellular calcein, hydroxyethidine, or PKH26, as well as surface markers; oxidative stress; pH; calcium flux. Total cellular protein can be measured with FITC or SR101.

Cell sorting

Cell sorting (operator-assisted) of up to 4 separate populations using up to 16 fluorochromes on the high-speed BD ARIAII or 6 fluorochromes on the MoFlo instrument.  High purity, high recovery, sterile cell sorting (single or multiple laser) may be performed on any cell population detectable by analysis (above). Up to four populations may be collected simultaneously, at sorting rates of up to 25,000 cells/second (90 million/hour).

BSL2+ level flow cytometry analysis and sorting

BSL2+ level flow cytometry analysis and sorting of potentially infectious biological materials, utilizing the BD ARIAII.  This provides an in-house resource for running non-characterized human-derived material, infected cell lines, or other potentially bio-hazardous samples.

Support functions

Consulting

Consulting with investigators to determine individual flow cytometry needs; to aid in development of appropriate and cost-effective protocols and techniques.  Resource staff assists in design of large multicolor fluorochromes panels and setting of compensation matrices, and provide advice on standard as well as specialized services, e.g. whether to use common reagents for surface phenotyping or specialized reagents and new techniques. 

Training and providing ongoing advice

Training and providing ongoing advice to investigators in the use of investigator-operated instrumentation so they may perform data acquisition and analysis without hands-on operator assistance; providing technical support for analysis of flow cytometry/cell sorting data for publication, presentation, and inclusion in grant applications.  Staff is conveniently located in order to provide immediate assistance and advice to investigators on user-friendly instruments. 

Facilitating collaborative efforts

Facilitating collaborative efforts among Wistar investigators, and where possible, with other Institute shared resources and other cytometry resources.  Resource operators are familiar with technical aspects of many of the investigators’ experiments, allowing them to facilitate sharing of reagents and techniques and to encourage collaborations among Wistar users.  Mr. Faust, manager, is well acquainted with a vast external network of flow cytometry laboratories and researchers in the field, and able to provide additional opportunities for collaborative sharing and development of techniques.  Additionally, as part of the Shared Resources Committee, he interacts on a regular basis with Wistar Shared Resource directors and is well-informed of resources available that may be used in conjunction with flow cytometry to augment or enhance research projects.

Providing quality assurance and feedback

Providing quality assurance and feedback to users by way of direct and immediate interaction of resource staff with investigators in the event of problems, through regularly scheduled user group meetings, and by way of occasional surveys to assess performance of staff as well as instruments (such as sorting rates, purity, and sterility) to gather suggestions for further improvements or enhancements.

Storing, archiving, and retrieving

Storing, archiving, and retrieving flow cytometric data and disseminating it through a local network and communication with the Institute network servers; working together with the Institute Information Systems to provide 24 hour accessibility and data security.

Administering FlowJo site license

Administering FlowJo site license for Wistar investigators to allow for low-cost post-acquisition analysis software, along with providing troubleshooting as needed.

Maintaining shared resource website

Maintaining shared resource website, which provides up-to-date information about the resource and its services, the latest enhancements, and helpful flow cytometry links in a manner that is convenient, efficient, timely, flexible, and accurate.

Maintenance

Maintenance of flow cytometry instrumentation is performed by the Shared Resource staff, which results in fewer disruptions to ongoing assays, maximum up-time operation, and elimination of the need for costly service contracts.

 

Instrumentation

View an overview of the instruments... Learn More >

How to Obtain Services

Contact the facility in person or by phone to schedule services... Learn More >