news
Defining the molecular forces that shape protein nanofibrils
N
N
Amyloid fibrils are best each. They found that (a) (b) (c) known as agents of disor the bending rigidities of α-Lactalbumin ders such as Alzheimer’s each type of molecule 30 and prion diseases, but varied over ~4 orders of 25 more recently they have magnitude and that a cor 20 15 been recognized as a com relation exists between 10 mon protein structure that cross-sectional moment 5 can form from a variety of inertia and bending 0 of polypeptides. These rigidity. The researchers Insulin B-chain polypeptides are diverse say that this correlation 25 in sequence and native strongly supports the idea 20 structure, and the factors that a similar set of forces 15 that govern the forma stabilizes all types of amy 10 tion of fibrils are poorly loid fibrils. 5 understood. The researchers also 0 Now, Christopher Dob examined different mod 0 2 4 6 8 10 12 son, Mark Welland, and els for the fibrils’ behavior Height (nm) their colleagues at the and found that the domi University of Cambridge Comparison of (a) AFM topographic images, (b) the distribution of heights, nant backbone interac (U.K.) have carried out and (c) the shapes of representative amyloid fibrils. Scale bars = 250 nm. tion contributing to their a detailed survey of the (Adapted with permission. Copyright 2007 American Association for the formation is their exten amyloid fibrils that form Advancement of Science.) sive hydrogen-bonding from a wide variety of network. They say that polypeptide chains. Their goal was to To derive the structures’ mechani the differing side-chain interactions of better define the material properties of cal properties, they probed the fibrils the individual sequences shape the more each type of fibril to understand what with atomic force microscopy (AFM) subtle differences seen in their AFM forces govern their formation. and recorded the height and shape of studies. (Science 2007, 318, 1900–1903)
Understanding noise in solid-state nanopores Despite having many advantages over their biologically derived kin, solid-state nanopores have always been plagued by problems with ionic current fluctuation, or “noise”, which limits their sensitivity and reliability. Now, Cees Dekker and colleagues at the Delft University of Technology (The Netherlands) have carried out a detailed study of the noise in solidstate nanopores. The researchers looked at both high-
© 2008 American Chemical Societ y
frequency and low-frequency noise in a variety of nanopores with similar diameters but different resistances. They found that the low-frequency noise varied much more from pore to pore than the higherfrequency version. The scientists also studied low-frequency noise in systems with various ionic strengths. They found that as conductivity increased with rising salt concentration, the noise power strongly de-
creased. They conclude that the noise level is related to the number of charge carriers on the nanopore. Finally, the researchers applied their results to DNA translocation through nanopores. They modeled S/N with their data and found that a better S/N was obtained in larger (>20 nm) nanopores under low-salt versus high-salt conditions. (Proc. Natl. Acad. Sci. U.S.A. 2008, 105, 417–421)
Ma r c h 1 , 2 0 0 8 / A n a l y t i c a l C h e m i s t r y
1349
news
DataChip tackles high-throughput drug screening Pharmaceutical companies have to de-
ing the response of a human carcinoma
the previously developed MetaChip (Proc.
termine the toxicity of a large number
cell line to toxic compounds. Three com-
Natl. Acad. Sci. U.S.A. 2005, 102, 983–
of potential drugs early in the discovery
pounds were each spotted at concentra-
987) so that the spots contained a sin-
process. To do so, they need in vitro as-
tions ≥1 mM onto a slide in a 14 × 40 ar-
gle human P450, a mixture of three pro-
says that avoid animal testing. Jonathan
ray. The array was then stamped on top
teins, or no P450 in a 20 × 54 array. Test
Dordick, Douglas Clark, and colleagues
of the DataChip. At the end of the incu-
compounds were added to the MetaChip.
at Solidus Biosciences, Rensselaer Poly-
bation period, the array was washed, the
The MetaChip was then stamped on a
technic Institute, and the University of
cells were grown for 3 days, and their vi-
DataChip with liver cells.
California Berkeley have developed a min-
ability was measured. The IC 50 values of
iaturized 3D array of cell cultures called
the compounds—the concentration of an
cultures, with the MetaChip, simulta
the data analysis toxicology assay chip
inhibitor needed for 50% inhibition—from
neously provided IC 50 values for nine
(DataChip) for high-throughput toxici-
the DataChip agreed well with those ob-
compounds and their metabolites. Close-
ty screening of drug candidates and their
tained from 96-well cell cultures.
ly matched responses from the DataChip
cytochrome P450-generated metabolites. The DataChip consists of an array of
Dordick, Clark, and colleagues next in-
The DataChip of 1080 individual cell
and 96-well plate assays confirmed that
corporated information from cytochrome
the ~2000-fold miniaturization of the array
cells trapped in a hydrogel matrix in vol-
P450, a large and diverse superfamily
didn’t hamper the cytotoxicity response.
