Quantifying Cell Binding Kinetics Mediated By Surface-Bound Blood Type B Antigen To Immobilized Antibodies

Cell adhesion is crucial to many biological processes, such as inflammatory responses, tumor metastasis and thrombosis formation. Recently a commercial surface plasmon resonance (SPR)-based BIAcore biosensor has been extended to determine cell binding mediated by surface-bound biomolecular interactions. How such cell binding is quantitatively governed by kinetic rates and regulating factors, however, has been poorly understood. Here we developed a novel assay to determine the binding kinetics of surface-bound biomolecular interactions using a commercial BIAcore 3000 biosensor. Human red blood cells (RBCs) presenting blood group B antigen and CM5 chip bearing immobilized anti-B monoclonal antibody (mAb) were used to obtain the time courses of response unit, or sensorgrams, when flowing RBCs over the chip surface. A cellular kinetic model was proposed to correlate the sensorgrams with kinetic rates. Impacts of regulating factors, such as cell concentration, flow duration and rate, antibody-presenting level, as well as pH value and osmotic pressure of suspending medium were tested systematically, which imparted the confidence that the approach can be applied to kinetic measurements of cell adhesion mediated by surface-bound biomolecular interactions. These results provided a new insight into quantifying cell binding using a commercial SPR-based BIAcore biosensor.

Measuring Binding Kinetics Of Surface-Bound Molecules Using The Surface Plasmon Resonance Technique

Surface plasmon resonance (SPR) technology and the Biacore biosensor have been widely used to measure the kinetics of biomolecular interactions in the fluid phase. In the past decade, the assay was further extended to measure reaction kinetics when two counterpart molecules are anchored on apposed surfaces. However, the cell binding kinetics has not been well quantified. Here we report development of a cellular kinetic model, combined with experimental procedures for cell binding kinetic measurements, to predict kinetic rates per cell. Human red blood cells coated with bovine serum albumin and anti-BSA monoclonal antibodies (mAbs) immobilized on the chip were used to conduct the measurements. Sensor-grams for BSA-coated RBC binding onto and debinding from the anti-BSA mAb-immobilized chip were obtained using a commercial Biacore 3000 biosensor, and analyzed with the cellular kinetic model developed. Not only did the model fit the data well, but it also predicted cellular on and off-rates as well as binding affinities from curve fitting. The dependence of flow duration, flow rate, and site density of BSA on binding kinetics was tested systematically, which further validated the feasibility and reliability of the new approach. Crown copyright (c) 2008 Published by Elsevier Inc. All rights reserved.

Surface-Bound Selectin-Ligand Binding Is Regulated By Carrier Diffusion

Two-dimensional (2D) kinetics of receptor-ligand interactions governs cell adhesion in many biological processes. While the dissociation kinetics of receptor-ligand bond is extensively investigated, the association kinetics has much less been quantified. Recently receptor-ligand interactions between two surfaces were investigated using a thermal fluctuation assay upon biomembrane force probe technique (Chen et al. in Biophys J 94:694-701, 2008). The regulating factors on association kinetics, however, are not well characterized. Here we developed an alternative thermal fluctuation assay using optical trap technique, which enables to visualize consecutive binding-unbinding transition and to quantify the impact of microbead diffusion on receptor-ligand binding. Three selectin constructs (sLs, sPs, and PLE) and their ligand P-selectin glycoprotein ligand 1 were used to conduct the measurements. It was indicated that bond formation was reduced by enhancing the diffusivity of selectin-coupled carrier, suggesting that carrier diffusion is crucial to determine receptor-ligand binding. It was also found that 2D forward rate predicted upon first-order kinetics was in the order of sPs > sLs > PLE and bond formation was history-dependent. These results further the understandings in regulating association kinetics of surface-bound receptor-ligand interactions.

