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Kelvin probe force microscopy (KPFM) should be a key tool for characterizing the device physics of nanoscale electronics because it can directly image electrostatic potentials. In practice, though, distant connective electrodes interfere with accurate KPFM potential measurements and compromise its applicability. A parameterized KPFM technique described here determines these influences empirically during imaging, so that accurate potential profiles can be deduced from arbitrary device geometries without additional modeling. The technique is demonstrated on current-carrying single-walled carbon nanotubes (SWNTs), directly resolving average resistances per unit length of 70 kΩ/μm in semimetallic SWNTs and 200 kΩ/μm in semiconducting SWNTs. © 2013 American Institute of Physics.

Taq DNA polymerase functions at elevated temperatures with fast conformational dynamics-regimes previously inaccessible to mechanistic, single-molecule studies. Here, single-walled carbon nanotube transistors recorded the motions of Taq molecules processing matched or mismatched template-deoxynucleotide triphosphate pairs from 22° to 85°C. By using four enzyme orientations, the whole-enzyme closures of nucleotide incorporations were distinguished from more rapid, 20-μs closures of Taq's fingers domain testing complementarity and orientation. On average, one transient closure was observed for every nucleotide binding event; even complementary substrate pairs averaged five transient closures between each catalytic incorporation at 72°C. The rate and duration of the transient closures and the catalytic events had almost no temperature dependence, leaving all of Taq's temperature sensitivity to its rate-determining open state.

Simple and fast methods for the detection of target genes with single-nucleotide specificity could open up genetic research and diagnostics beyond laboratory settings. We recently reported a biosensor for the electronic detection of unamplified target genes using liquid-gated graphene field-effect transistors employing an RNA-guided catalytically deactivated CRISPR-associated protein 9 (Cas9) anchored to a graphene monolayer. Here, using unamplified genomic samples from patients and by measuring multiple types of electrical response, we show that the biosensors can discriminate within one hour between wild-type and homozygous mutant alleles differing by a single nucleotide. We also show that biosensors using a guide RNA-Cas9 orthologue complex targeting genes within the protospacer-adjacent motif discriminated between homozygous and heterozygous DNA samples from patients with sickle cell disease, and that the biosensors can also be used to rapidly screen for guide RNA-Cas9 complexes that maximize gene-targeting efficiency., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Single-molecule measurements of protein dynamics help unveil the complex conformational changes and transitions that occur during ligand binding and catalytic processes. Using high-resolution single-molecule nanocircuit techniques, we have investigated differences in the conformational dynamics and transitions of lysozyme interacting with three ligands: peptidoglycan substrate, substrate-based chitin analogue, and indole derivative inhibitors. While processing peptidoglycan, lysozyme followed one of the two mechanistic pathways for the hydrolysis of the glycosidic bonds: a concerted mechanism inducing direct conformational changes from open to fully closed conformations or a nonconcerted mechanism involving transient pauses in intermediate conformations between the open and closed conformations. In the presence of either chitin or an indole inhibitor, lysozyme was unable to access the fully closed conformation where catalysis occurs. Instead, lysozymes' conformational closures terminated at slightly closed, "excited" conformations that were approximately one-quarter of the full hinge-bending range. With the indole inhibitor, lysozyme reached this excited conformation in a single step without any evidence of rate-liming intermediates, but the same conformational motions with chitin involved three hidden, intermediate processes and features similar to the nonconcerted peptidoglycan mechanism. The similarities suggest that these hidden processes involve attempts to accommodate imperfectly aligned polysaccharides in the active site. The results provide a detailed glimpse of the enzyme-ligand interplay at the crux of molecular recognition, enzyme specificity, and catalysis.

Purpose: We introduce an automated, quantitative image analysis package for gamma camera and single photon emission computed tomography quality control. Our focus was to produce consistent methods that are feasible in clinical settings and use standard phantoms., Methods: Four gamma cameras were used to acquire planar images of four-quadrant bar phantoms and projection views of an American College of Radiology (ACR) phantom as part of a standard gamma camera quality control program. Images were sent to QC-Track® (Atirix Medical Systems, Inc., Minneapolis, MN, USA), which automatically placed predetermined regions of interest (ROIs) and performed analysis. For the bar phantom, a standard deviation (SD)-based modulation transfer function was calculated for a circular ROI in each quadrant. The bar widths at various MTF values were reported using linear interpolation as applicable. For the ACR phantom, the contrast-to-noise ratio (CNR) for each sphere, a modulation for each rods section, and a percent deviation for uniformity ROIs was calculated. Spheres corresponding to a CNR of 3, and the rod size at various modulations were also reported using linear interpolation. Visual analysis was performed by three medical physicists to evaluate interobserver variability and correlation to quantitative values., Results: Analysis of the bar phantom showed predictable differences with changes in matrix size and bar width and showed consistency over similar acquisitions over the course of the study. Analysis of the ACR Phantom showed increasing CNR and modulation with increasing sphere and rod diameter, as expected. For both phantoms, quantitative values from linear interpolation correlated well with visual analysis., Conclusion: Our automated method for quantitative image analysis is consistent and shows increased precision and sensitivity when compared to standard visual methods. Thresholds correspond well with visual analysis and previous guidelines for observer visibility (e.g., Rose criterion), making our framework suitable for routine use in a nuclear medicine department., (© 2020 The Authors. Journal of Applied Clinical Medical Physics published by Wiley Periodicals LLC on behalf of American Association of Physicists in Medicine.)

