Bovine Serum Albumin Layer Research Articles

Protein modified upconversion nanoparticles for imaging-guided combined photothermal and photodynamic therapy

In this work, we develop a multifunctional nano-platform by coating upconversion nanoparticles (UCNPs) with bovine serum albumin (BSA), obtaining UCNP@BSA nanoparticles with great solubility and stability in physiological environments. Two types of dye molecules, including a photosensitizer, Rose Bengal (RB), and an NIR-absorbing dye, IR825, can be simultaneously loaded into the BSA layer of the UCNP@BSA nanoparticles. In this carefully designed UCNP@BSA-RB&; IR825 system, RB absorbs green light emitted from UCNPs under 980-nm excitation to induce photodynamic cancer cell killing, while IR825 whose absorbance shows no overlap with upconversion excitation and emission wavelengths, offers nanoparticles a strong photothermal perform under 808-nm laser irradiation. Without showing noticeable dark toxicity, the obtained dual-dye loaded nanoparticles are able to kill cancer via combined photothermal and photodynamic therapies, both of which are induced by NIR light with high tissue penetration, by a synergetic manner both in vitro and in vivo. In addition, the intrinsic paramagnetic and optical properties of Gd3+-doped UCNPs can further be utilized for in vivo dual modal imaging. Our study suggests that UCNPs with well-designed surface engineering could serve as a multifunctional nano-platform promising in cancer theranostics.

Quenching of the emission of peroxydisulfate system by ferrocene functionalized chitosan nanoparticles: A sensitive “signal off” electrochemiluminescence immunosensor

In this paper, a novel sensitive “signal off” electrochemiluminescence (ECL) immunosensor was constructed by ferrocene functionalized chitosan nanoparticles (Fc-CsNPs) as tag for quenching peroxydisulfate system emission for the first time. Herein, CsNPs were prepared by a facile counterion complex coacervation method, and then numerous Fc and detection antibodies (Ab2) were immobilized together via covalent cross-linking. The sensing platform was constructed by assembling carboxyl polyamidoamine (PAMAM) dendrimers onto bovine serum albumin (BSA) modified electrode. Herein, BSA layer provided a multifunctional interface with good biocompatibility and sufficient active sites to produce conjugates with PAMAM. Then, PAMAM may play dual roles that not only as carriers to immobilize numerous capture antibodies but also as enhancer to significantly increase the ECL of peroxydisulfate system, thus, greatly improving the immunosensors’ sensitivity. After a sandwich-type immunoreaction, a remarkable decrease of ECL signal was observed due to the ECL quenching of peroxydisulfate by Fc-CsNPs labeled Ab2. As a result, the logarithm of change of ECL intensity has a direct relationship with the logarithm of N-terminal pro-B-type natriuretic peptide concentration in the range of 0.001–2pg/mL with a detection limit of 0.33fg/mL (S/N=3).

Potential-Assisted Adsorption of Bovine Serum Albumin onto Optically Transparent Carbon Electrodes

This article describes the effect of the applied potential on the adsorption of bovine serum albumin (BSA) to optically transparent carbon electrodes (OTCE). To decouple the effect of the applied potential from the high affinity of the protein for the bare surface, the surface of the OTCE was initially saturated with a layer of BSA. Experiments described in the article show that potential values higher than +500 mV induced a secondary adsorption process (not observed at open-circuit potential), yielding significant changes in the thickness (and adsorbed amount) of the BSA layer obtained. Although the process showed a significant dependence on the experimental conditions selected, the application of higher potentials, selection of pH values around the isoelectric point (IEP) of the protein, high concentrations of protein, and low ionic strengths yielded faster kinetics and the accumulation of larger amounts of protein on the substrate. These experiments, obtained around the IEP of the protein, contrast with the traditional hypothesis that enhanced electrostatic interactions between the polarized substrate and the (oppositely charged) protein are solely responsible for the enhanced adsorption. These results suggest that the potential applied to the electrode is able to polarize the adsorbed layer and induce dipole-dipole interactions between the adsorbed and the incoming protein. This mechanism could be responsible for the potential-dependent oversaturation of the surface and could bolster to the development of surfaces with enhanced catalytic activity and implants with improved biocompatibility.