Mechanism study on the frequent variations of cell-bound microcystins in cyanobacterial blooms in Lake Taihu: Implications for water quality monitoring and assessments

Although Microcystis-based toxins have been intensively studied, previous studies using laboratory cultures of Microcystis strains are difficult to explain the phenomenon that microcystin concentrations and toxin variants in natural blooms differ widely and frequently within a short-term period. The present study was designed to unravel the mechanisms for the frequent variations of intracellular toxins related to the differences in cyanobacterial colonies during bloom seasons in Lake Taihu, China. Monitoring of Microcystis colonies during warm seasons indicated that the variations in microcystins in both concentrations and toxin species were associated with the frequent alteration of Microcystis colonies in Lake Taihu. High concentration of microcystins in the blooms was always associated with two Microcystis colonies, Microcystis flos-aquae and Microcystis aeruginosa, whereas when Microcystis wesenbergii was the dominant colonial type, the toxin production of the blooms was low. Additionally, environmental factors such as temperature and nutrition were also shown to have an effect on the toxin production of the blooms, and may also potentially influence the Microcystis species present. The results of the present study provides insight into a new consideration for quick water quality monitoring, assessment and risk alert in cyanobacterium- and toxin-contaminated freshwaters, which will be beneficial not only for water agencies but also for public health. (C) 2009 Elsevier Ltd. All rights reserved.

The role of cell-surface-bound phosphatases in species competition within natural phytoplankton assemblage: an in situ experiment

Despite it is widely acknowledged that the ability to hydrolyze dissolved organic matter using extracellular phosphatases is diverse in fresh water phytoplankton, the competition within single species related to presence and quantity of cell-surface-bound phosphatases has not been examined in natural conditions yet. Here, we studied phytoplankton species competition in a freshwater reservoir during an in situ experiment. A natural plankton community, with the exclusion of large zooplankton, was enclosed in permeable dialysis bags inside two large containers of different bioavailable phosphate concentrations. Phytoplankton species biomass and the abundance of bacteria were determined in purpose to compare the development of enclosed microbial communities. Total and cell-surface-bound phosphatase activities in the phytoplankton were investigated using the Fluorescently Labelled Enzyme Activity (FLEA) technique that allows for direct microscopic detection of phosphatase-positive cells and, with image cytometry, enables quantification of phosphatase hydrolytic capacity. Production of extracellular phosphatases was not completely inhibited or stopped in the phosphate-enriched environment, phytoplankton cells only showed the activity less often. Under the phosphate-nonenriched conditions, the production of phosphatases was enhanced, but active species did not proliferate amongst phytoplankton assemblage. Further, specific growth rates of the phosphatase-positive species in the non-enriched environment were lower than the same phosphatase-positive species in phosphate-enriched environment. Interestingly, the phosphatase-positive cells of Ankyra ancora increased their size in both treatments equally, although the population in phosphate-enriched environment grew much faster and the cell-specific phosphatase activity was lower. We hypothesize that brand new daughter cells had sufficient phosphorus reserves and therefore did not employ extracellular phosphatases until they matured and needed extra bioavailable phosphorus to support their metabolism before cell division. Based on presented in situ experiment, we propose that the ability to hydrolyze organic polymers and particles with cell-surface-hound phosphatases is advantageous for longer persistence of given population in a phosphate-scarce environment; although phosphatase-positive species cannot dominate the reservoir phytoplankton solely because of specific phosphorus-scavenging strategy.

Weak measurement of qubit oscillations with strong response detectors: Violation of the fundamental bound imposed on linear detectors

Qubit measurement by mesoscopic charge detectors has received great interest in the community of mesoscopic transport and solid-state quantum computation, and some controversial issues still remain unresolved. In this work, we revisit the continuous weak measurement of a solid-state qubit by single electron transistors (SETs) in nonlinear-response regime. For two SET models typically used in the literature, we find that the signal-to-noise ratio can violate the universal upper bound "4," which is imposed quantum mechanically on linear-response detectors. This different result can be understood by means of the cross correlation of the detector currents by viewing the two junctions of the single SET as two detectors. Possible limitation of the potential-scattering approach to this result is also discussed.