Advances in bioconjugation, the ability to link biomolecules to each other, small molecules, surfaces, and more, can spur the development of advanced materials and therapeutics. We have discovered that pyrocinchonimide, the dimethylated analogue of maleimide, undergoes a surprising transformation with biomolecules. The reaction targets amines and involves an imide transfer, which has not been previously reported for bioconjugation purposes. Despite their similarity to maleimides, pyrocinchonimides do not react with free thiols. Though both lysine residues and the N -termini of proteins can receive the transferred imide, the reaction also exhibits a marked preference for certain amines that cannot solely be ascribed to solvent accessibility. This property is peculiar among amine-targeting reactions and can reduce combinatorial diversity when many available reactive amines are available, such as in the formation of antibody-drug conjugates. Unlike amides, the modification undergoes very slow reversion under high pH conditions. The reaction offers a thermodynamically controlled route to single or multiple modifications of proteins for a wide range of applications.

As biosensing devices shrink smaller and smaller, they approach a scale in which single molecule electronic sensing becomes possible. Here, we review the operation of single-enzyme transistors made using single-walled carbon nanotubes. These novel hybrid devices transduce the motions and catalytic activity of a single protein into an electronic signal for real-time monitoring of the protein's activity. Analysis of these electronic signals reveals new insights into enzyme function and proves the electronic technique to be complementary to other single-molecule methods based on fluorescence. As one example of the nanocircuit technique, we have studied the Klenow Fragment (KF) of DNA polymerase I as it catalytically processes single-stranded DNA templates. The fidelity of DNA polymerases makes them a key component in many DNA sequencing techniques, and here we demonstrate that KF nanocircuits readily resolve DNA polymerization with single-base sensitivity. Consequently, template lengths can be directly counted from electronic recordings of KF's base-by-base activity. After measuring as few as 20 copies, the template length can be determined with <1 base pair resolution, and different template lengths can be identified and enumerated in solutions containing template mixtures., Competing Interests: The authors declare no conflict of interest.

A single point defect surrounded on either side by quasi-ballistic, semimetallic carbon nanotube is a nearly ideal system for investigating disorder in one-dimensional (1D) conductors and comparing experiment to theory. Here, individual single-walled nanotubes (SWNTs) are investigated before and after the incorporation of single point defects. Transport and local Kelvin Probe force microscopy independently demonstrate high-resistance depletion regions over 1.0 μm wide surrounding one point defect in semimetallic SWNTs. Transport measurements show that conductance through such wide depletion regions occurs via a modified, 1D version of Poole-Frenkel field-assisted emission. Given the breadth of theory dedicated to the possible effects of disorder in 1D systems, it is surprising that a Poole-Frenkel mechanism appears to describe defect scattering and resistance in this semimetallic system.

DNA polymerases exhibit a surprising tolerance for analogs of deoxyribonucleoside triphosphates (dNTPs), despite the enzymes' highly evolved mechanisms for the specific recognition and discrimination of native dNTPs. Here, individual DNA polymerase I Klenow fragment (KF) molecules were tethered to a single-walled carbon nanotube field-effect transistor (SWCNT-FET) to investigate accommodation of dNTP analogs with single-molecule resolution. Each base incorporation accompanied a change in current with its duration defined by τclosed. Under Vmax conditions, the average time of τclosed was similar for all analog and native dNTPs (0.2 to 0.4 ms), indicating no kinetic impact on this step due to analog structure. Accordingly, the average rates of dNTP analog incorporation were largely determined by durations with no change in current defined by τopen, which includes molecular recognition of the incoming dNTP. All α-thio-dNTPs were incorporated more slowly, at 40 to 65% of the rate for the corresponding native dNTPs. During polymerization with 6-Cl-2APTP, 2-thio-dTTP, or 2-thio-dCTP, the nanocircuit uncovered an alternative conformation represented by positive current excursions that does not occur with native dNTPs. A model consistent with these results invokes rotations by the enzyme's O-helix; this motion can test the stability of nascent base pairs using nonhydrophilic interactions and is allosterically coupled to charged residues near the site of SWCNT attachment. This model with two opposing O-helix motions differs from the previous report in which all current excursions were solely attributed to global enzyme closure and covalent-bond formation. The results suggest the enzyme applies a dynamic stability-checking mechanism for each nascent base pair.