umes as small as 20 nL. The investi-
of enzymes that break down toxic com-
(Proc. Natl. Acad. Sci. U.S.A. 2008, 105,
gators evaluated the array by measur-
pounds, into the DataChip. They modified
59–63)
Dielectrophoresis with reconfigurable oil barriers Mark Burns, Jennifer Linder electric field lines to create the man, and Prasanna Thwar of field gradients. These objects can the University of Michigan have be, for example, a narrow chan designed a new system for elec nel between two insulating sur trodeless dielectrophoresis (DEP) faces or insulating posts placed in that uses small menisci of oil to the middle of the flow. Both of t = 1.5 s t=0 t = 3.15 s shape the field gradient. DEP these choices must be built with differentially polarizes dielectric microfabrication techniques. particles and controls their move In the current work, the scien ment by directing them either tists instead added to the system toward the location of maximal two to four channels that pro field gradient (positive DEP) or trude from the side walls of the toward the location of minimum main channel and pipe oil into t = 12.15 s t = 8.15 s t = 20.5 s field gradient (negative DEP). It the system. With careful con is a common method to control Time-sequence photographs of a single fluorescent polysty- trol, the oil will form a bulging the flow of particles in microflu rene particle trapped by an oil-menisci-created electric field. meniscus that extends into the idic devices. The particle is circled in each image. (Adapted with permischannel and shapes the electric In traditional DEP, a highsion. Copyright 2007 Wiley-VCH Verlag GmbH & Co. KGaA.) field lines. The researchers note frequency ac voltage is applied that their method provides flex on a planar microelectrode or metallic problem has led to the development of ibility and dynamic control and even structure patterned on the interior of a “electrodeless” DEP, where a voltage is allows for adjustment of the dimensions microfluidic channel, but electrolysis at applied with two electrodes placed at of the electric field gradients during the the electrode surface limits the maxi the ends of the channel and insulating course of an experiment. (Electrophoresis mum voltage that can be applied. This objects are placed in the path of the 2007, 28, 4572–4581) 1350
A n a l y t i c a l C h e m i s t r y / Ma r c h 1 , 2 0 0 8
news
A multiwell, multicantilever, protein detection system Arun Majumdar and colleagues at the University of California Berkeley, the University of Southern California, and Lawrence Berkeley National Laboratory have fabricated a new label-free proteinbinding sensor chip. Each chip contains 80–120 reaction wells with 4–8 inde pendent microcantilever sensors. Previ ously, a maximum of eight cantilevers has been displayed on a single chip. The researchers explored several dif ferent combinations of agents for func tionalizing the active surfaces and pas sivating inactive surfaces. They found that the best combination consisted of either 3,3′-dithiobis-[sulfosuccinimidyl propionate] (DTSSP) or an N-hydroxy succinimide (NHS)-thiol/polyethylene glycol (PEG)-thiol mixture for func tionalization and PEG-silane as the passivation medium. As a proof of principle, the scien
tists attached an antibody for prostate-specific antigen (PSA) to the surface of their cantilevers through the DTSSP or NHSthiol/PEG-thiol linkages and then blocked the inactive surfaces with PEG-silane. Binding of PSA was monitored by illuminating the gold surface of the cantilevers with a laser and recording their individual deflections with a CCD camera. The researchers calculated the limit of detection of their device to be 0.2 ng/mL, which is an order of magnitude lower than the 2 ng/mL clinical cutoff level. They note that their device could be used for any binding pair and that it may also eventually be used to carry out multiplexed assays. (Nano Lett. 2008, DOI 10.1021/nl072740c)
(a)
Micropipette Si
Glass (b)
(c)
Laser 100 µm
(a) Schematic of multiple cantilevers in each reaction well. Laser light is reflected off each cantilever and is used to monitor its deflection. (b) The size of a single chip. (c) SEM image of three cantilevers in a reaction well.
Proteomic profiling method not suitable for detecting prostate cancer The use of SELDI TOFMS for early iden-
across laboratories. In addition, the de-
could not distinguish healthy from can-
tification of cancers has been controver-
cision algorithm could correctly identi-
cerous samples nor could it discriminate
sial. As a result of these and other ques-
fy samples from cancer patients when the
between samples from more aggressive
tions about diagnostic approaches, the
samples originated from the patient co-
versus less aggressive cancers.
National Cancer Institute Early Detec-
hort used to derive the algorithm.