Variational calculations of neutral bound excitons in GaAs quantum-well wires

The binding energy of an exciton bound to a neutral donor (D-0,X) in GaAs quantum-well wires is calculated variationally as a function of the wire width for different positions of the impurity inside the wire by using a two-parameter wavefunction. There is no artificial parameter added in our calculation. The results we have obtained show that the binding energies are closely correlated to the sizes of the wire, the impurity position, and also that their magnitudes are greater than those in the two-dimensional quantum wells compared. In addition, we also calculate the average interparticle distance as a function of the wire width. The results are discussed in detail.

Recombination property of nitrogen-acceptor-bound states in ZnO

The recombination property of nitrogen (N)-related acceptor-bound states in ZnO has been investigated by photoluminescence (PL), time-resolved PL, and selective PL. Several possible recombination processes were discussed by analyzing the relaxation and recombination properties under large Coulomb interaction. It is strongly suggested that bound exciton emission dominates the recombination process related to the N acceptor. The recombination lifetime is 750 ps and the binding energy is 67 meV for N-acceptor-bound exciton at low temperature. (c) 2006 American Institute of Physics.

Variational calculations of ionized-donor-bound excitons in GaAs-AlxGa1-xAs quantum wells

Using a two-parameter wave function, we calculate variationally the binding energy of an exciton bound to an ionized donor impurity (D+,X) in GaAs-AlxGa1-xAs quantum wells for the values of the well width from 10 to 300 Angstrom, when the dopant is located in the center of the well and at the edge of the well. The theoretical results confirm that the previous experimental speculation proposed by Reynolds tit al. [Phys. Rev. B 40, 6210 (1989)] is the binding energy of D+,X for the dopant at the edge of the well. in addition, we also calculate the center-of-mass wave function of the exciton and the average interparticle distances. The results are discussed in detail.

Binding energy of ionized-donor-bound excitons in the GaAs-AlxGa1-xAs quantum wells

The binding energy of an exciton bound to an ionized donor impurity (D+,X) located st the center or the edge in GaAs-AlxGa1-xAs quantum wells is calculated variationally for the well width from 10 to 300 Angstrom by using a two-parameter wave function, The theoretical results are discussed and compared with the previous experimental results.

Binding energy of excitons bound to neutral donors in two-dimensional semiconductors

The binding energies of excitons bound to neutral donors in two-dimensional (2D) semiconductors within the spherical-effective-mass approximation, which are nondegenerate energy bands, have been calculated by a variational method for a relevant range of the effective electron-to-hole mass ratio sigma. The ratio of the binding energy of a 2D exciton bound to a neutral donor to that of a 2D neutral donor is found to be from 0.58 to 0.10. In the limit of vanishing sigma and large sigma, the results agree fairly well with previous experimental results. The results of this approach are compared with those of earlier theories.

Linewidth of the infrared absorption spectra due to bound-to-continuum transition in GaAs/AlxGa1-xAs multiple quantum well structures

We have observed an extremely narrow absorption spectrum due to bound-to-continuum transition in GaAs/AlxGa1-xAs multiple quantum wells (MQWs). Its linewidth is only about one tenth of the values reported previously. Our calculation indicates that the broadening of the excited state in the continuum has little contribution to the absorption linewidth. We have grown a sample whose MQW region contains two kinds of wells with a minor thickness inhomogeneity. Its resultant absorption linewidth is six times as large as that of homogeneous well sample, which is in good agreement with our theoretical analysis. Thus we can suggest that the wider absorption spectra reported by many authors may be due to the well width inhomogeneity. (C) 1998 American Institute of Physics. [S0003-6951(98)03430-5]

The second dissociation threshold bound levels of hydrogen molecule

The near-threshold highly bound states of all three stable isotopic variants of molecular hydrogen have been studied. Numerous perturbations and unexpected transitions are observed as far as 1cm(-1) just below the second dissociation threshold. This complex structure may arise from a combination of nonadiabatic coupling between B, B', C electronic states, perturbations due to. ne and hyperfine interactions, and strong shape resonances. The perturbed near-threshold states and vibrational continuum exhibit finegrained structure, differing greatly between isotopes because of varying nonadiabatic coupling.