Single-molecule techniques can monitor the kinetics of transitions between enzyme open and closed conformations, but such methods usually lack the resolution to observe the underlying transition pathway or intermediate conformational dynamics. We have used a 1 MHz bandwidth carbon nanotube transistor to electronically monitor single molecules of the enzyme T4 lysozyme as it processes substrate. An experimental resolution of 2 μs allowed the direct recording of lysozyme's opening and closing transitions. Unexpectedly, both motions required 37 μs, on average. The distribution of transition durations was also independent of the enzyme's state: either catalytic or nonproductive. The observation of smooth, continuous transitions suggests a concerted mechanism for glycoside hydrolysis with lysozyme's two domains closing upon the polysaccharide substrate in its active site. We distinguish these smooth motions from a nonconcerted mechanism, observed in approximately 10% of lysozyme openings and closings, in which the enzyme pauses for an additional 40-140 μs in an intermediate, partially closed conformation. During intermediate forming events, the number of rate-limiting steps observed increases to four, consistent with four steps required in the stepwise, arrow-pushing mechanism. The formation of such intermediate conformations was again independent of the enzyme's state. Taken together, the results suggest lysozyme operates as a Brownian motor. In this model, the enzyme traces a single pathway for closing and the reverse pathway for enzyme opening, regardless of its instantaneous catalytic productivity. The observed symmetry in enzyme opening and closing thus suggests that substrate translocation occurs while the enzyme is closed.

Using a model system of single, isolated carbon nanotubes loaded with high-capacitance metal-oxide films, we have quantitatively investigated electrochemical composites on the single-nanotube scale. Electrochemical charging and discharging of a model MnO2 storage material was used to probe interfacial charge transfer and surface impedances at the nanotube interface. We found that one single-walled carbon nanotube has an apparent surface resistivity of 30 mΩ cm(2), approximately 4 times smaller than for a multiwalled carbon nanotube and 50 times smaller than the 1.5 Ω cm(2) resistivity of Pt or graphite films. The improvement originates in the electrochemical-transport properties of microelectrodes shrunk to a nanotube's dimensions rather than any unique nanotube property like curvature, bandstructure, or surface chemistry. In explaining the enhanced performance of certain nanotube-containing composites, the results overturn widely held assumptions about nanotubes' roles while also providing guidelines for optimizing effective composites.

Single molecule bioelectronic circuits provide an opportunity to study chemical kinetics and kinetic variability with bond-by-bond resolution. To demonstrate this approach, we examined the catalytic activity of T4 lysozyme processing peptidoglycan substrates. Monitoring a single lysozyme molecule through changes in a circuit's conductance helped elucidate unexplored and previously invisible aspects of lysozyme's catalytic mechanism and demonstrated lysozyme to be a processive enzyme governed by 9 independent time constants. The variation of each time constant with pH or substrate crosslinking provided different insights into catalytic activity and dynamic disorder. Overall, ten lysozyme variants were synthesized and tested in single molecule circuits to dissect the transduction of chemical activity into electronic signals. Measurements show that a single amino acid with the appropriate properties is sufficient for good signal generation, proving that the single molecule circuit technique can be easily extended to other proteins.

Bioconjugating single molecules of the Klenow fragment of DNA polymerase I into electronic nanocircuits allowed electrical recordings of enzymatic function and dynamic variability with the resolution of individual nucleotide incorporation events. Continuous recordings of DNA polymerase processing multiple homopolymeric DNA templates extended over 600 s and through >10,000 bond-forming events. An enzymatic processivity of 42 nucleotides for a template of the same length was directly observed. Statistical analysis determined key kinetic parameters for the enzyme's open and closed conformations. Consistent with these nanocircuit-based observations, the enzyme's closed complex forms a phosphodiester bond in a highly efficient process >99.8% of the time, with a mean duration of only 0.3 ms for all four dNTPs. The rate-limiting step for catalysis occurs during the enzyme's open state, but with a nearly 2-fold longer duration for dATP or dTTP incorporation than for dCTP or dGTP into complementary, homopolymeric DNA templates. Taken together, the results provide a wealth of new information complementing prior work on the mechanism and dynamics of DNA polymerase I.