Effects of organic macromolecular conditioning on gypsum scaling of forward osmosis membranes

This study analyzed the membrane surface conditioning due to the adsorption of different organic macromolecules and investigated how such conditioning affected the gypsum scaling behavior in forward osmosis (FO) membrane processes. A commercially available cellulose acetate FO membrane was used as the base membrane. The membrane surface was conditioned respectively by representative organic macromolecules, namely bovine serum albumin (BSA), Aldrich humic acid (AHA), and sodium alginate, prior to gypsum scaling experiments. Results of direct observation and flux decline experiments showed that the chemistry of organic macromolecules greatly influenced the gypsum scaling of an FO membrane thus conditioned. Specifically, the AHA and alginate conditioning increased gypsum crystal sizes, shortened the crystal nucleation time, and accelerated the flux decline. In contrast, the BSA conditioning noticeably hindered the formation of gypsum crystals and thus slowed down the flux decline caused by gypsum scaling. Further investigation of the conditioning layer properties by quartz crystal microbalance with dissipation and the scaling layer morphologies by SEM suggested that the conditioning layers of AHA and alginate, each containing a high density of carboxylate functional groups along their long chain structures, exhibited strong adsorption of calcium ions as well as accelerated surface or heterogeneous crystallization; in comparison, the conditioning layer of BSA, which has a low carboxylate density and a three-dimensional structure, showed a low level of calcium ion adsorption and a slowdown in gypsum crystallization.

Controlling Mixed-Protein Adsorption Layers on Colloidal Alumina Particles by Tailoring Carboxyl and Hydroxyl Surface Group Densities

We show that different ratios of bovine serum albumin (BSA) and lysozyme (LSZ) can be achieved in a mixed protein adsorption layer by tailoring the amounts of carboxyl (-COOH) and aluminum hydroxyl (AlOH) groups on colloidal alumina particles (d50 ≈ 180 nm). The particles are surface-functionalized with -COOH groups, and the resultant surface chemistry, including the remaining AlOH groups, is characterized and quantified using elemental analysis, ζ potential measurements, acid-base titration, IR spectroscopy, electron microscopy, nitrogen adsorption, and dynamic light scattering. BSA and LSZ are subsequently added to the particle suspensions, and protein adsorption is monitored by in situ ζ potential measurements while being quantified by UV spectroscopy and gel electrophoresis. A comparison of single-component and sequential protein adsorption reveals that BSA and LSZ have specific adsorption sites: BSA adsorbs primarily via AlOH groups, whereas LSZ adsorbs only via -COOH groups (1-2 -COOH groups on the particle surface is enough to bind one LSZ molecule). Tailoring such groups on the particle surface allows control of the composition of a mixed BSA and LSZ adsorption layer. The results provide further insight into how particle surface chemistry affects the composition of protein adsorption layers on colloidal particles and is valuable for the design of such particles for biotechnological and biomedical applications.

On the effects of atmospheric-pressure microplasma array treatment on polymer and biological materials

This paper reports the first systematic investigation on the effects of atmospheric-pressure helium microplasma array treatment on the surface chemistries of model organic materials and a biological coating. These materials include polystyrene (PS), low-density polyethylene (LDPE) and ethylene tetrafluoroethylene (ETFE), and bovine serum albumin (BSA). The plasma treatment introduced a range of oxygen functionalities into the surface of the polymers, with oxygen incorporation reaching “saturation” after relatively short treatment times. PS and LDPE surfaces were more readily oxidised and to a greater depth compared to ETFE. The polymer surfaces became smoother at short plasma treatment times due to removal of adventitious hydrocarbon, but became rougher at longer treatment times as a result of etching of low molecular weight, volatile material from the surface. Atmospheric-pressure helium microplasma array treatment of a BSA layer resulted in the majority of the protein being removed from the underlying (PS) surface. The plasma treatment reduced the surface roughness of the BSA coating at short treatment times, but at longer treatment times, the surface roughness increased and the surfaces exhibited granular structures. All of the hydrophobic polymers became hydrophilic after the plasma treatment. The hydrophilicity of the surfaces decreased upon storage (hydrophobic recovery) and none of the polymers reverted to their original hydrophobic state, even after 500 h of storage. The knowledge presented in this paper may be useful in the development of new manufacturing processes based on atmospheric-pressure plasma and in the field of plasma medicine, particularly with respect to cleaning and sterilisation methods. In addition, it provides a foundation for future efforts to establish the mechanisms behind interactions of atmospheric-pressure plasmas with materials.

