Dr. Baron explains that throughout his seven years at RML, he and others have contemplated how to use fluorescent tracking to learn more about TSEs, but they struggled to develop an effective method to do so.
"When I started working on TSEs, I thought about them as being similar to intracellular bacterial pathogens--something that replicates within an animal or human host cell," says Dr. Baron. "I wanted to know how such a pathogen binds to the host cell, and how it enters, replicates and spreads to other cells."
Dr. Baron says researchers have tracked infectious prion protein moving through other parts of animal bodies up to the brain, but no one had ever tracked the protein movement within animal brain cells. One of the most difficult aspects of the experiment, he says, was finding a way to fluorescently tag the TSE prion proteins without altering them--while still allowing researchers to identify the prions as they penetrated the cells and spread within the long projections that nerve cells develop to send signals to other nerve cells.
"This was difficult from a technical aspect because the scrapie pathogen is largely a corrupted form of a host cell protein," Dr. Baron said. "It can be hard to detect the corrupted prion protein in living infected cells and distinguish it from its normal counterpart."
He explains that once researchers learned how to mark the prion proteins, they added them to a culture of nerve cells and then began watching and taking photo images with a confocal microscope. Confocal microscopy uses laser light to scan many thin sections of a fluorescent sample, resulting in a clean three-dimensional image. The painstaking job of analyzing and deciphering about 1,
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Contact: Ken Pekoc
kpekoc@niaid.nih.gov
406-375-9690
NIH/National Institute of Allergy and Infectious Diseases
24-May-2005