Single-molecule studies of enzymes open a window into their dynamics and kinetics. A single molecule of the catalytic domain of cAMP-dependent protein kinase A (PKA) was attached to a single-walled carbon nanotube device for long-duration monitoring. The electronic recording clearly resolves substrate binding, ATP binding, and cooperative formation of PKA's catalytically functional, ternary complex. Using recordings of a single PKA molecule extending over 10 min and tens of thousands of binding events, we determine the full transition probability matrix and conversion rates governing formation of the apo, intermediate, and closed enzyme configurations. We also observe kinetic rates varying over 2 orders of magnitude from one second to another. Anti-correlation of the on and off rates for PKA binding to the peptide substrate, but not ATP, demonstrates that regulation of enzyme activity results from altering the stability of the PKA-substrate complex, not its binding to ATP. The results depict a highly dynamic enzyme offering dramatic possibilities for regulated activity, an attribute useful for an enzyme with crucial roles in cell signaling.

Single-molecule experimental methods have provided new insights into biomolecular function, dynamic disorder, and transient states that are all invisible to conventional measurements. A novel, nonfluorescent single-molecule technique involves attaching single molecules to single-walled carbon nanotube field-effective transistors (SWNT FETs). These ultrasensitive electronic devices provide long-duration, label-free monitoring of biomolecules and their dynamic motions. However, generalization of the SWNT FET technique first requires design rules that can predict the success and applicability of these devices. Here, we report on the transduction mechanism linking enzymatic processivity to electrical signal generation by a SWNT FET. The interaction between SWNT FETs and the enzyme lysozyme was systematically dissected using eight different lysozyme variants synthesized by protein engineering. The data prove that effective signal generation can be accomplished using a single charged amino acid, when appropriately located, providing a foundation to widely apply SWNT FET sensitivity to other biomolecular systems.

The dynamic processivity of individual T4 lysozyme molecules was monitored in the presence of either linear or cross-linked peptidoglycan substrates. Single-molecule monitoring was accomplished using a novel electronic technique in which lysozyme molecules were tethered to single-walled carbon nanotube field-effect transistors through pyrene linker molecules. The substrate-driven hinge-bending motions of lysozyme induced dynamic electronic signals in the underlying transistor, allowing long-term monitoring of the same molecule without the limitations of optical quenching or bleaching. For both substrates, lysozyme exhibited processive low turnover rates of 20-50 s(-1) and rapid (200-400 s(-1)) nonproductive motions. The latter nonproductive binding events occupied 43% of the enzyme's time in the presence of the cross-linked peptidoglycan but only 7% with the linear substrate. Furthermore, lysozyme catalyzed the hydrolysis of glycosidic bonds to the end of the linear substrate but appeared to sidestep the peptide cross-links to zigzag through the wild-type substrate., (© 2012 American Chemical Society)

Tethering a single lysozyme molecule to a carbon nanotube field-effect transistor produced a stable, high-bandwidth transducer for protein motion. Electronic monitoring during 10-minute periods extended well beyond the limitations of fluorescence techniques to uncover dynamic disorder within a single molecule and establish lysozyme as a processive enzyme. On average, 100 chemical bonds are processively hydrolyzed, at 15-hertz rates, before lysozyme returns to its nonproductive, 330-hertz hinge motion. Statistical analysis differentiated single-step hinge closure from enzyme opening, which requires two steps. Seven independent time scales governing lysozyme's activity were observed. The pH dependence of lysozyme activity arises not from changes to its processive kinetics but rather from increasing time spent in either nonproductive rapid motions or an inactive, closed conformation.

A variation of scanning gate microscopy (SGM) is demonstrated in which this imaging mode is extended into an electrostatic spectroscopy. Continuous variation of the SGM probe's electrostatic potential is used to directly resolve the energy spectrum of localized electronic scattering in functioning, molecular scale devices. The technique is applied to the energy-dependent carrier scattering that occurs at defect sites in carbon nanotube transistors, and fitting energy-resolved experimental data to a simple transmission model determines the electronic character of each defect site. For example, a phenolic type of covalent defect is revealed to produce a tunnel barrier 0.1 eV high and 0.5 nm wide.

Individual single-walled carbon nanotubes (SWCNTs) become sensitive to H(2) gas when their surfaces are decorated with Pd metal, and previous reports measure typical chemoresistive increases to be approximately 2-fold. Here, thousand-fold increases in resistance are demonstrated in the specific case where a Pd cluster decorates a SWCNT sidewall defect site. Measurements on single SWCNTs, performed both before and after defect incorporation, prove that defects have extraordinary consequences on the chemoresistive response, especially in the case of SWCNTs with metallic band structure. Undecorated defects do not contribute to H(2) chemosensitivity, indicating that this amplification is due to a specific but complex interdependence between a defect site's electronic transmission and the chemistry of the defect-Pd-H(2) system. Dosage experiments suggest a primary role is played by spillover of atomic H onto the defect site.

