Nonradioactive Northern blotting with biotinylated and digoxigenin-labeled RNA probes

The application of nonradioactive RNA probes for Northern blotting offers the advantage of a rapid turn-around time for results without the loss of sensitivity for target mRNA detection. However, a problem that has impeded the widespread use of nonradioactive RNA probes for use in Northern blotting is the difficulty in stripping these probes from nylon membranes after hybridization. In this report we describe two protocols for stripping digoxigenin (Dig)-labeled RNA probes from nylon membranes. One protocol utilizes a phosphate-buffered formamide stripping solution to remove nonchemically modified (regular) RNA probes while the other method utilizes strippable probes that were produced with a chemically modified nucleotide (CTP) and removed by a specific stripping solution. This latter method was developed by Ambion Inc. and is called Strip-EZ. We also describe a protocol for the detection of two separate rat mRNAs using both biotin and digoxigenin-labeled RNA probes that does not require stripping the membrane after hybridization. Finally, we describe the use of another new labeling technology, called Chem-Link, that quickly and conveniently labels RNA for use in Northern blotting.     

Clinical utility of the polymerase chain reaction in the diagnosis of extrapulmonary tuberculosis

This study examines the diagnostic utility of the polymerase chain reaction (PCR) in 156 patients (five human immunodeficiency virus (HIV) seropositive) suspected of extrapulmonary tuberculosis. The results of PCR in 226 samples from 11 different sites were compared with the results of microscopy and culture. Positive culture results were predicted in 86% of samples by PCR but in only 31% by microscopy. Specificity of PCR was 92%. In cases with culture-proven tuberculosis, PCR identified all 11 microscopy positive cases and 19 of 24 (79%) of the microscopy-negative cases. In four patients, PCR excluded the diagnosis of tuberculosis in microscopy-positive samples, which were later shown to contain mycobacteria other than Mycobacterium tuberculosis or laboratory contaminants. In 20 patients (microscopy, PCR and culture negative) a trial of antituberculous drugs was given, but patients showed no improvement and treatment was stopped. In 17 patients, all culture negative (in nine PCR was positive, three of whom also had positive microscopy) the diagnosis was probable tuberculosis based on clinical findings and response to treatment. This polymerase chain reaction has a much higher sensitivity than microscopy and can facilitate therapeutic decisions for those with suspected extrapulmonary tuberculosis.     

Error handling in a parallel LR substring parser

Parallel parsing is currently receiving attention but there is little discussion about the adaptation of sequential error handling techniques to these parallel algorithms. We describe a noncorrecting error handler implemented with a parallel LR substring parser. The parser used is a parallel version of Cormack's LR substring parser. The applicability of noncorrecting error handling for parallel parsing is discussed. The error information provided for a standard set of 118 erroneous Pascal programs is analysed. The programs are run on the sequential LR substring parser.     

Quantitative ECT: Comparison of Recovery Coefficient and Linearity of Detector Response for Single Photon and Coincidence Detection (F-18, B+ 511 keV)

The response characteristics of two tomographic systems were compared for imaging of positron emitters: a) a SPECT system with a 3/8 in crystal and 511 keV detector shielding, equipped with a specially designed 511 keV collimator, and b) a PET V system using coincidence detection. SPECT transverse plane resolution was 19 mm FWHM and 35 mm FWTM for a radius of rotation of 16 cm. Corresponding resolution for PET was 14 mm FWHM and 28 mm FWTM. Transverse images through a phantom containing cylindrical sources of various cross sections and uniform activity were obtained for each detector. The measured count density or recovery coefficient was found to decrease with source size, the dependence being similar for both systems. The theoretical values for recovery coefficients were calculated by convolution of a Gaussian fit to the SPECT resolution (FWHM, FWTM) values with the uniform cross section of each source. This simple mathematical model confirmed that the recovery coefficient dependence on source size was primarily related to the limited resolution of the detector. Experimental measurements demonstrated that the SPECT resolution for volume sources was sufficient for quantitation, although some limitations exist with respect to source sizes smaller than the detector resolution.     

Free fatty acid inhibition of the insulin induction of glucose-6-phosphate dehydrogenase in rat hepatocyte monolayers

Rat hepatocytes in monolayer culture were utilized to determine if the decrease in glucose-6-phosphate dehydrogenase (G6PD) activity resulting from the ingestion of fat can be mimicked by the addition of fatty acids to a chemically, hormonally defined medium. G6PD activity in cultured hepatocytes was induced several-fold by insulin. Dexamethasone or T3 did not amplify the insulin induction of G6PD. Glucose alone increased G6PD activity in cultured hepatocytes from fasted donors by nearly 500%. Insulin in combination with glucose induced G6PD an additional two-fold. The increase in G6PD activity caused by glucose was greater in hepatocytes isolated from 72 hr-fasted rats as compared to fed donor rats. Such a response was reminiscent of the “overshoot” phenomenon in which G6PD activity is induced well above the normal level by fasting-refeeding rats a high glucose diet. Addition of linoleate to the medium resulted in a significant suppression of insulin’s ability to induce G6PD, but linoleate had no effect on the induction of G6PD activity by glucose alone. A shift to the right in the insulin-response curve for the induction of G6PD also was detected for the induction of malic enzyme and acetyl-CoA carboxylase. Arachidonate (0.25 mM) was a significantly more effective inhibitor of the insulin action than linoleate was. Apparently rat hepatocytes in monolayer culture can be utilized as a model to investigate the molecular mechanism by which fatty acids inhibit the production of lipogenic enzymes. In part, this mechanism of fatty acid inhibition involves desensitization of hepatocytes to the lipogenic action of insulin.     