Competition of Bovine Serum Albumin Adsorption and Bacterial Adhesion onto Surface-Grafted ODT: In Situ Study by Vibrational SFG and Fluorescence Confocal Microscopy

The interaction of hydrophilic and hydrophobic ovococcoid bacteria and bovine serum albumin (BSA) proteins with a well ordered surface of octadecanethiol (ODT) self assembled monolayer (SAM) has been studied in different situations where proteins were either preadsorbed on ODT or adsorbed simultaneously with bacterial adhesion as in life conditions. The two situations lead to very different antimicrobial behavior. Bacterial adhesion on preadsorbed BSA is very limited, while the simultaneous exposure of ODT SAM to proteins and bacteria lead to a markedly weaker antimicrobial effect. The combination of sum frequency generation spectroscopy and fluorescence confocal microscopy experiments allow one to draw conclusions on the factors that govern the ODT SAM or BSA film interaction with bacteria at the molecular level. On the hydrophobic ODT surface, interaction with hydrophobic or hydrophilic biomolecules results in opposite effects on the SAM, namely, a flattening or a raise of the terminal methyl groups of ODT. On an amphiphilic BSA layer, the bacterial adhesion strength is weakened by the negative charges carried by both BSA and bacteria. Surprisingly, preadsorbed BSA that cover part of the bacteria cell walls increase the adhesion strength to the BSA film and reduce hydrophobic interactions with the ODT SAM. Finally, bacterial adhesion on a BSA film is shown to modify the BSA proteins in some way that change their interaction with the ODT SAM. The antimicrobial effect is much stronger in the case of a preadsorbed BSA layer than when BSA and bacteria are in competition to colonize the ODT SAM surface.

Arrays of nanoelectromechanical biosensors functionalized by microcontact printing

The biofunctionalization of nanoelectromechanical systems (NEMS) is critical for the development of new classes of biosensors displaying improved performance and higher levels of integration. In this paper we propose a modified microcontact process (μCP) in order to biofunctionalize arrays of NEMS with a probe molecule on the active sensing areas together with an anti-fouling layer on the passive areas in a single, self-aligned step. We demonstrate the adequate functionalization/anti-fouling of arrays of freestanding nanocantilevers as dense as 105 nanostructures cm−2 by using both fluorescence microscopy and dynamic measurements of the structures’ resonant frequency. The proper bioactivity of an antibody deposited onto the cantilevers and the blocking property of a bovine serum albumin layer are both assessed by incubating specific and non-specific tagged secondary antibodies followed by fluorescence imaging. Furthermore, measurement of the resonant frequency of the nanocantilevers before and after functionalization and biological recognition demonstrate that using μCP for device functionalization does not damage the nanostructures and preserves the mechanical sensing capability of our NEMS.

Modeling colloid deposition on a protein layer adsorbed to iron-oxide-coated sand

Our recent study reported that conformation change of granule-associated Bovine Serum Albumin (BSA) may influence the role of the protein controlling colloid deposition in porous media (Flynn et al., 2012). The present study conceptualized the observed phenomena with an ellipsoid morphology model, describing BSA as an ellipsoid taking a side-on or end-on conformation on granular surface, and identified the following processes: (1) at low adsorbed concentrations, BSA exhibited a side-on conformation blocking colloid deposition; (2) at high adsorbed concentrations, BSA adapted to an end-on conformation promoted colloid deposition; and (3) colloid deposition on the BSA layer may progressively generate end-on molecules (sites) by conformation change of side-on BSA, resulting in sustained increasing deposition rates. Generally, the protein layer lowered colloid attenuation by the porous medium, suggesting the overall effect of BSA was inhibitory at the experimental time scale. A mathematical model was developed to interpret the ripening curves. Modeling analysis identified the site generation efficiency of colloid as a control on the ripening rate (declining rate in colloid concentrations), and this efficiency was higher for BSA adsorbed from a more dilute BSA solution.

Zwitterionic lipid (DPPC)–protein (BSA) complexes at the air–water interface

Complexation of zwitterionic lipid, dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and protein, bovine serum albumin (BSA) at the air–water interface has been studied by surface pressure (π) – mean molecular area (A) isotherms and X-ray reflectivity. Although BSA has isoelectric point nearly at pH≈4.8, possibility of complex formation with lipid molecules has been investigated from low (≈4.0) to high (≈9.0) pH range in presence of divalent cation, Ca2+ in the water subphase. Both the isotherm and reflectivity analysis show that the interaction of BSA with lipid monolayer takes place from that low to high subphase pH range, i.e., complexation occurs both below and above of the isoelectric point. Only one layer of BSA forms below the lipid monolayer and the probable reasons for such complex formation have been proposed.