Motivated by recent experiments, we investigate how NO3-SWNT interactions become energetically favorable with varying oxidation state of a single-walled carbon nanotube (SWNT) using first-principles calculations. Chemisorption becomes less endothermic with respect to physisorption when the SWNT oxidation state is elevated. Importantly, the dissociative incorporation of an oxygen atom into the SWNT sidewall becomes highly favorable when the SWNT oxidation state is elevated from electron density depletion in the vicinity, as caused experimentally using electrochemical potential. The elevation of the SWNT oxidation state through accumulating local charge transfer from the surrounding molecules does not have the same effect. Our investigation reveals the crucial effects of the SWNT oxidation state in understanding the molecule-SWNT interaction.

We investigate electronic devices consisting of individual, metallic, single-walled carbon nanotubes contacted by Pt electrodes in a field effect transistor configuration, focusing on improvements to the metal-nanotube contact resistance as the devices are annealed in inert environments including ultrahigh vacuum. At moderate temperatures (T < 880 K), thermal processing results in high resistance contacts with thermally activated barriers. Higher temperatures (T > 880 K) achieve nearly transparent contacts. In the latter case, analytical surface measurements reveal the catalytic decomposition of hydrocarbons into graphene layers on the Pt surface, suggesting that improved electronic behavior is primarily due to the formation of an all-carbon nanotube-graphite interface rather than to the improvement of the nanotube-Pt one.

Dual color four-wave-mixing (FWM) microscopy is used to spatially resolve the third-order optical response from individual carbon nanotubes. Good signal-to-noise is obtained from single-walled carbon nanotubes (SWNT) sitting on substrates, when the excitation beams are resonant with electronic transitions of the nanotube, by detecting the FWM response at the anti-Stokes frequency. Whereas the coherent anti-Stokes (CAS) signal is sensitive to both electronic and vibrational resonances of the material, it is shown that the signal from individual SWNTs is dominated by the electronic response. The CAS signal is strongly polarization dependent, with the highest signals found parallel with the enhanced electronic polarizibility along the long axis of the SWNT.

Real-time monitoring of carbon nanotube conductance during electrochemical and chemical etching reveals the electronic signatures of individual bond alteration events on the nanotube sidewall. Tracking the conductance of multiple single-molecule experiments through different synthetic protocols supports putative mechanisms for sidewall derivatization. Insights gained from these mechanistic observations imply the formation of sidewall carboxylates, which are useful as handles for bioconjugation. We describe an electronic state required for efficacious chemical treatment. Such real-time monitoring can improve carboxylate yields to 45 % or more. The experiments illustrate the power of molecular nanocircuits to uncover and direct the mechanisms of chemical reactions.

Using point-functionalized carbon nanotube devices, we demonstrate continuous, multihour monitoring of a single carboxylate group interacting with its immediate environment. The conductance of the nanotube device directly transduces single-molecule attachments and detachments in the presence of the carboxylate-selective reagent 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Because only one carboxylate is present in the device, it can be studied through hundreds of reactions, providing the statistical accuracy to directly determine a 12 s lifetime of the carboxy-EDC complex. An additional instability of the complex is transduced in real time and observed to have a median lifetime of 2 ms. By determining a turnover time in good agreement with bulk measurements and simultaneously illuminating additional dynamics, these results demonstrate this platform's potential for complementing optical methods in single-molecule research.

We used covalent attachments to single-walled carbon nanotubes (SWNTs) to fabricate single-molecule electronic devices. The technique does not rely on submicrometer lithography or precision mechanical manipulation, but instead uses circuit conductance to monitor and control covalent attachment to an electrically connected SWNT. Discrete changes in the circuit conductance revealed chemical processes happening in real time and allowed the SWNT sidewalls to be deterministically broken, reformed, and conjugated to target species. By controlling the chemistry through electronically controlled electrochemical potentials, we were able to achieve single chemical attachments. We routinely functionalized pristine, defect-free SWNTs at one, two, or more sites and demonstrated three-terminal devices in which a single attachment controls the electronic response.

The chemical reactivity of carbon nanotubes in H2SO4 is investigated using individual, single-walled carbon nanotubes (SWNTs) incorporated into electronic devices. Exploiting the device conductance as a sensitive indicator of chemical reactions, discrete oxidation and reduction events can be clearly observed. During oxidation, a SWNT opens circuits to a nanometer-scale tunnel junction with residual conduction similar to Frenkel-Poole charge emission. When electrochemically reduced, a SWNT returns to its original conductance. This redox cycle can be repeated many times, suggesting a novel chemical method of reversibly switching SWNT conductivity.