Comparison of single and coastal superphosphate for subterranean clover on phosphorus leaching soils

Coastal superphosphate, a partially acidulated rock phosphate (PARP), is being considered as an alternative fertilizer to single superphosphate for pastures in high rainfall (> 800 mm annual average) areas of south-western Australia. The effectiveness of single and coastal superphosphate, as P fertilizers, was measured in two field experiments using dry herbage yield of subterranean clover (Trifolium subterraneum). The experiments were started in April 1990 and were terminated at the end of 1993. In the years after P applications, soil samples were collected each January to measure Colwell soil-test P, which was related to plant yields measured later on that year, to provide soil P test calibrations.Relative to freshly-applied single superphosphate, the effectiveness of freshly-applied coastal superphosphate and the residues of previously-applied single and coastal superphosphate were less effective in some years (from 3% as effective to equally effective), and up to 100% more effective in other years. This large range in effectiveness values in different years is attributed to different climatic conditions. Soil P test calibrations were different for soils treated with single or coastal superphosphate. The calibrations were also different for different yield assessments (harvests) in the same year, and in different years. Consequently soil P testing can only provide a very crude estimate of the current P status of the soils.     

Effect of Zwitterionic Surfactants on Interparticle Forces, Rheology, and Particle Packing of Silicon Nitride Slurries

Phosphocholine (PC) zwitterionic surfactants, with different hydrocarbon chain lengths (C₆C₆PC to C₉C₉PC), were absorbed on the surface of silicon nitride near the isoelectric point (pH 6). Adsorption of the surfactants changed the lateral and normal surface forces, the rheology, and the consolidation behavior of the particles. The normal force between two silicon nitride surfaces as a function of separation and the lateral (friction) forces were measured using an atomic force microscope (AFM). These measurements indicated that surfactant adsorption reduced the magnitude of the long-range attractive van der Waals force and produced a repulsive short-range force. Although the adsorbed layers provided a barrier to particle contact, they could be ejected with a critical force that increased with the hydrocarbon chain length. The effect of an adsorbed layer on the viscosity and consolidation of slurries was also measured. The viscosity of all slurries decreased with increasing shear rate, indicative of attractive particle networks. The highest viscosity was observed for slurries formulated at the isoelectric point without added surfactant. Much lower viscosities were observed when the surfactant concentration was greater than the critical micelle concentration (cmc). A relative density of 0.46 was obtained via pressure filtration at 4 MPa without a surfactant, and between 0.46 to 0.59 (C₆C₆PC to C₉C₉PC, respectively) for surfactant concentrations greater than the cmc. Comparing force measurements with rheology and packing density provides a basis for discussing the role of interparticle forces in ceramic powder processing via colloidal routes.     

Deformation Bands in Ceria-Stabilized Tetragonal Zirconia/Alumina : II, Stress-Induced Aging at Room Temperature

A stress-induced aging phenomenon is observed to occur at room temperature in deformation bands introduced into a 8.5 mol% ceria-stabilized tetragonal zirconia/alumina (CeTZP/A1₂O₃) composite by flexural loading. The aging occurs with time after unloading and in laboratory air. Over a period of 100 days, the concentration of monoclinic zirconia within a deformation band increases and, in addition, the wedge-shaped deformation band grows with time. Accompanying these two changes are an increase in the tensile stress in the remaining tetragonal zirconia within the deformation band and a consequential increase in the overall compressive stress within the band. The average value of the monoclinic concentration within the deformation band is found to increase parabolically with time, suggesting the mechanism responsible for the observed aging is diffusion limited. Away from the deformation bands, no aging is observed to occur, suggesting aging is stress dependent. Although a water-vapor-mediated mechanism cannot be ruled out, it is proposed that the observed aging is in fact due to a tensile stress assisted chemical reduction of Ce⁴⁺ to Ce³⁺ whose rate is controlled by the indiffusion of oxygen vacancies driven by the tensile stress gradient. It is further proposed that the deformation band grows with time, since the region ahead of the band is under tension and hence subject to an enhanced rate of reduction     

云环境下基于本体的用户行为分析引擎研究

针对当前大型互联网站提升用户体验的需求,本研究创造性地提出基于本体的用户行为分析引擎的思想,它通过上下文感知技术,实时获取用户访问页面时的上下文信息(以下也称为动态行为),基于本体对用户行为建模,建立基于规则的上下文推理算法,对监测到的动态用户行为进行实时推理,及时反馈结果信息给用户。实验表明:本研究理论上独辟蹊径,设计实现上结合了云存储技术,能够一定程度改善用户体验效果。     

Aromatic bisphenol A-based elastomers via hydrosilylation chemistry

The preparation of highly aromatic elastomers from a bisphenol A-based divinyl-terminated resin and polymerization with various aromatic silane containing compounds utilizing a room temperature hydrosilylation reaction is demonstrated. The polymers exhibit high thermal and oxidative stability with 5% weight losses around 430 and 350°C and char yields ranging from 35% to 40%. The thermosets maintained their elastomeric properties with good hardness and mechanical properties as measured by elongation measurements. The toughness of the thermosets was not improved with the inclusion of aromatic moieties but the hardness did appear to increase with the addition of more aromatic groups.