What once took days now takes hours thanks to Oregon Opportunity-funded laboratory
PORTLAND, Ore. – Cory Bystrom, Ph.D., remembers – not so fondly – how it once took him and two laboratory technicians a full day to prepare a total of 60 gel samples.
Thanks to the mapping of the human, mouse and other genomes in recent years, the process grew increasingly labor-intensive as demand for faster, more precise protein identification swelled. Technology couldn't keep up with discoveries.
But the labor intensity of the procedure is now a thing of the past for researchers in the Oregon Health & Science University School of Medicine. The Proteomics Shared Resource, an unassuming but state-of-the-art lab on the fifth floor of OHSU's Medical Research Building, can process more than three times as many protein samples in half a day, and much of it can be done by one person.
"This can do 180 samples in six hours," said Bystrom, the lab's director, motioning to the large, blue-hooded 2DiDx spotcutter and sample preparation workstation as he scans computer images of raw sample data collected the day before. "For us, it's absolutely great. If you were to do this manually, you're going to do about 20 a day."
The 2DiDx workstation is one of several sophisticated devices occupying nearly every corner of the lab, a research core of the OHSU Office of Research Development and Administration that opened Feb. 17. Funded by a grant from the Oregon Opportunity, the public-private, $500 million biomedical research funding initiative, the facility also contains a highly sensitive, laser-activated flouroimaging machine, which can provide detailed digital images of stained proteins suspended in gels, as well as a mass spectrometer the size of an upright piano, and a liquid chromatography machine.
And the latest software helps Bystrom and his research associate, Deb McMillen, manage the samples, analyze the proteins and mine genome databases around the world to help identify them.
But protein samples begin their journey toward identification in the 2DiDx workstation. The machine is fully automated and can prepare samples unattended with the help of a robotic arm that slowly pinpoints protein samples in gels visualized with fluorescent stains to enhance their identity, cuts the samples from the gels, and breaks them down with enzymes, preparing them for closer examination with the mass spectrometer. It can process up to 192 samples at a time.
"Each spot represents a protein," Bystrom explains, pointing to a gel sample. "This gel has hundreds to thousands of identifiable proteins on it. We use the robot to help us cut the spots. The (robot) head moves over, finds the spot and pokes a hole in the gel."
After the proteins are broken down at the 2DiDx workstation, the "few dozen to few hundred" peptides are sent to the mass spectrometer. There, high-voltage electrostatic charges and a pump flowing at 200 billionths of a liter per minute blast the liquid samples through a needle-nosed nozzle with an opening that is one-tenth the width of a human hair.
The result is a fine, liquid aerosol that generates peptide ions, which are then analyzed to help scientists determine their composition.
"You want to get the most out of the least amount of material. We need to use liquid chromatography to spread out the complex mixture of peptides," Bystrom said. The mass spectrometer "focuses on one peptide, breaks the peptide into pieces, and measures the fragments to get an idea of the amino acid sequence."
The process generates "enormous amounts of data," Bystrom said, so the lab uses the newest software to manage the samples, analyze the amino acid sequences produced in mass spectrometry and search international genome databases, where the sequences are compared to identify the proteins.
Such data crunching has been essential since last year's completion of the Human Genome Project, the massive effort led by the U.S. Department of Energy and the National Institutes of Health to identify the human genetic sequence. And as the genomes of organisms used in research become known, demand for services like those provided by the Proteomics Shared Resource will only increase.
"In the proteomics world, things are changing so rapidly," Bystrom said. "We in this lab have to be flexible in dealing with a wide range of samples, everything from bacteria to humans."
The term proteomics was coined in 1994 as an easy way to describe the concept of all proteins expressed by a genome, according to Bystrom. The term is now generally accepted to describe the cooperative use of a range of analytical tools to examine a population of proteins.
"The field of proteomics has been blown wide open by the completion of the human genome project, and the Proteomics Shared Resource allows us to be a leader in the protein field technologically," said Dan Dorsa, Ph.D., OHSU vice president for research and professor of physiology and pharmacology in the School of Medicine. "Until now, the processing, analysis and identification of proteins has been a time-consuming procedure, and creation of the proteomics core dramatically lowers this research hurdle."
Dorsa called the new proteomics lab the "perfect response" to the Oregon Opportunity initiative. "This new service will allow OHSU to remain competitive, and will undoubtedly increase OHSU's opportunities for collaboration with commercial entities."
Source: Eurekalert & othersLast reviewed: By John M. Grohol, Psy.D. on 21 Feb 2009
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