DNA-Modified Electrodes Fabricated Using Copper-Free Click Chemistry for Enhanced Protein Detection

A method of DNA monolayer formation has been developed using copper-free click chemistry that yields enhanced surface homogeneity and enables variation in the amount of DNA assembled; extremely low-density DNA monolayers, with as little as 5% of the monolayer being DNA, have been formed. These DNA-modified electrodes (DMEs) were characterized visually, with AFM, and electrochemically, and were found to facilitate DNA-mediated reduction of a distally bound redox probe. These low-density monolayers were found to be more homogeneous than traditional thiol-modified DNA monolayers, with greater helix accessibility through an increased surface area-to-volume ratio. Protein binding efficiency of the transcriptional activator TATA-binding protein (TBP) was also investigated on these surfaces and compared to that on DNA monolayers formed with standard thiol-modified DNA. Our low-density monolayers were found to be extremely sensitive to TBP binding, with a signal decrease in excess of 75% for 150 nM protein. This protein was detectable at 4 nM, on the order of its dissociation constant, with our low-density monolayers. The improved DNA helix accessibility and sensitivity of our low-density DNA monolayers to TBP binding reflects the general utility of this method of DNA monolayer formation for DNA-based electrochemical sensor development

A Multiplexed, Two-Electrode Platform for Biosensing Based on DNA-Mediated Charge Transport

We have developed a thin layer, multiplexed biosensing platform that features two working-electrode arrays for detecting small molecules, nucleic acid sequences, and DNA-binding proteins. DNA duplexes are patterned onto the primary electrode array, while a secondary electrode array is used both to initiate DNA monolayer formation and for electrochemical readout via DNA-mediated charge transport (DNA CT) chemistry. Electrochemical reduction of Cu(phendione)_2^(2+) (phendione is 1,10-phenanthroline-5,6-dione) at the secondary electrodes induces covalent attachment via click chemistry of ethynyl-labeled DNA probe duplexes onto the primary electrodes that have been treated with azide-terminated alkylthiols. Electrochemical impedance spectroscopy and cyclic voltammetry confirm that catalyst activation at the secondary electrode is essential to maintain the integrity of the DNA monolayer. Electrochemical readout of DNA CT processes that occur at the primary electrode is accomplished also at the secondary electrode. The two-electrode system enables the platform to function as a collector–generator using either ferrocyanide or ferricyanide as mediators with methylene blue and DNA charge transport. Electrochemical measurements at the secondary electrode eliminate the need for large background corrections. The resulting sensitivity of this platform enables the reliable and simultaneous detection of femtomoles of the transcription factors TATA-binding protein and CopG on a single multiplexed device

G328.4+0.2 : A large and luminous Crab-like supernova remnant

We report on radio continuum and HI observations of the radio source G328.4+0.2 using the Australia Telescope Compact Array. Our results confirm G328.4+0.2 to be a filled-center nebula with no surrounding shell, showing significant linear polarization and an almost flat spectral index. These results lead us to conclude that G328.4+0.2 is a Crab-like, or ``plerionic'', supernova remnant (SNR), presumably powered by an unseen central pulsar. HI absorption towards G328.4+0.2 puts a lower limit on its distance of 17.4 +/- 0.9 kpc, making it the largest (D=25 pc) and most luminous (L_R = 3e35 erg/s) Crab-like SNR in the Galaxy. We infer G328.4+0.2 to be significantly older than the Crab Nebula, but powered by a pulsar which is fast spinning (P<20 ms) and which has a comparatively low magnetic field (B<1e12 G). We propose G328.4+0.2, G74.9+1.2 and N157B as a distinct group of large-diameter, high-luminosity Crab-like SNRs, all powered by fast-spinning low-field pulsars.Comment: 7 pages, 3 embedded EPS figures, uses emulateapj.sty. Accepted to ApJ. Abstract corrected so that distance is now in kpc, not pc

Label-free electrochemical detection of human methyltransferase from tumors

The role of abnormal DNA methyltransferase activity in the development and progression of cancer is an essential and rapidly growing area of research, both for improved diagnosis and treatment. However, current technologies for the assessment of methyltransferase activity, particularly from crude tumor samples, limit this work because they rely on radioactivity or fluorescence and require bulky instrumentation. Here, we report an electrochemical platform that overcomes these limitations for the label-free detection of human DNA(cytosine-5)-methyltransferase1 (DNMT1) methyltransferase activity, enabling measurements from crude cultured colorectal cancer cell lysates (HCT116) and biopsied tumor tissues. Our multiplexed detection system involving patterning and detection from a secondary electrode array combines low-density DNA monolayer patterning and electrocatalytically amplified DNA charge transport chemistry to measure selectively and sensitively DNMT1 activity within these complex and congested cellular samples. Based on differences in DNMT1 activity measured with this assay, we distinguish colorectal tumor tissue from healthy adjacent tissue, illustrating the effectiveness of this two-electrode platform for clinical applications

