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AB Sciex and ISB Collaborating on Development of Libraries, Methods for SWATH Technology

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AB Sciex said this week that it has signed a multi-year collaboration agreement with the Institute of Systems Biology to develop methods related to its SWATH mass spectrometry technology.

Specifically, the collaboration will focus on configuring the selected-reaction monitoring mass spec libraries collected by ISB via its SRMAtlas project for use with the SWATH technique, Robert Moritz, director of proteomics at ISB, told ProteoMonitor.

The collaboration will also investigate the use of AB Sciex's Selexion ion mobility technology with SWATH, he added.

SWATH is a data-independent acquisition technique enabled by AB Sciex's TripleTOF 5600+ mass spectrometer. In DIA mass spec, instruments select broad m/z windows and fragment all precursor ions in that window, allowing for collection of MS/MS spectra on all ions in a sample.

The technique was first introduced in a 2004 Nature Methods paper by Scripps Institute researcher John Yates III, who implemented it on a ThermoElectron LTQ instrument. Since then it has been employed by a number of proteomics researchers using a variety of different platforms.

As University of Washington researcher Mike MacCoss told ProteoMonitor last year, what has set SWATH apart from these previous versions of the technique is AB Sciex's emphasis on applying it to targeted proteomics and providing researchers with the tools to do so (PM 6/8/2012).

The ISB collaboration can be seen as another step in that effort, with the company leveraging the institute's targeted proteomics data to provide users with spectral libraries they can use in SWATH analysis.

SWATH doesn't use conventional proteomic search database engines but, rather, makes peptide IDs by matching experimentally acquired spectra to reference spectra, meaning that researchers need reference spectra libraries for the samples they want to analyze.

"The goal," Moritz said, "is to utilize our existing data and to develop new spectral libraries so that new [SWATH] users can quickly adopt the technology without having to go through the pain of having to develop those libraries for themselves."

Specifically, the ISB is configuring the SRM-MS spectra contained in its SRMAtlas database for use with SWATH.

The SRMAtlas is a collaborative effort between the ISB and the Swiss Federal Institute of Technology to develop sets of SRM-MS assays for proteomic research. The project launched in October 2009 with funding from the National Human Genome Research Institute, which provided $2.7 million in direct funds under the American Recovery and Reinvestment Act. It also received €2.7 million ($4.1 million) from the European Research Council.

In 2010, the ISB and ETH Zurich announced they had completed an initial version of the library comprising more than 170,000 SRM assays – one each for at least five proteotypic peptides for each of the 20,300 human genes currently annotated as protein-encoding (PM 9/24/2010).

Since then the researchers have continued to add data to the library, including assays for modified proteins and proteins in a variety of organisms. Currently, Moritz said, they are in the process of releasing an N-glycoform version of the human library and are also working on libraries for mouse, yeast, Drosophila, Escherichia coli, and Mycobacterium tuberculosis.

"We've been compiling these libraries and building in more and more data so they become more comprehensive over time," Moritz said. "And it's these libraries that we will now configure for SWATH analysis and make available to any researcher for free use."

The process is simplified by the comprehensiveness of the existing SRMAtlas data, Moritz said, noting that the researchers would not need to reacquire spectra on the TripleTOF 5600 but could simply reconfigure the existing SRM data.

"In our development [work] over the last few years, to really future-proof our data collection we really comprehensively analyzed each of the peptides that we looked at across different instruments and across multiple parameters on the same instrument," he said. "We made a very conscious decision to be very comprehensive with our data collection so that newer technologies could take advantage of the data."

While his team will focus on reconfiguring its SRM data for use with the SWATH method, because the data is freely available, other vendors or researchers can take it and optimize it for their own DIA methods if they desire, Moritz said. In addition to AB Sciex, Agilent, Bruker, Thermo Fisher Scientific, and Waters all either have or are developing DIA methods for their instruments.

The ISB-AB Sciex announcement follows the release last month of another large spectral library intended to ease targeted proteomic method development – that one a paper in Nature presenting ion trap and triple quadrupole-based reference fragment ion spectra for peptides covering 97 percent of the predicted yeast proteome (PM 1/25/2013). Mortiz was an author on that paper, along with his SRMAtlas collaborator and ETH Zurich researcher Ruedi Aebersold.

In addition to repurposing the SRMAtlas data for the SWATH method, the collaboration will look at applying AB Sciex's Selexion ion mobility technology to SWATH, Moritz said.

The Selexion technology is considered by many researchers to be a form of field asymmetric ion mobility spectrometry, or FAIMS, although AB Sciex prefers to maintain that it is a distinct technology, perhaps because of FAIMS' somewhat troubled past.

FAIMS works by running mixtures of ions at atmospheric pressure between two electrodes and subjecting them to alternating high- and low-field conditions by applying an asymmetric RF waveform, which separates them based on differences in ion mobility under high and low electric fields. Depending on their mass and charge, some ions will drift into the electrodes while others will pass through the device, allowing, for instance, a researcher to select low-abundance peptides for analysis from a tryptic digest while weeding out higher-abundance ions.

The technology can be inserted between a traditional liquid chromatography system and a mass spec instrument, allowing for an additional level of separation.

Moritz is currently working under a National Science Foundation grant to develop FAIMS capabilities for high-resolution mass spectrometry and said that his lab has found that "with a simple application of FAIMS we can immediately double the [number of protein] identifications."

The Selexion technology is not yet available on the TripleTOF 5600, but, Moritz said, the company plans to release a version of technology for that instrument later this year and his team will be "taking our knowledge of FAIMS and applying it to the AB Sciex version and seeing what benefits we can get for SWATH with FAIMS on there as well."

Financial and other terms of the agreement were not disclosed.

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