The temperature-dependent noise of individual, single-walled carbon nanotubes is measured here in a variety of different gases and liquids. The ambient environment is found to have only a weak relationship with device noise, even in cases where adsorption significantly changes the dc resistance. Correspondingly, a 450 K degassing procedure typically reduces the device noise by only 1 order of magnitude. An important exception to this finding is a pronounced, 100-fold increase in noise observed near gas-liquid phase transitions of the ambient. Wide-range temperature scans clearly identify the condensation of N(2), H(2), and CH(4) onto metallic nanotubes, but not the sublimation of CO(2). The observations suggest that nanotube devices can directly transduce ambient density fluctuations, though without an inherent gas specificity. Even so, the method is a particularly sensitive characterization of nanotube chemical interactions, one which is successful even for the extreme case of inert gases adsorbed on metallic nanotubes.

The prevailing conception of carbon nanotubes and particularly single-walled carbon nanotubes (SWNTs) continues to be one of perfectly crystalline wires. Here, we demonstrate a selective electrochemical method that labels point defects and makes them easily visible for quantitative analysis. High-quality SWNTs are confirmed to contain one defect per 4 microm on average, with a distribution weighted towards areas of SWNT curvature. Although this defect density compares favourably to high-quality, silicon single-crystals, the presence of a single defect can have tremendous electronic effects in one-dimensional conductors such as SWNTs. We demonstrate a one-to-one correspondence between chemically active point defects and sites of local electronic sensitivity in SWNT circuits, confirming the expectation that individual defects may be critical to understanding and controlling variability, noise and chemical sensitivity in SWNT electronic devices. By varying the SWNT synthesis technique, we further show that the defect spacing can be varied over orders of magnitude. The ability to detect and analyse point defects, especially at very low concentrations, indicates the promise of this technique for quantitative process analysis, especially in nanoelectronics development.

Carbon nanotubes display either metallic or semiconducting properties. Both large, multiwalled nanotubes (MWNTs), with many concentric carbon shells, and bundles or "ropes" of aligned single-walled nanotubes (SWNTs), are complex composite conductors that incorporate many weakly coupled nanotubes that each have a different electronic structure. Here we demonstrate a simple and reliable method for selectively removing single carbon shells from MWNTs and SWNT ropes to tailor the properties of these composite nanotubes. We can remove shells of MWNTs stepwise and individually characterize the different shells. By choosing among the shells, we can convert a MWNT into either a metallic or a semiconducting conductor, as well as directly address the issue of multiple-shell transport. With SWNT ropes, similar selectivity allows us to generate entire arrays of nanoscale field-effect transistors based solely on the fraction of semiconducting SWNTs.

We investigate the limits of high energy transport in multiwalled carbon nanotubes (MWNTs). In contrast to metal wires, MWNTs do not fail in the continuous, accelerating manner typical of electromigration. Instead, they fail via a series of sharp, equally sized current steps. We assign these steps to the sequential destruction of individual nanotube shells, consistent with the MWNT's concentric-shell geometry. Furthermore, the initiation of this failure is very sensitive to air exposure. In air failure is initiated by oxidation at a particular power, whereas in vacuum MWNTs can withstand much higher power densities and reach their full current carrying capacities.

The thermoelectric power (TEP) of single-walled carbon nanotubes (SWNTs) is extremely sensitive to gas exposure history. Samples exposed to air or oxygen have an always positive TEP, suggestive of holelike carriers. However, at fixed temperature the TEP crosses zero and becomes progressively more negative as the SWNTs are stripped of oxygen. The time constant for oxygen adsorption/desorption is strongly temperature dependent and ranges from seconds to many days, leading to apparently "variable" TEP for a given sample at a given temperature. The saturated TEP can be accounted for within a model of strong oxygen doping of the semiconducting nanotubes.

The electronic properties of single-walled carbon nanotubes are shown here to be extremely sensitive to the chemical environment. Exposure to air or oxygen dramatically influences the nanotubes' electrical resistance, thermoelectric power, and local density of states, as determined by transport measurements and scanning tunneling spectroscopy. These electronic parameters can be reversibly "tuned" by surprisingly small concentrations of adsorbed gases, and an apparently semiconducting nanotube can be converted into an apparent metal through such exposure. These results, although demonstrating that nanotubes could find use as sensitive chemical gas sensors, likewise indicate that many supposedly intrinsic properties measured on as-prepared nanotubes may be severely compromised by extrinsic air exposure effects.