Investigation of spin-orbit effects in the excitation of noble gases by spin-polarized electrons

We have measured the integrated Stokes parameters of the light emitted following the impact excitation by polarized electrons of the np5 (n+1)p [5/2] 3 states in Ne (n=2), Ar (n=3), Kr (n=4), Xe (n=5), and the np5 (n+1) p[5/2] 2 state in Kr. The near-threshold linear-polarization fractions η1 were consistent with zero for all of the J=3 states measured, providing no evidence of Mott scattering. For J=2 excitation, η1 shows the clear influence of the intermediately coupled nature of the state. At threshold, the measured circular polarization η2 and linear polarization η3 for the J=3 states agree well with the LS-coupled threshold predictions. These results provide the basis for optical measurements of electron polarization

Исследование влияния природы растворителя, используемого при получении электродной мембраны, на работу Ni-селективного электрода при определении ионов никеля (LL) в водных средах

Исследовано влияние природы растворителей (бензиловый спирт и хлорфенол), используемых для изготовления полимерной электродной мембраны твердофазного никельселективного электрода на основе никельгексационоферратов ({Ni2[Fe(CN)6]}), иммобилизованных в полиэтилентерефталатную (ПЭТФ) матрицу для определения ионов никеля в водных средах. Установлено, что мембраны, полученные на основе бензилового спирта имеют более высокие электродные показатели по сходимости и воспроизводимости потенциалов, стабильности работы, низком дрейфе и времени отклика электрода по сравнению с мембранами на основе хлорфенола и могут быть рекомендованы к использованию в никель селективных электродах.The influence of the nature of the solvents (benzyl alcohol and chlorophenol) used to fabricate a polymer electrode membrane of a solid-phase nickel selective electrode based on nickelhexacetanoferrates ({Ni2 [Fe (CN) 6]}) immobilized in a polyethylene terephthalate (PET) matrix for the determination of nickel ions in aqueous media . It has been established that membranes obtained on the basis of benzyl alcohol have higher electrode indices for the convergence and reproducibility of potentials, stability of operation, low drift and electrode response time compared to chlorophenol-based membranes, and can be recommended for use in nickel selective electrodes

X-Ray Observations of the supernova remnant G21.5-0.9

We present the analysis of archival X-ray observations of the supernova remnant (SNR) G21.5-0.9. Based on its morphology and spectral properties, G21.5-0.9 has been classified as a Crab-like SNR. In their early analysis of the CHANDRA calibration data, Slane et al. (2000) discovered a low-surface-brightness, extended emission. They interpreted this component as the blast wave formed in the supernova (SN) explosion. In this paper, we present the CHANDRA analysis using a total exposure of ~150 ksec. We also include ROSAT and ASCA observations. Our analysis indicates that the extended emission is non-thermal -- a result in agreement with XMM observations. The entire remnant of radius ~ 2'.5 is best fitted with a power law model with a photon index steepening away from the center. The total unabsorbed flux in the 0.5-10 keV is 1.1E-10 erg/cm2/s with an 85% contribution from the 40" radius inner core. Timing analysis of the High-Resolution Camera (HRC) data failed to detect any pulsations. We put a 16% upper limit on the pulsed fraction. We derive the physical parameters of the putative pulsar and compare them with those of other plerions (such as the Crab and 3C 58). G21.5-0.9 remains the only plerion whose size in X-rays is bigger than in the radio. Deep radio observations will address this puzzle.Comment: 23 pages including 11 figures and 3 tables; accepted by ApJ June 22, 2001; to appear in Oct 20, 2001 issue of Ap

Faint HI 21-cm Emission Line Wings at Forbidden-Velocities

We present the results of a search for faint HI 21-cm emission line wings at velocities forbidden by Galactic rotation in the Galactic plane using the Leiden/Dwingeloo HI Survey data and the HI Southern Galactic Plane Survey data. These ``forbidden-velocity wings (FVWs)'' appear as protruding excessive emission in comparison with their surroundings in limited (< 2 deg) spatial regions over velocity extent more than ~20 km/s in large-scale (l-v) diagrams. Their high-velocities imply that there should be some dynamical phenomena associated. We have identified 87 FVWs. We present their catalog, and discuss their distribution and statistical properties. We found that 85% of FVWs are not coincident with known supernova remnants (SNRs), galaxies, or high-velocity clouds. Their natures are currently unknown. We suspect that many of them are fast-moving HI shells and filaments associated with the oldest SNRs that are essentially invisible except via their HI line emission. We discuss other possible origins.Comment: 41 pages, 14 figures, to be published in apj