Biomedical subjects

Tinglu Yang

Publications and source records attributed to Tinglu Yang.

7 recordsLinked to original sources

PubMed2005

Effect of average phospholipid curvature on supported bilayer formation on glass by vesicle fusion.

The adsorption of large unilamellar vesicles composed of various combinations of phosphatidylcholine, phosphatidylethanolamine (PE), monomethyl PE, and dimethyl PE (PE-Me2) onto a glass surface was studied using fluorescence microscopy. The average lipid geometry within the vesicles, described mathematically by the average intrinsic curvature, C(0,ave), was methodically altered by changing the lipid ratios to determine the effect of intrinsic curvature on the ability of vesicles to rupture and form a supported lipid bilayer. We show that the ability of vesicles to create fluid planar bilayers is dependent on C(0,ave) and independent of the identity of the component lipids. When the C(0,ave) was approximately -0.1 nm(-1), the vesicles readily formed supported lipid bilayers with almost full mobility. In contrast, when the C(0,ave) ranged from approximately -0.2 to approximately -0.3 nm(-1), the adsorbed vesicles remained intact upon the surface. The results indicate that the average shape of lipid molecules within a vesicle (C(0,ave)) is essential for determining kinetically viable reactions that are responsible for global geometric changes.

Absorption↗

PubMed2005

Fluid and air-stable lipopolymer membranes for biosensor applications.

The behavior of poly(ethylene glycol) (PEG) conjugated lipids was investigated in planar supported egg phosphatidylcholine bilayers as a function of lipopolymer density, chain length of the PEG moiety, and type of alkyl chains on the PEG lipid. Fluorescence recovery after photobleaching measurements verified that dye-labeled lipids in the membrane as well as the lipopolymer itself maintained a substantial degree of fluidity under most conditions that were investigated. PEG densities exceeding the onset of the mushroom-to-brush phase transition were found to confer air stability to the supported membrane. On the other hand, substantial damage or complete delamination of the lipid bilayer was observed at lower polymer densities. The presence of PEG in the membrane did not substantially hinder the binding of streptavidin to biotinylated lipids present in the bilayer. Furthermore, above the onset of the transition into the brush phase, the protein binding properties of these membranes were found to be very resilient upon removal of the system from water, rigorous drying, and rehydration. These results indicate that supported phospholipid bilayers containing lipopolymers show promise as rugged sensor platforms for ligand-receptor binding.

Biosensing Techniques↗

PubMed2005

Glycolipids as receptors for Bacillus thuringiensis crystal toxin.

The development of pest resistance threatens the effectiveness of Bacillus thuringiensis (Bt) toxins used in transgenic and organic farming. Here, we demonstrate that (i) the major mechanism for Bt toxin resistance in Caenorhabditis elegans entails a loss of glycolipid carbohydrates; (ii) Bt toxin directly and specifically binds glycolipids; and (iii) this binding is carbohydrate-dependent and relevant for toxin action in vivo. These carbohydrates contain the arthroseries core conserved in insects and nematodes but lacking in vertebrates. We present evidence that insect glycolipids are also receptors for Bt toxin.

Animals↗

PubMed2004

Creating fluid and air-stable solid supported lipid bilayers.

Solid supported lipid bilayers are rapidly delaminated when drawn through the air/water interface. We have discovered that a close packed monolayer of specifically bound protein prevents this process. The protection mechanism worked in two ways. First, when protein-protected bilayers were drawn through the air/water interface, a thin bulk water layer was visible over the entire bilayer region, thereby preventing air from contacting the surface. Second, a stream of nitrogen was used to remove all bulk water from a protected bilayer, which remained fully intact as determined by fluorescence microscopy. The condition of this dried bilayer was further probed by fluorescence recovery after photobleaching. It was found that lipids were not two-dimensionally mobile in dry air. However, when the bilayer was placed in a humid environment, 91% of the bleached fluorescence signal was recovered, indicating long-range two-dimensional mobility. The diffusion coefficient of lipids under humid conditions was an order of magnitude slower than the same bilayer under water. Protected bilayers could be rehydrated after drying, and their characteristic diffusion coefficient was reestablished. Insights into the mechanism of bilayer preservation were suggested.

Air↗

PubMed2003

Investigations of bivalent antibody binding on fluid-supported phospholipid membranes: the effect of hapten density.

Investigations of ligand-receptor binding between bivalent antibodies and membrane-bound ligands are presented. The purpose of these studies was to explore binding as a function of hapten density in a two-dimensionally fluid environment. A novel microfluidic strategy in conjunction with total internal reflection fluorescence microscopy was designed to achieve this. The method allowed binding curves to be acquired with excellent signal-to-noise ratios while using only minute quantities of protein solution. The specific system investigated was the interaction between anti-DNP antibodies and phospholipid membranes containing DNP-conjugated lipids. Binding curves for ligand densities ranging from 0.1 to 5.0 mol % were obtained. Two individual dissociation constants could be extracted from the data corresponding to the two sequential binding events. The first dissociation constant, K(D1), was 2.46 x 10(-)(5) M, while the second was K(D2) = 1.37 x 10(-)(8) mol/m(2). This corresponded to a positively cooperative binding effect with an entropic difference between the two events of 62.3 +/- 2.7 J/(mol.K). Furthermore, the percentage of monovalently and bivalently bound protein was determined at each ligand density.

2,4-Dinitrophenol↗

PubMed2002

A microfluidic device with a linear temperature gradient for parallel and combinatorial measurements.

Methods for obtaining combinatorial and array-based data as a function of temperature are needed in the chemical and biological sciences. It is presently quite difficult to employ temperature as a variable using standard wellplate formats simply because it is very inconvenient to keep each well at a distinct temperature. In microfluidics, however, the situation is very different due to the short length scales involved. In this article, it is shown how a simple linear temperature gradient can be generated across dozens of parallel microfluidic channels simultaneously. This result is exploited to rapidly obtain activation energies from catalytic reactions, melting point transitions from lipid membranes, and fluorescence quantum yield curves from semiconductor nanocrystal probes as a function of temperature. The methods developed here could quite easily be extended to protein crystallization, phase diagram measurements, chemical reaction optimization, or multivariable experiments.

Alkaline Phosphatase↗

PubMed2002

Design and characterization of immobilized enzymes in microfluidic systems.

Herein we report the fabrication, characterization, and use of total analytical microsystems containing surface-immobilized enzymes. Streptavidin-conjugated alkaline phosphatase was linked to biotinylated phospholipid bilayers coated inside poly(dimethylsiloxane) microchannels and borosilicate microcapillary tubes. Rapid determination of enzyme kinetics at many different substrate concentrations was made possible by carrying out laminar flow-controlled dilution on-chip. This allowed Lineweaver-Burk analysis to be performed from a single experiment with all the data collected simultaneously. The results revealed an enzyme turnover number of 51.1 +/- 3.2 s(-1) for this heterogeneous system. Furthermore, the same enzyme immobilization strategy was extended to demonstrate that multiple chemical reactions could be performed in sequence by immobilizing various enzymes in series. Specifically, the presence of glucose was detected by two coupled steps employing immobilized avidinD-conjugated glucose oxidase and streptavidin-conjugated horseradish peroxidase.

Bacterial Proteins↗