Parallel human genome analysis: microarray-based expression monitoring of 1000 genes

Microarrays containing 1046 human cDNAs of unknown sequence were printed on glass with high-speed robotics. These 1.0-cm2 DNA "chips" were used to quantitatively monitor differential expression of the cognate human genes using a highly sensitive two-color hybridization assay. Array elements that displayed differential expression patterns under given experimental conditions were characterized by sequencing. The identification of known and novel heat shock and phorbol ester-regulated genes in human T cells demonstrates the sensitivity of the assay. Parallel gene analysis with microarrays provides a rapid and efficient method for large-scale human gene discovery.     

The in vitro effect of platelet-derived growth factor isoforms on the proliferation of bovine corneal stromal fibroblasts depends on cell density

PURPOSE: Recently, it has been shown that corneal stromal fibroblasts express the mRNA for PDGF-beta-type receptors, while corneal epithelial cells express the mRNA for the PDGF B-chain, suggesting a role of PDGF isoforms in the regulation of corneal homeostasis and wound healing via an unidirectional epithelial to stromal paracrine interaction. The purpose of this study was to characterize the proliferative response of cultured bovine corneal stromal fibroblasts to PDGF isoforms. METHODS: Bovine corneal stromal fibroblasts were seeded at a cell density of 60 cells/mm2 (low density) and 120 cells/mm2 (high density) and were cultured under serum-free conditions. Except for corresponding controls, PDGF AA, BB and AB (obtained by separate expression of cloned genes in E. coli) were added in concentrations ranging from 3.125 to 100 ng/ml. Cell numbers were determined after an incubation period of 6 days using a cell counter. RESULTS: Stromal fibroblasts, when cultured at a high density, revealed constant cell numbers during the whole incubation period. Under these culture conditions, stimulation with PDGF AA, BB and AB led to a significant dose-dependent increase in cell proliferation. When cultured at a low cell density, stromal fibroblasts revealed a significant reduction of cell numbers after 6 days of incubation. This reduction was prevented by PDGF AA and AB isoforms in a dose-dependent manner. In contrast, PDGF BB was not effective. CONCLUSION: The results of the "high-density" assays suggest that PDGF isoforms act as mitogens for stromal fibroblasts during wound healing, when density of fibroblasts is high. The results of the "low-density" assays support the idea that PDGF AA and AB can prevent cell loss during corneal homeostasis when density of keratocytes is low.     

Single molecule fluorescence imaging of the photoinduced conversion and bleaching behavior of the fluorescent protein Kaede

Photoconversion and photobleaching behavior of the fluorescent protein Kaede immobilized in polyacrylamide gel matrix at room temperature was studied by single molecule wide-field fluorescence microscopy. Photobleaching kinetics of Kaede molecules upon excitation at 488 nm showed slight heterogeneity, suggesting the presence of different protein conformations and/or the distribution of local environments in the gel matrix. Statistical analysis of intensity trajectories of single molecules revealed four major types of fluorescence dynamics behavior upon short illumination by a violet light pulse (405 nm). In particular, two types of photoswitching behavior were observed: the green-to-red photoconversion (4% of Kaede molecules) and the photoactivation of green fluorescence without emission of red fluorescence (13%). Two other major groups show neither photoconversion nor red emission and demonstrate photoinduced partial deactivation (43%) and partial revival (30%) of green fluorescence. The significantly lower green-to-red conversion ratio as compared with bulk measurements in aqueous solution might be induced by the immobilization of the protein molecules within a polyacrylamide gel. Contrary to Ando et al. (Proc Natl Acad Sci 2002;99:12651-12656), we found a significant increase in green fluorescence emission upon illumination with 405-nm light, which is typical for GFP and related proteins.     

Turbulent Opposing Mixed Convection Heat Transfer in a Vertical Flat Channel with One-Side Heating

The paper presents experimental data on turbulent local heat transfer in a flat vertical channel with one-side heating for opposing mixed convection in wide ranges of airflow parameters (Re = 4 × 10³–4 × 10⁴; Gr_q = 1.7 × 10⁸–1.4 × 10¹⁰; pressures p = 0.2, 0.4, 0.6 MPa). Analysis has been performed by application of different buoyancy parameters. General correlation was suggested to predict the heat transfer rate. The experimental data were compared with the available correlations for vertical tubes.     

A novel order reduction method for nonlinear dynamical system under external periodic excitations

The concept of approximate inertial manifold (AIM) is extended to develop a kind of nonlinear order reduction technique for non-autonomous nonlinear systems in second-order form in this paper.Using the modal transformation,a large nonlinear dynamical system is split into a 'master' subsystem,a 'slave' subsystem,and a 'negligible' subsystem.Accordingly,a novel order reduction method (Method I) is developed to construct a low order subsystem by neglecting the 'negligible' subsystem and slaving the 'slave' sub...     

Temperature and Magnetic Field Effects on the Transport Controlled Charge State of a Single Quantum Dot

Individual InAs/GaAs quantum dots are studied by micro-photoluminescence. By varying the strength of an applied external magnetic field and/or the temperature, it is demonstrated that the charge state of a single quantum dot can be tuned. This tuning effect is shown to be related to the in-plane electron and hole transport, prior to capture into the quantum dot, since the photo-excited carriers are primarily generated in the barrier.     

No common major gene for apolipoprotein A-I and HDL3-C levels: evidence from bivariate segregation analysis

Apolipoprotein A-I (apo A-I) is the most abundant protein in high-density lipoprotein (HDL) particles, and it plays an important role in HDL metabolism. Both apo A-I and HDL cholesterol (HDL-C) levels are inversely associated with risk of cardiovascular disease. Segregation analyses suggest apo A-I levels are under the control of one or more major loci. Since HDL particles are heterogeneous in their composition and size, genetic influence on its subfractions (i.e., HDL2 and HDL3) could vary. A previous report showed evidence of a major locus controlling HDL3-C levels in a subset of the current study population. Because quantitative trait loci involved in complex diseases are likely to have pleiotropic effects on several related traits, it is possible to have a common major gene involved in regulating apo A-I and HDL3-C levels. We performed a bivariate segregation analysis of apo A-I and HDL3-C levels in 1,006 individuals from 137 families ascertained through probands undergoing elective, diagnostic coronary angiography at the Johns Hopkins Hospital. The results showed significant genetic correlation between these two traits, but the hypothesis of a common major gene was rejected. Bivariate segregation analysis favored a model with two genes controlling apo A-I and a third gene independently controlling HDL3-C, and the genetic correlation between these two traits is due to residual additive polygenes. Overall, results from this study suggest that there are distinct genetic mechanisms for apo A-I and HDL3-C levels. Future studies, especially linkage analysis, should consider distinct genetic mechanisms and multiple major gene loci.     

Time correlated fluorescence characterization of an asymmetrically focused flow in a microfluidic device

In this study, we explore and model the behavior of a prototype microfluidic device which employs two non-mixing fluids (sheath and inlet fluids) displaying an asymmetric focused flow, in the presence of a fluorescent dye. Fluorescence correlation spectroscopy is employed, allowing the precise measure of flow speeds across the channels and of the concentration profile of the central focused flux along the flow direction. The system is modeled via a standard Navier–Stokes finite-element approach, coupled to convection–diffusion equations for the solute. Simulations reproduce accurately the shape, the position, and the width of the velocity and concentration profiles along the central channel and across the transversal and vertical sections of the microfluidic device. The observed asymmetric flow with respect to the center of the channel is reproduced numerically with an error in the position determination smaller than 1%.