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The Flexar liquid chromatography platform comes in three formats: conventional-pumping; “near” ultra-high pressure LC (UHPLC); and “true” UHPLC. (Source: PerkinElmer) |
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Today’s researchers want more options from chromatography (see "Redesigning Protein Purification"). So when
PerkinElmer (Waltham, Mass.) started to develop its new Flexar liquid chromatography (LC) platform, Alessandro Baldi, PhD, the company’s chromatography marketing manager, says that they focused on one thing: choice. "We understand that customers want to pick the right LC technology and performance based on their needs of throughput, existing analytical methods, and budget."
To provide those options, PerkinElmer’s Flexar provides three tiers of performance. The conventional-pumping Flexar provides up to 6,000 pounds per square inch (psi). The "near" ultra-high pressure LC (UHPLC) Flexar goes up to 10,000 psi. Finally, the "true" UHPLC model can take pumping pressure up to 15,000 psi. "With our ‘near’ UHPLC," says Baldi, "a customer can port a conventional LC application to a new level of throughput—often an increase of 3- to 4-fold—without lots of disruption." Flexar’s "near" UHPLC also increases sensitivity, sometimes by 10 to 20 times. Moving to "true" UHPLC does take some modifications in approach and method development, but this can be well worth the time for applications—such as protein digestion—where every peak might contain valuable information.
Thermo Fisher Scientific (Waltham, Mass.) is also advancing chromatography with its Exactive Benchtop LC-MS. According to Lester Taylor, marketing director of life sciences, the Exactive provides resolutions of 100,000 at one scan per second or 10,000 at 10 scans per second. This LC-MS instrument also provides a sensitivity of 500 femtograms with a signal-to-noise ratio of 10 to 1. Taylor says that the Exactive provides a range of benefits, including ultra-fast method development and "multiple-residue analysis without sensitivity compromise." Taylor adds that the high resolution provides the ability to "screen and quantify compounds without setting up defined methods."
The net result of these new instruments is increased throughput and more options than ever before.
Redesigning Protein Purification
To purify proteins, researchers often use affinity tags in concert with liquid chromatography (LC). Bio-Rad (Hercules, Calif.) recently made the tag-cleavage process faster and easier with its Profinity eXact Fusion Tag System. This Escherichia coli-based system, says Tanis Correa, Bio-Rad’s senior product manager of chromatography, "provides parallel purification and on-column cleavage of affinity tags without a protease." Structure and activity of native proteins are maintained, and the need for additional materials, like cleavage enzymes and purification resin for post-cleavage enzyme removal, is eliminated. The Profinity eXact resin includes immobilized protease on the chromatography beads. As Correa says, "With a 30-minute cleavage incubation, Profinity eXact offers a significant improvement over the traditional tag-cleavage workflow in which incubations can take as long as 48 hours to generate a precise cut."
Kevin D. Ridge, PhD, of the University of Texas Health Science Center in Houston, used this system to express and purify the G-protein alpha subunit and a mutant form of arrestin from the vertebrate visual system. Ridge says, "My laboratory rates [this] system very highly as it provides facile isolation of highly purified and stable protein preparations in essentially a single step."
Eksigent (Dublin, Calif.) also recently added a new option for protein research with its cHiPLC-nanoflex System. This docking station holds up to three microfluidic chips containing nanoLC columns. This can be used with Eksigent’s nanoLC systems or the Tempo from Applied Biosystems (Foster City, Calif.). According to Remco van Soest, vice president of product management at Eksigent, "Since the microfabricated chips used for columns and traps are manufactured with semiconductor manufacturing methods, part-to-part variability is minimized—leading directly to improved precision that stands out from the competition and allowing Eksigent to provide a specification for column-to-column retention time of 2% relative standard deviation or better."
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About the Author
Mike May is a publishing consultant for science and technology based in Houston, Texas.
This article was published in Drug Discovery & Development magazine: Vol. 12, No. 6, May, 2009, p. 10.