Food allergy presents a problem for many parts of society, including sensitive subjects, schools, health authorities, and the food industry. Once food allergy is diagnosed, dietary avoidance is the principle method of management. Because trace levels of peanuts can elicit an adverse to fatal reaction, unintentional exposure to the offending allergens may have devastating consequences to sensitive individuals. However, determination of trace amounts of unintentional peanut contamination in our food supply is very difficult. Recently, we developed polyclonal antibodies specific to peanut proteins that do not cross-react with 22 legumes, tree nuts, or other common snack ingredients. An antiserum containing the polyclonal antibodies was used to develop a sensitive enzyme-linked immunosorbent assay (ELISA) for determination of peanut proteins in snack foods. This study reports the first successful ELISA test to detect trace amounts of peanut allergens in a variety of foods. The concentration of peanut protein that inhibited 50% of antibody-antigen binding (IC50) was 12 ng/mL, the linear range was 1-63 ng/mL, and the detection limit was 400 ng/g (ppb) for the various foods tested. Recoveries ranged from 68 to 90%, with coefficients of variation of 2-22%, depending on the commodity. Using this new procedure, allergy-related complaint samples from various food groups were analyzed, and undeclared peanut proteins were identified in some products.

Oligosaccharyltransferase catalyzes the transfer of a preassembled high mannose oligosaccharide from a dolichol-oligosaccharide donor to consensus glycosylation acceptor sites in newly synthesized proteins in the lumen of the rough endoplasmic reticulum. The Saccharomyces cerevisiae oligosaccharyltransferase is an oligomeric complex composed of six non-identical subunits (alpha-zeta). The alpha, beta, gamma, and delta subunits of the oligosaccharyltransferase are encoded by the OST1, WBP1, OST3, and SWP1 genes, respectively. Here we describe the functional characterization of the OST2 gene that encodes the epsilon-subunit of the oligosaccharyltransferase. Genomic disruption of the OST2 locus was lethal in haploid yeast showing that expression of the Ost2 protein is essential for viability. Overexpression of the Ost2 protein suppresses the temperature-sensitive phenotype of the wbp1-2 allele and increases in vivo and in vitro oligosaccharyltransferase activity in a wbp1-2 strain. An analysis of a series of conditional ost2 mutants demonstrated that defects in the Ost2 protein cause pleiotropic underglycosylation of soluble and membrane-bound glycoproteins. Microsomal membranes isolated from ost2 mutant yeast show marked reductions in the in vitro transfer of high mannose oligosaccharide from exogenous lipid-linked oligosaccharide to a glycosylation site acceptor tripeptide. Surprisingly, the Ost2 protein was found to be 40% identical to the DAD1 protein (defender against apoptotic cell death), a highly conserved protein initially identified in vertebrate organisms. The protein sequence of ost2 mutant alleles revealed mutations at highly conserved residues in the Ost2p/DAD1 protein sequence.

Oligosaccharyltransferase mediates the transfer of a preassembled high mannose oligosaccharide from a lipid-linked oligosaccharide donor to consensus glycosylation acceptor sites in newly synthesized proteins in the lumen of the rough endoplasmic reticulum. The Saccharomyces cerevisiae oligosaccharyltransferase is an oligomeric complex composed of six nonidentical subunits (alpha-zeta), two of which are glycoproteins (alpha and beta). The beta and delta subunits of the oligosaccharyltransferase are encoded by the WBP1 and SWP1 genes. Here we describe the functional characterization of the OST1 gene that encodes the alpha subunit of the oligosaccharyltransferase. Protein sequence analysis revealed a significant sequence identity between the Saccharomyces cerevisiae Ost1 protein and ribophorin I, a previously identified subunit of the mammalian oligosaccharyltransferase. A disruption of the OST1 locus was not tolerated in haploid yeast showing that expression of the Ost1 protein is essential for vegetative growth of yeast. An analysis of a series of conditional ost1 mutants demonstrated that defects in the Ost1 protein cause pleiotropic underglycosylation of soluble and membrane-bound glycoproteins at both the permissive and restrictive growth temperatures. Microsomal membranes isolated from ost1 mutant yeast showed marked reductions in the in vitro transfer of high mannose oligosaccharide from exogenous lipid-linked oligosaccharide to a glycosylation site acceptor tripeptide. Microsomal membranes isolated from the ost1 mutants contained elevated amounts of the Kar2 stress-response protein.

