Multiplexed microfluidic enzyme assays for simultaneous detection of lipolysis products from adipocytes

Microfluidics has enabled new cell biology experiments. Incorporating chemical monitoring of cellular secretion into chips offers the potential to increase information content and utility of such systems. In this work, an integrated, multilayer polydimethylsiloxane microfluidic chip was developed to simultaneously measure fatty acids and glycerol secreted from cultured adipocytes on chip in near real time. Approximately 48,000 adipocytes were loaded into a cell chamber in a reversibly sealed chip. Cells were perfused at 0.75 μL/min. Cell perfusate was split and directed to separate, continuously operating fluorescent enzyme assay channel networks. The fluorescent assay products were detected simultaneously near the outlet of the chip. The fatty acid and glycerol assays had linear dynamic ranges of 150 and 110 μM and limit of detection (LOD) of 6 and 5 μM, respectively. Surface modifications including pretreatment with sodium dodecyl sulfate were utilized to prevent adsorption of fatty acids to the chip surface. Using the chip, basal fatty acid and glycerol concentrations ranged from 0.18 to 0.7 nmol?×?10⁶ cell^?1 min^?1 and from 0.23 to 0.85 nmol?×?10⁶ cell^?1 min^?1, respectively. Using valves built into the chip, the perfusion solution was switched to add 20 μM isoproterenol, a β-adrenergic agonist, which stimulates the release of glycerol and fatty acids in adipocytes. This manipulation resulted in a rapid and stable 1.5- to 6.0-fold increase of non-esterified fatty acid (NEFA) and glycerol. The ratio of NEFA:glycerol released increased with adipocyte age. These experiments illustrate the potential for performing multiple real-time assays on cells in culture using microfluidic devices.     

Control of a coupled map lattice model for vortex shedding in the wake of a cylinder

The flow behind a vibrating flexible cable at low Reynolds numbers can exhibit complex wake structures such as lace-like patterns, vortex dislocations and frequency cells. These structures have been observed in experiments and numerical simulations, and are predicted by a previously developed low-order coupled map lattice (CML). The discrete (in time and space) CML models consist of a series of diffusively coupled circle map oscillators along the cable span. Motivated by a desire to modify the complex wake patterns behind flexible vibrating cables we have studied the addition of control terms into the highly efficient CML models and explored the resulting dynamics. Proportional, adaptive proportional and discontinuous non-linear (DNL) control methods were used to derive the control laws. The first method employed occasional proportional feedback. The adaptive method used spatio-temporal feedback control. The DNL method used a discontinuous feedback linearization procedure, and the controller was designed for the resulting linearized system using eigenvalue assignment. These techniques were applied to a modeled vortex dislocation structure in the wake of a vibrating cable in uniform freestream flow. Parallel shedding patterns were achieved for a range of forcing frequency-forcing amplitude combinations studied to validate the control theory. The adaptive proportional and DNL methods were found to be more effective than the proportional control method due to the incorporation of a spatially varying feedback gain across the cylinder span. The DNL method was found to be the most efficient controller of the low-order CML model. The required control level across the cable span was correlated to the 1/1 lock-on behavior of the temporal circle map.     

The development of information storage materials — How microscopy can help?

The response of giant magnetoresistance (GMR) devices depends critically on the film microstructure, with parameters such as layer thickness and interfacial abruptness being crucial. This paper presents results obtained using high resolution electron microscopy (HREM), chemical mapping and atom probe microanalysis. Local variations in the magnetic properties are induced by the microstructure and also when the films are patterned to form small elements. These lead to changes in the magnetization reversal mechanism. Some results of the studies of the magnetization reversal carried out using in situ in Lorentz transmission electron microscopy (LTEM) magnetizing experiments are also included.     

Reversibly sealed multilayer microfluidic device for integrated cell perfusion and on-line chemical analysis of cultured adipocyte secretions

A three-layer microfluidic device was developed that combined perfusion of cultured cells with on-line chemical analysis for near real-time monitoring of cellular secretions. Two layers were reversibly sealed to form a cell chamber that allowed cells grown on coverslips to be loaded directly into the chip. The outlet of the chamber was in fluidic contact with a third layer that was permanently bonded. Perfusate from the cell chamber flowed into this third layer where a fluorescence enzyme assay for non-esterified fatty acid (NEFA) was performed on-line. The device was used to monitor efflux of NEFAs from ∼6,200 cultured adipocytes with 83 s temporal resolution. Perfusion of murine 3T3-L1 cultured adipocytes resulted in an average basal concentration of 24.2 ± 2.4 μM NEFA (SEM, n = 6) detected in the effluent corresponding to 3.31 × 10⁻⁵ nmol cell⁻¹ min⁻¹. Upon pharmacological treatment with a β-adrenergic agonist to stimulate lipolysis, a 6.9 ± 0.7-fold (SEM, n = 6) sustained increase in NEFA secretion was observed. This multilayer device provides a versatile platform that could be adapted for use with other cell types to study corresponding cellular secretions in near real-time.     

Temperature dependence of the penetration depth in Sr2RuO4: evidence for nodes in the gap function

We report measurements of the magnetic penetration depth in single crystals of Sr2RuO4 down to 0.04 K using a tunnel-diode based, self-inductive technique. We observe a power law temperature dependence below 0.8 K, with no sign of a second phase transition nor of a crossover predicted for a multiband superconductor. A power law dependence suggests that the gap function has nodes, inconsistent with candidate p-wave states. We argue that nonlocal effects, rather than impurity scattering, can explain the observed T2 dependence instead of the T-linear behavior expected for line nodes.     

Derivations, isomorphisms, and second cohomology of generalized Witt algebras

Generalized Witt algebras, over a field of characteristic , were defined by Kawamoto about 12 years ago. Using different notations from Kawamoto's, we give an essentially equivalent definition of generalized Witt algebras over , where the ingredients are an abelian group , a vector space over , and a map which is linear in the first variable and additive in the second one. In this paper, the derivations of any generalized Witt algebra
, with the right kernel of being , are explicitly described; the isomorphisms between any two simple generalized Witt algebras are completely determined; and the second cohomology group for any simple generalized Witt algebra is computed. The derivations, the automorphisms and the second cohomology groups of some special generalized Witt algebras have been studied by several other authors as indicated in the references.