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Gel electrophoresis and MEKC join forces on chip
50 mm
SDS µCGE
but “the type of coating materials you matography column. That’s about Gel- and CE-based protein separations get for glass are much more mature and $400. The problem is, [glass chips] have come together at last on a plastic evolved than they are for polymers,” exdon’t last as long as chromatography chip. In this issue of Analytical Chemplains Soper. More effort is needed to istry (pp 3519–3527), Steven Soper and columns,” he says. “Plastic chips are develop coatings for polymers. “The about an order of magnitude lower in Hamed Shadpour of Louisiana State cost to produce. So, instead of spending [original] thought was you could use University describe a PMMA microfluthese polymers in their native form for $400, you can spend $40 or less on a idic chip on which proteins are separatdoing electrophoretic or chromatoed in two dimensions by SDS microcap- plastic chip that pretty much behaves graphic separations, [but] that’s turning the same way as the glass chips do.” illary gel electrophoresis (µCGE) and out not to be the case,” he says. They chose PMMA over other polymicellar electrokinetic chromatography Initially, the investigators labeled the mers because of its properties. PMMA (MEKC). With the new chip, a protein proteins with the Alexa Fluor mixture can be separated in 633 fluorophore and used ~12 min. (a) (b) laser-induced fluorescence Protein mixtures are typicalfor detection. But, because ly separated by isoelectric fomodifying proteins with fluocusing followed by SDS-PAGE. rophores is cumbersome and But the methods are labor-intime-consuming, they are tensive and involve unwieldy now focusing on detection slabs of gels. Efforts have been methods that don’t require made to ease the process by labels. One alternative is condeveloping multidimensional ductivity, in which the bulk CE separations, in both capilconductance of the fluid is lary and microfluidic formats. MEKC measured as proteins migrate In their approach, Soper and through the channel. MS is Shadpour have combined a bit also an option for detection. of both the gel- and CE-based Soper and Shadpour are methods. now using their chip to sepaThe investigators chose SDS µCGE, which separates (a) A PMMA microfluidic chip separates proteins by SDS µCGE and rate membrane proteins from stem cells. With >3000 proteins on the basis of size in MEKC. (b) The fluorescence image shows the interface between membrane proteins in the a gel inside a capillary, and the two dimensions. mixture, Soper says they are MEKC, which separates probehaves relatively well in separations be- close to reaching the chip’s predicted teins on the basis of how they interact peak capacity of 1000. cause proteins don’t readily stick to it with micelles, to increase the orthogoSoper points out that the chip lends and hurt the resolution. However, nality, resolution, and efficiency of proitself well to subsequent analyses. CurSoper points out, “The little caveat is, tein separations. First, Soper and Shadrently, if researchers want to analyze while PMMA works well in terms of pour determined that SDS µCGE and many proteins separated from a complex electrophoresis, as compared to other MEKC were orthogonal: The methods mixture on gels, they need to cherrypolymers, it does not perform as well as operate on different selection criteria pick the bands or spots containing the for separation but were compatible with glass. Glass seems to give you the best proteins out of the gels, digest them, performance characteristics for both each other. The investigators put SDS and then carry out an LC/MS analysis. chromatographic and electrophoretic µCGE in a 30-mm-long channel as the Soper says, “With this [chip], we comfirst dimension on the chip because it is separations.” pletely obviate the need for cherry-pickTo enhance PMMA’s performance, slower than MEKC, which took 10 s ing. The material comes off the column, the investigators coated the surfaces of per separation in their system. The and it can be sent to downstream prothe channels with methyl hydroxyethyl MEKC channel was 10 mm in length. cessors to digest the protein and make it cellulose, which acted as a dynamic Soper explains that they chose the ready for LC analysis and mass spectral coating (as opposed to a static coating, polymer PMMA over glass because of identification.” a which is permanently attached to the cost. “A glass-based microfluidic chip —Rajendrani Mukhopadhyay surface). Dynamic coatings aren’t new, will cost the same amount as a chroJ U N E 1 , 2 0 0 6 / A N A LY T I C A L C H E M I S T R Y
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