Research at Fox Chase »Research Facilities »Cell Biology

Richard R. Hardy, PhD Director Jim Oesterling, BS Scientific Technician

Reimann Building, Room R385 215-728-2463 Richard.Hardy@fccc.edu

Function

The Flow Cytometry and Cell Sorting Facility offers research scientists at Fox Chase the capability of analyzing cells by up to fourteen independent measurements (2 light scatter, 12-fluorescence).

Major applications of flow cytometry include discrimination and isolation (sorting) of cell subpopulations (at up to 30,000 cells/ sec); isolation of rare variant (1/107) cell lines, characterization of tumors or cell lines, single cell cloning of cell lines by direct deposition into 96-well plates, measurement of calcium flux, and cell cycle analysis. Facility personnel operate the 12-color cell sorter, whereas users run their own samples on two analysis-only instruments.

Facility personnel also provide users with advice on experimental design (including availability of reagents and appropriate fluorochromes), sample preparation, and data interpretation.

Description

The facility includes a Becton Dickinson FACS-VantageSE/DiVa flow cytometer equipped with three lasers, one UV-capable. Up to four streams (independent populations) can be sorted simultaneously.

Sorting is monitored by two CCD video cameras, facilitating sorts lasting for extended periods (>1 hour). Sterile sorting is routinely provided at no extra charge. The UV laser facilitates measurement of Ca++ flux, using the calcium sensitive reporter, Indo-1. There are also two user-operated analysis-only instruments: 1) a three-laser 10-color Becton Dickinson LSR-II; and 2) a 3-color Becton Dickinson FACScan flow analyzer, particularly suited for DNA cell cycle analysis and simple fluorescence analysis. Scheduling time is done through an online calendar application. Rapid analytical throughput (particularly for complex multi-color samples) is facilitated by an in-house developed "Flow-LIMS" that allows users to create "protocols" describing their experiments in advance in a web browser interface on any lab computer, then collect data into the system which completely annotates each sample. All data collected with this system are available from a permanent archive, accessible on any networked computer. The desktop application "FlowJo", available through the facility, inter-operates with the Flow-LIMS to provide state-of-the-art analytic capabilities.

Finally, the facility makes available to users a fluorescence microscope for imaging stained samples and a Milteny Biotek MACS magnetic cell separation device. The latter is particularly useful for enriching or depleting cell populations prior to final isolation by sorting on the Vantage.