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ANALYTICAL CURRENTS
XPS analysis of DNA films Although DNA is now routinely immobilized
Other techniques, such as radiolabeling,
Knowing the film thickness, they could
onto surfaces to create biosensors and mi-
have been used to characterize immobilized
correct the XPS peaks for attenuation and
croarrays, it is difficult to quantify the ele-
DNA, but safety concerns are driving re-
calculate the concentrations of the four
mental composition of the DNA and how
searchers to explore label-free techniques,
principal elements—nitrogen, carbon,
much of it actually binds to the surface. To
such as ellipsometry, surface plasmon res-
phosphorus, and oxygen (hydrogen is not
obtain this information, which is crucial for
onance (SPR), and XPS. Petrovykh and co-
detected by XPS)—in the DNA film. The
determining the efficiency of the immobi-
workers chose XPS to reduce complica-
carbon to nitrogen and oxygen to nitrogen
lization process, Dmitri Petrovykh and co-
tions with nonspecific adsorption, which
ratios indicated that the thickest films were
workers at the University of Maryland, the
can occur with ellipsometry and SPR.
not hydrated and that little nonspecific ad-
Naval Research Laboratory, and the Nation-
ssDNA molecules consisting of only thy-
sorption of other species occurred. On the basis of film thickness and ele-
al Institute of Standards and Technology
mine bases, which have a simpler chemical
developed and validated an X-ray photo-
structure than the other three bases, were
mental concentrations, the researchers
electron spectroscopy (XPS) method, using
immobilized onto gold surfaces via a thiol
calculated absolute and relative surface
films of thiol-modified single-stranded DNA
group by a process that appears to be similar
coverages. After a 20-h immobilization
(ssDNA) molecules on gold substrates as a
to the well-studied self-assembly of alkanethi-
process, the absolute DNA coverage was
model system. The method should be appli-
ols. The researchers performed XPS analysis
3.7 1013 molecules/cm2. The results are
cable to DNA films prepared under a wide
on the gold-DNA samples and compared the
in agreement with those obtained under
range of conditions, allowing for the direct
results with those from clean gold samples
the same conditions by radiolabeling and
quantitative comparison of them.
in order to determine the film thickness.
F TIR. (Langmuir 2004, 20, 429–440)
Nanospheres for separating DNA Mari Tabuchi and colleagues at the University of Tokushima and several other universities and institutes in Japan have developed a new technique for separating DNA fragments with nanospheres. The approach, which involves two pressurization steps in conjunction with microchip electrophoresis, is faster than conventional separation methods and covers a wider range of DNA fragment sizes. At the heart of the new technique are core-shell type nanospheres, 30 nm in diam, created by multimolecular micellization and polymerization of poly(ethylene glycol) (PEG) with poly(lactic acid) (PLA). The hydrophobic PLA segments contain methacryloyl groups at their chain ends and form the core of the sphere, which is covered with the flexible hydrophilic PEG chains. The closely packed structure of the nanospheres makes them suitable packing material for separations in microchannels. An essential component of the nano114 A
Hydrophilic PEG chain
Hydrophobic PLA chain
sphere system is a pressurization technique. Samples are injected Polymerizable Water under pressure (2.5 kPa for 1 s) double bond PEG shell into a microchip channel filled with a 1% solution of the nanospheres. Crosslinking Just before the electrophoretic sepCrosslinked aration, secondary pressure (2.5 kPa PLA core for 1 s) is applied to the nanoNanosphere spheres to speed up the separation. A broad band forms, which is then Nanospheres can be combined with a double-presfocused by the electrophoretic surization technique to speed up DNA separations. process. When samples are injected (Adapted with permission. Copyright 2004 Macmillan without pressurization, an intense Publishing Ltd.) band does not form and no peaks appear. Similarly, when samples are inject- Likewise, the researchers could separate 100–1000 bp fragments in 60 s using the ed into a conventional polymer medium, nanospheres vs 130 s in methylcellulose. such as methylcellulose, no bands form In contrast to conventional methods, which even with the pressurization technique. are typically limited to separating DNA Using the new nanosphere system, fragments
