Overview
Flow Cytometry provides a flexible method to characterize cells in suspension. Fluorescence activated cell sorting allows the user to select living cells on the basis of characteristics measured by flow cytometry. Among the many cellular features and functions that can be measured are expression of proteins outside and within cells, analysis of DNA content, viability and apoptosis, multiple drug resistance pump activity, enzyme activity, T-cell activation, T-cell receptor specificity, cytokine expression, phagocytosis and oxidative burst activity are examples.
One of the major missions of the Flow Cytometry Facility is to train investigators, their students and staff to use analytical flow cytometry to its fullest advantage in their research. Basic and advanced training in the use of the Coulter XL four-color flow cytometer is offered continuously. This user operated cytometer is available to trained users 24 hours a day, seven days a week. Rigorous quality control is performed daily on this instrument and users are provided with validated settings for the common fluorochromes used in multi-parameter cytometry. Consultation concerning experimental design, analysis and assay development is available to UPCI investigators without charge.
The Flow Cytometry Facility is also dedicated to providing high quality cell separation by fluorescence activated cell sorting. The flagship sorter is a Cytomation MoFlo high speed sorter capable of sorting speeds in excess of 30,000 events per second. This instrument was purchased through a National Institutes of Health shared instrumentation grant. It is equipped with two lasers (nine lines) and ten PMTs and can be used for up to eight-color cytometry, simultaneously using as many as three laser lines. The Flow Cytometry Facility has partnered with Cytomation to develop a first of its kind biocontainment unit for the MoFlo designed to protect both the operator and the sample. The sorting laboratory is operated as a BSL-2+ facility and can sort live human cells and cells transfected with retroviral constructs.
The Facility offers the use of a high speed workstation equipped with a selection of flow cytometry analysis programs. The workstation can read data from a variety of media and is equipped with a 32X CD burner and 250MB zip drive for efficient archival of flow cytometry data files.
The most recent addition to the facility is an Amnis ImageStream 100 imaging flow cytometer, purchased with a Shared Instrumentation Grant from the National Institutes of Health. Imaging flow cytometry combines advantages of image analysis and conventional flow cytometry, enabling fluorescence to be associated with cellular features.