A simple, sensitive, and precise enzyme-linked immunosorbent assay (ELISA) is described for the quantitation of captan as its degradation product tetrahydrophthalimide (THPI) in foods using polyclonal antibodies. Three hapten analogues of THPI with different alkyl spacer arm lengths were synthesized. Immunogens and coating proteins were prepared by coupling these haptens to human serum albumin and ovalbumin, respectively. A 5-carbon spacer arm appeared to be optimum for the production of antibodies. Heterologous coating proteins did not improve the sensitivity, but reduction of homologous coating protein concentration did improve the sensitivity, resulting in a concentration of test compound required to inhibit binding by 50% of 15.5 ng/mL. The antiserum is specific for captan, captafol, and THPI, but not other structurally related compounds. The minimum detection limit was 1 ng/mL; the linearity was 1-200 ng/mL. The overall recoveries of captan and THPI from 11 commodities spiked at 4 levels were 92 and 100%, respectively. The intra-assay and interassay coefficients of variation were 9.1 and 16.8% for apple blanks and 5.9 and 4.2% for apple spiked with 3 ppm THPI, respectively. The ELISA described is suitable for measuring captan and THPI at levels comparable to those typically found in fruit.

Retrorectal masses are rare and of insidious onset. We report a consecutive series of six such cases (males = 4, females = 2). The main presenting complaint was back pain and the most reliable physical sign was a palpable mass posteriorly on rectal examination (all cases). C T scan was the most radiologically informative investigation. Surgical intervention was undertaken using both anterior (trans-abdominal) and posterior (retrorectal) approaches. The majority of the masses excised were benign and all patients, to date, remain well.

We have used the membrane-impermeable, thiol-cleavable, crosslinker 3,3'-dithio bis (sulfosuccinimidylpropionate) to identify proteins that are in the vicinity of membrane-bound ribosomes of the RER. A specific subset of RER proteins was reproducibly crosslinked to the ribosome. Immunoblot analysis of the crosslinked products with antibodies raised against signal recognition particle receptor, ribophorin I, and the 35-kD subunit of the signal sequence receptor demonstrated that these translocation components had been crosslinked to the ribosome, but each to a different extent. The most prominent polypeptide among the crosslinked products was a 180-kD protein that has recently been proposed to be a ribosome receptor (Savitz, A.J., and D.I. Meyer, 1990. Nature (Lond.). 346: 540-544). RER membrane proteins were reconstituted into liposomes and assayed with radiolabeled ribosomes to determine whether ribosome binding activity could be ascribed to the 180-kD protein. Differential detergent extraction was used to prepare soluble extracts of microsomal membrane vesicles that either contained or lacked the 180-kD protein. Liposomes reconstituted from both extracts bound ribosomes with essentially identical affinity. Additional fractionation experiments demonstrated that the bulk of the ribosome binding activity present in detergent extracts of microsomal membranes could be readily resolved from the 180-kD protein by size exclusion chromatography. Taken together, we conclude that the 180-kD protein is in the vicinity of membrane bound ribosomes, yet does not correspond to the ribosome receptor.

Optimal treatment for unresectable carcinoma of the pancreas remains controversial. This study was done to examine the relationship between perioperative jaundice and postoperative morbidity, and type of palliative biliary bypass and postoperative morbidity and jaundice clearance. Seventy-six patients with obstructive jaundice secondary to carcinoma of the head of the pancreas were studied. Forty-nine patients underwent one of four different types of palliative bypass: 1, cholecystojejunostomy (n = 22); 2, choledochojejunostomy (n = 11); 3, choledochoduodenostomy (n = 9), and 4, cholecystoduodenostomy (n = 7). Age, sex and preoperative health status were similar for all operative groups, as well as for those with and without postoperative morbidity. The postoperative complication rate was 33 per cent and there was one postoperative death. Length of preoperative jaundice and peak preoperative bilirubin levels were independent of morbidity. Postoperative morbidity was similar for each type of bypass used and no significant difference was found when cholecystoenteric (1 and 4) and choledochoenteric (2 and 3) bypass were compared. The results of this study support the view that postoperative morbidity is not directly related to the presence of jaundice preoperatively. Furthermore, the rate of jaundice clearance and the occurrence of postoperative complications are not dependent on the type of bypass used.

A variety of low molecular weight aliphatic chlorinated hydrocarbons have a history of use in the processing of foods, and their presence in potable water supplies has also been demonstrated. In this study, the relative uptake of four different aliphatic chlorinated hydrocarbons (methylene chloride, dichloroethane, chloroform and trichloroethylene) was studied after administration, by intragastric intubation, at the same dose level as corn-oil or aqueous solutions. Serial blood samples were collected over a 5-h period and uptake was assessed as the calculated area under blood concentration-time curves. A significant decrease in the rate and extent of uptake was observed for the compounds when administered as an oil solution as compared with an aqueous solution. Lower peak concentrations and an increase in the time taken to reach peak concentration, after dosing with corn oil solutions, was also observed.