In situ electroporation of surface-bound siRNAs in microwell arrays

Gene silencing using RNA interference (RNAi) has become a prominent biological tool for gene annotation, pathway analysis, and target discovery in mammalian cells. High-throughput screens conducted using whole-genome siRNA libraries have uncovered rich sets of new genes involved in a variety of biological processes and cellular models of disease. However, high-throughput RNAi screening is not yet a mainstream tool in life science research because current screening platforms are expensive and onerous. Miniaturizing the RNAi screening platform to reduce cost and increase throughput will enable its widespread use and harness its potential for rapid genome annotation. With this aim, we have combined semi-conductor microfabrication and nanolitre dispensing techniques to develop miniaturized electroporation-ready microwell arrays loaded with siRNA molecules in which multiplexed gene knockdown can be achieved. Arrays of microwells are created using high-aspect ratio biocompatible photoresists on optically transparent and conductive Indium-Tin Oxide (ITO) substrates with integrated micro-electrodes to enable in situ electroporation. Non-contact inkjet microarraying allows precise dispensing of nanolitre volumes into the microwell structures. We have achieved parallel electroporation of multiple mammalian cells cultured in these microwell arrays and observed efficient knockdown of genes with surface-bound, printed siRNAs. Further integration of microfabrication and non-contact nanolitre dispensing techniques described here may enable single-substrate whole-genome siRNA screening in mammalian cells.     

Strain relief and disorder in commensurate water layers formed on Pd(111)

Water adsorbs and desorbs intact on Pd(111), forming a hydrogen-bonded wetting layer whose structure we examine by low energy electron diffraction (LEED) and He atom scattering (HAS). LEED shows that water forms commensurate (√3 × √3)R30° clusters that aggregate into a partially ordered, approximately (7 × 7) superstructure as the layer completes. HAS indicates that the water layer remains disordered on a local (approximately 10 ?) scale. Based on workfunction measurements and density functional theory simulations we propose that water forms small, flat domains of a commensurate (√3 × √3)R30° water network, separated by disordered domain boundaries containing largely H-down water. This arrangement allows the water layer to adapt its density and relieve the lateral strain associated with adsorbing water in the optimum flat atop adsorption site. We discuss different possibilities for the structure of these domain walls and compare this strain relief mechanism to the highly ordered, large unit cell structures formed on surfaces such as Pt(111).     

The metabolism of methylarsine oxide and sulfide

Methylarsine oxide and sulfide are more toxic to Candida humicola than arsenite; the sulfide is rapidly metabolized to trimethylarsine (Me₃As) and methylarsine (MeAsH₂) and the oxide to dimethylarsinic acid [Me₂AsO(OH)]. Cell-free extracts of C. humicola also convert the oxide to Me₂AsO(OH). The glutathione (RSH) derivative Me₂AsSR is metabolized by C. humicola to Me₃As and Me₂AsH, but some other Me₂AsSR′ compounds are unaffected. Studies involving the interaction of the arsenic(III) compounds with natural ecosystems and other micro-organisms such as Scopulariopsis brevicaulis, Straptococcus sanguis, Escherichia coli, and Veillonella alcalescens are described.     

Search for the KL-->pi 0 pi 0 e+e- decay in the KTeV experiment

The recent discovery of a large CP violating asymmetry in KL-->pi+pi-e+e- mode has prompted us to seach for the associated KL-->pi 0 pi 0 e+e- decay mode in the KTeV-E799 experiment at Fermilab. In 2.7 x 10(11) K(L) decays, one candidate event has been observed with an expected background of 0.3 event, resulting in an upper limit for the KL-->pi 0 pi 0 e+e- branching ratio of 6.6 x 10(-9) at the 90% C.L.     

Measurement of the wavelength dependence of beam divergence for photonic crystal fiber

We report measurements of the wavelength dependence of beam divergence for single-mode photonic crystal fiber. These measurements confirm predictions of strongly wavelength-dependent beam divergence, consistent with the effective-index model for the photonic crystal cladding material.     

Search for the rare decay K(L)-->pi(0)e(+)e(-)

The KTeV/E799 experiment at Fermilab has searched for the rare kaon decay K(L)-->pi(0)e(+)e(-). This mode is expected to have a significant CP violating component. The measurement of its branching ratio could support the standard model or could indicate the existence of new physics. This Letter reports new results from the 1999-2000 data set. One event is observed with an expected background at 0.99+/-0.35 events. We set a limit on the branching ratio of 3.5x10(-10) at the 90% confidence level. Combining with the previous result based on the data set taken in 1997 yields the final KTeV result: BR(K(L)-->pi(0)e(+)e(-))<2.8x10(-10) at 90% C.L.     

Measurements of the decay KL-->e+ e- gamma

The E799-II (KTeV) experiment at Fermilab has collected 83 262 K(L)-->e+ e- gamma(gamma) events above a background of 79 events. We measure a decay width, normalized to the K(L)-->pi0pi0pi(D)0 (pi0-->gammagamma, pi0-->gammagamma, pi(D0-->e+ e- gamma(gamma)) decay width, of Gamma(K(L)-->e+e-gamma(gamma))/Gamma(K(L)-->pi0pi0pi(D)0)=(1.3302+/-0.0046(stat)+/-0.0102(syst)) x 10(-3). We also measure parameters of two K(L)gamma*gamma form factor models. In the Bergstr?m-Massó-Singer parametrization, we find Calpha(K*)= -0.517 +/- 0.030(stat) +/- 0.022(syst). We separately fit for the first parameter of the D'Ambrosio-Isidori-Portolés model and find alpha(DIP)= -1.729 +/- 0.043(stat) +/- 0.028(syst).     

Phase behavior of attractive and repulsive ramp fluids: integral equation and computer simulation studies

Using computer simulations and a thermodynamically self-consistent integral equation we investigate the phase behavior and thermodynamic anomalies of a fluid composed of spherical particles interacting via a two-scale ramp potential (a hard core plus a repulsive and an attractive ramp) and the corresponding purely repulsive model. Both simulation and integral equation results predict a liquid-liquid demixing when attractive forces are present, in addition to a gas-liquid transition. Furthermore, a fluid-solid transition emerges in the neighborhood of the liquid-liquid transition region, leading to a phase diagram with a somewhat complicated topology. This solidification at moderate densities is also present in the repulsive ramp fluid, but in this case inhibits the fluid-fluid separation.