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Agilent, Duquesne U Form Mass Spec Center of Excellence

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By Tony Fong

Agilent Technologies and Duquesne University this week launched a mass spectrometry center of excellence at the school to further mass spec-based research in the western Pennsylvania region.

The COE pulls together the mass spec-based instruments and resources at Duquesne into two dedicated laboratories, and serves as a regional showcase for Agilent to display its mass spec technologies. As part of the collaboration between the school and Agilent, Duquesne researchers also will be developing methods for Agilent instruments and evaluating new technologies.

According to Duquesne officials, the COE could also become a mass spec core facility for the school down the road.

The facility is outfitted with three Agilent mass specs: a 6500 Series Accurate-Mass Quadrupole Time-of-Flight LC/MS; a 6400 Series Triple Quadrupole LC/MS; and a 6200 Series Accurate-Mass TOF MS.

In addition, the COE has an Agilent 1200 Series Rapid Resolution LC system and 1200 Series HPLC-Chip/MS systems. Agilent's Spectrum Mill for MassHunter Workstation software is also being used for protein and peptide identifications.

While the COE was officially "christened" this week, it had its beginning about six years ago, Skip Kingston, a professor of chemistry and biochemistry at Duquesne and the co-principal investigator at the center, told ProteoMonitor. It was then that he received a National Science Foundation grant for the purchase of the school's first TOF high-resolution mass spec, an 1100 Series MALDI/LC TOF instrument from Agilent.

About a year and a half ago, Kingston and Mitchell Johnson, an associate professor of chemistry and biochemistry and the co-PI at the COE, received a second NSF grant, allowing them to purchase additional mass specs.

With it, the school acquired Agilent's 6500 and 6400 platforms. The Agilent 6200 instrument is an upgrade to the original 1100 platform purchased in 2004.

Separate from the COE, an ICP-MS platform, also from Agilent, was purchased for analysis of inorganic materials.

The decision to purchase Agilent instruments was made after Kingston and his colleagues conducted an extensive evaluation of instruments by all the major mass spec vendors, Kingston said.

"The dean has said it was probably the most careful evaluation of equipment for a large purchase that a university has ever done," he said, adding the process took about two months.

"We went over all of major manufacturers' equipment, we looked at all of the specifications and [had] a list of very rigorous criteria," he said. "Agilent technology rose to the top and we purchased the top of the line equipment from them."

For its part, Agilent provided a "steep discount" to the school on the instruments, said David Seybert, dean of the Bayer School of Natural and Environmental Sciences.

An Agilent official did not respond to a request for an interview for this story. According to Kingston, though, Agilent made the mass spec facility a COE because "we have put forth so much effort over a number of years into research in mass spectrometry [and] have a unique capability."

Agilent's collaboration with Duquesne is likely also part of the company's stated goal of building up its academic and government business. For the past two years, Agilent has repeatedly said that one of its main priorities, especially in the life sciences, is to increase its revenue stream from the academic and government sector, which currently stands at about 5 percent of its total revenues.

The collaboration benefits Agilent in another way — Kingston and his colleagues will be testing the company's platforms and tools and providing feedback on modifications to the instruments and tweaks to software, he said.

And while the Duquesne researchers could have bought and installed the instruments without the center being named a COE, the designation benefits the school and the center by giving them more visibility within the region and establishes the mass spec facility as a leading mass spec resource for other institutions, school officials said.

"We actually have a consortium of other colleges and universities in the western Pennsylvania region who are really part of the collaborative group that will be using that center," Seybert said. "So we make it available for their students to come in and gain experience on" the mass specs.

Ultimately, the goal is to turn the center into a core facility. While Kingston and Johnson are the co-PIs for the facility, they are exploring possibly hiring a lab director and dedicated staff. As requests from outside Duquesne to use the equipment increase, the PIs are also creating a fee structure.

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"This is a major resource for the western Pennsylvania region, so we have a lot of smaller colleges now that will be coming in [and] teaching classes for their students," Kingston said. "We're working on a very ambitious management structure and that hasn't been finished yet."

Too Sensitive for a Lab

In order to store all the new equipment, the school "had to redesign part of the building to handle these new sophisticated instruments," Kingston said. That included building a new cleanroom in addition to two that were already available.

"These instruments are so sensitive, you can't put them in a standard laboratory," Kingston said. And because they consume massive amounts of liquid nitrogen, a 1,200-gallon liquid nitrogen tank was installed outside the building where the instruments are housed to supply a continuous uninterrupted flow of the chemical to the machines.

The new platforms will be used to further research that Duquesne staff has been investigating, including projects in proteomics.

"We didn't have nearly the capabilities that these instruments provide, so we didn't do proteomics work as we didn't have instrumentation capable of doing this type of proteomics," Seybart said, adding the new platforms enable the school to do more clinical research and collaborate with nearby institutions such as the Children's Institute of Pittsburgh, with whom Kingston is working to investigate biomarkers for autism.

In prior epigenomic research, Kingston identified a biomarker for autism. The task now is to discover and identify protein biomarkers for the condition.

In one project, he and his colleagues are trying to discover diagnostic biomarkers in children with autism that may be associated with immunological response to the environment. About 70 percent of autistic children also have immune dysregulation, Kingston said.

Over a period of weeks, the children are being placed in cleanrooms to sleep, "and we're looking for both their proteomic and their biological components of toxins, and then we're looking at the end to see if we've reset their immune systems."

A second autism-related project is targeted at the discovery of biomarkers associated with damage from toxins. Kingston and his co-researchers are examining the proteomic content of autistic children and comparing that to non-autistic children. In particular, Kingston and his colleagues are interested in their red blood cell content, he said.

"We're also measuring 70 toxins," Kingston said. "We'll also have the toxin load in each of these children."

Also, Kingston is working with researchers at the Allegheny General Hospital Allegheny Singer Research Institute to investigate inherited "knock-out diseases" in which the body stops making a necessary enzyme. Their work is focused on Fabry's disease where a deficiency in alpha glactosidase A can lead to a malfunctioning of blood vessels, tissues, or organs.

Patients with Fabry's disease need to undergo enzyme replacement therapy.

Kingston and researchers at AGHASRI are working to create a mass spec-based assay to measure the enzymes and proteins in patients with the disease.

The work is still in the proof-of-concept stage, but "we have proved that the mass spectrometers measure these enzymes and proteins far better than the older techniques of fluorescence," Kingston said. "It's about an order of magnitude better."

Partha Basu, an associate professor of chemistry and biochemistry who runs Duquesne's Center for Metals in Biological Systems, is pursuing environmental proteomics, the study of how the proteome of organisms such as bacteria respond to environment changes. In February, he and colleagues published a study in Metallomics based on a proteomic investigation of roxarsone degradation by Alkaliphilus oremlandi strain OhILAs.

The study was conducted using MALDI mass spectrometry and 2D gels, which identified about 90 proteins. Using the new LC/MS system, he and his colleagues have "inferred" about 400 proteins so far, he said, resulting in more pathway and expression data.

"So the magnitude is different," he said.

In addition to proteomics, the COE will support research in metabolomics and trace lipid analysis; how veterinary drugs affect the food chain; food safety analysis; and factors affecting the immune system, according to a statement from Duquesne and Agilent.

In the statement, Johnson, who is conducting research primarily in metabolomics and lipidomics, said, "The difference between what we can do now and what we could do before is incredible. It's moved us into an entirely new echelon of research that is now possible at Duquesne."

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