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Flow Cytometry

Fluorescence-Activated Cell Sorter

Cheryl Baird, Principal Investigator

A PNNL researcher using the newest flow cytometer PNNL researchers use the newest flow cytometer, FACS-ARIA, to isolate and characterize antibodies. The machine is unparalleled in speed and utility, requires less operator time, and provides a higher fluorescent and sensitivity resolution.

Flow Cytometry allows high-throughput, single-cell investigation. Routinely used in the Microbial Cell Dynamics Laboratory (MCDL) at Pacific Northwest National Laboratory (PNNL), it rapidly interrogates bacteria, yeast, and mammalian cell populations on a single-cell basis. Fluorescent-activated cell sorting (FACS) has become an indispensable tool in biology. Recent technical advances allow fine resolution between cell types by measuring many optical parameters in parallel. FACS provides the ability to separate a heterogeneous suspension of cells into a purified fraction on the basis of fluorescence and light scattering properties.

PNNL is one of a handful of laboratories in the world that uses flow cytometry to isolate and characterize antibodies. PNNL was the first to create an immense library of nonimmune antibodies to be displayed on the surface of the yeast, Saccharonmyces cerevisiae. One advantage of displaying antibodies on the surface of a yeast cell is its compatibility with flow cytometry. This unique combination of technologies allows antibodies present in this library, which bind particular antigens of interest to the investigator, to be readily isolated after addition of fluorescently labeled antigen.

PNNL researcher using the flow cytometer A library of yeast cells is incubated with fluorescently labeled antigens and loaded into the BD FACS-Aria cell sorter. Those cells with antibodies capable of binding the antigen are visualized and isolated from the rest of the cellular population for further characterization.

To facilitate this research, PNNL has purchased the latest in flow cytometers, the FACS-ARIA from BD Biosciences. This state-of-the-art equipment provides a single-cell sorting capability at a rate of up to 70,000 events onto a variety of formats. PNNL will use the FACS-ARIA to quantitate, analyze, and isolate cells of interest. The increased speed and sensitivity available with this instrument will allow PNNL researchers to develop high-throughput methodologies to meet the challenges of the DOE's Genomics: Genomes to Life (GTL) program.

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