Semmes and colleagues note that a
tion Research Network organized a vali-
In the first of two new papers pub-
major challenge for biomarker research
dation study of serum proteomic profiling
lished by Semmes et al., the decision al-
is minimizing the incidence of false dis-
to diagnose prostate cancer. Two recent
gorithm could not correctly distinguish
covery from biased samples. For that, re-
papers published by a collaboration in-
between healthy and cancer patients
searchers must spend the time and effort
volving 29 researchers at 10 institutions
when used to analyze geographically di-
to put together enough “ideal” specimens.
present the results from the second stage
verse samples that were not from the co-
The authors say that their results do
of the validation study.
hort used to derive the algorithm. This
not indicate whether a particular meth-
In a previous publication, O. John
prompted the group to look for evidence
od works and do not imply that previous
Semmes from Eastern Virginia Medical
of sample bias, which they found. In the
studies were wrong. However, they do
School and colleagues determined in the
companion paper, the authors compiled a
conclude that the SELDI TOFMS method
first stage of the study that with frequent
new cohort of specimens that eliminated
used in their study has no diagnostic val-
instrument calibration and automated
the sample bias, and they derived anoth-
ue. Finally, they recommend that all other
sample preparation, the SELDI TOFMS
er decision algorithm from the new sam-
biomarker assays be rigorously validated.
method was sufficiently reproducible
ple cohort. However, this new algorithm
(Clin. Chem. 2008, 54, 44–52; 53–60)
Ma r c h 1 , 2 0 0 8 / A n a l y t i c a l C h e m i s t r y
13 51
news
Whole-organism imaging MS
MALDI MS of transition-metal catalysts
Most imaging MS (MALDI IMS) studies involve the analysis of organs or tissue sections. To gain a more complete understanding of processes taking place within an entire organism, however, Tuhin Sinha and colleagues at Vanderbilt University developed a method to apply 3D MALDI IMS to whole-animal tissue sections. They also created a tool to combine MALDI IMS images with other types of data, such as those from MRI experiments.
x
x
0 2.6
0
for organic transformations, but often their exact structures and mechanisms of action are poorly understood by researchers. Because these catalysts TUHIN SINHA
1 150
2.1
2.1
Transition-metal complexes can serve as catalysts
have so many relevant applications, including pharmaceutical and polymer synthesis, great effort has been made to understand and tailor them for specific reactions.
z
0
y
2.1
2.6
z
0
y
2.1 0
0
Transition-metal complexes are notoriously airand water-sensitive. MALDI MS can be used to char-
The left image in each graph represents aligned MALDI IMS and MRI data obtained from a tissue slice through a mouse head. The MRI data are depicted in black and white, whereas the levels of a protein detected with MALDI IMS are represented by the colored area. On the right in each graph is a reconstructed optical image of the mouse head. Data for (left cube) Pea15 and (right cube) Fabp5 are shown.
acterize the complexes, but transferring samples
With in vivo MRI, the researchers imaged the heads of mice with brain tumors. The mice were then sacrificed, perfused with saline, and frozen. MALDI IMS images were acquired from selected tissue sections throughout whole mice with a lateral resolution of 150– 300 μm. To generate spatially resolved 3D volume reconstructions of entire mice, Sinha and colleagues performed several postprocess ing steps that matched the mass spectra to the targeting image, which was aligned with optical images of the tissue slices. MRI scans of mouse brains also were aligned with optical images of the tissue slices so they could be compared with the MALDI IMS results. In overlays, regions of high protein concentration detected in the MALDI IMS studies corresponded well to MRI contrast varia tions. In addition, brain regions within the tumor area were signifi cantly different in terms of the measured MALDI and MRI param eters compared with other regions. (Nat. Methods 2008, 5, 57–59)
a glovebox and a MALDI MS instrument. Their de-
from an inert-atmosphere glovebox to the spectrometer often leads to rapid decomposition. To address this problem, Deryn Fogg and colleagues at the University of Ottawa have interfaced sign projects the mass spectrometer loading chamber directly into the glovebox. The scientists also explored the issue of finding an appropriate matrix for ionization of transition metals and found that the charge-transfer ionization method works the best. The researchers note that their assembly also could be used for assaying air-sensitive reactions to monitor reaction progress. (Angew. Chem., Int. Ed. 2008, 47, 303–306)
Bard and Moerner win the 2008 Wolf Prize in Chemistry Allen Bard of the University of Texas Austin and William Moerner of Stanford University have been awarded the 2008 Wolf Prize in Chemistry. Bard was editorin-chief of the Journal of the American Chemical Society from 1982 to 2001. He
1352
pioneered the development of the scanning electrochemical microscope, which allows for high-resolution chemical imaging of surfaces and the study of chemical reactions at the nanoscale. Moerner was the first to perform optical detection
A n a l y t i c a l C h e m i s t r y / Ma r c h 1 , 2 0 0 8
and spectroscopy of a single molecule in condensed matter. The Wolf Prize is a $100,000 award created to honor scientists and artists “for achievements in the interest of mankind and friendly relations among peoples.”