Slow and remanent electric polarization of adsorbed BSA layer evidenced by neutron reflection

Using neutron reflectivity together with an appropriate electrochemical cell, we have studied the effects of transverse electric field on the Bovine Serum Albumin (BSA) monolayer initially adsorbed at the interface of the aqueous solution and a conductive doped-silicon wafer. Depending on the sign of the initial potential, a second layer is adsorbed on top of the first whereas a subsequent reversal of potential has no effect. We show that this behavior reveals the slow and remanent electric polarization of the first BSA layer and suggest an analogy with spin glasses based on the dipolar structure of this protein.

Plasma Polymerized Pyrrole Films for Biological Applications: Correlation Between Protein Adsorption Properties and Characteristics

AbstractThe present work describes the fabrication, characterization, and optimization of plasma polymerized pyrrole films, PPpy, prepared by continuous wave for application as the adhesion layers of bovine serum albumin (BSA). Plasma conditions used throughout the work produced significant differences in the chemical, physical, and electrochemical properties of the resultant PPpy films. Structural analysis and property characterizations of PPpy were carried out using Fourier transform infrared spectroscopy, X‐ray photoelectron spectroscopy, and electrochemical workstation. Surface plasmon resonance spectroscopy was used to investigate the adsorption kinetics of BSA onto PPpy films. Moreover, when immersed in solutions or after BSA adsorption, the differences of electron transfer at the interfaces between the PPpy and the electrolyte solutions were determined by electro impedance spectroscopy. While there is a clear dependence of the protein adsorption affinity on the plasma polymerization condition employed during the film preparation. The relationship between film structure, electrochemical properties, polymer stability in aqueous buffer, and protein binding affinities are discussed.magnified image

Microwave-assisted hydrothermal synthesis of biocompatible silver sulfide nanoworms

In this study, silver sulfide nanoworms were prepared via a rapid microwave-assisted hydrothermal method by reacting silver nitrate and thioacetamide in the aqueous solution of the Bovine Serum Albumin (BSA) protein. The morphology, composition, and crystallinity of the nanoworms were characterized by field emission scanning electron microscopy (FESEM), X-ray powder diffraction (XRD), transmission electron microscopy (TEM), selected area electron diffraction (SAED), X-ray energy dispersive spectroscopy (EDS), and Fourier transform infrared (FTIR) spectroscopy. The results show that the nanoworms were assembled by multiple adjacent Ag2S nanoparticles and stabilized by a layer of BSA attached to their surface. The nanoworms have the sizes of about 50 nm in diameter and hundreds of nanometers in length. The analyses of high-resolution TEM and their correlative Fast Fourier Transform (FFT) indicate that the adjacent Ag2S nanoparticles grow by misoriented attachment at the connective interfaces to form the nanoworm structure. In vitro assays on the human cervical cancer cell line HeLa show that the nanoworms exhibit good biocompatibility due to the presence of BSA coating. This combination of features makes the nanoworms attractive and promising building blocks for advanced materials and devices.

Effects of bovine serum albumin on the corrosion behaviour of AISI 316L, Co–28Cr–6Mo, and Ti–6Al–4V alloys in phosphate buffered saline solutions

The corrosion behaviour of AISI 316L, wrought Co–28Cr–6Mo and Ti–6Al–4V was studied in aerated solutions of phosphate buffered saline (PBS) at various concentrations of bovine serum albumin (BSA) at 37 °C. Open circuit potential, potentiodynamic polarization, linear polarization resistance (LPR) and electrochemical impedance spectroscopy (EIS) experiments along with X-ray photoelectron spectroscopy (XPS) on Co–28Cr–6Mo oxide layer were conducted to study the interaction of BSA and passive layers and to measure the corrosion rates. Ti–6Al–4V alloy had the lowest corrosion rate and the highest breakdown potential. It was shown that BSA has enhanced the alloy passive film stability at higher concentrations.

Optimization of a label‐free biosensor vertically characterized based on a periodic lattice of high aspect ratio SU‐8 nano‐pillars with a simplified 2D theoretical model

AbstractWe have recently demonstrated a biosensor based on a lattice of SU8 pillars on a 1 μm SiO2/Si wafer by measuring vertically reflectivity as a function of wavelength. The biodetection has been proven with the combination of Bovine Serum Albumin (BSA) protein and its antibody (antiBSA). A BSA layer is attached to the pillars; the biorecognition of antiBSA involves a shift in the reflectivity curve, related with the concentration of antiBSA. A detection limit in the order of 2 ng/ml is achieved for a rhombic lattice of pillars with a lattice parameter (a) of 800 nm, a height (h) of 420 nm and a diameter(d) of 200 nm. These results correlate with calculations using 3D‐finite difference time domain method. A 2D simplified model is proposed, consisting of a multilayer model where the pillars are turned into a 420 nm layer with an effective refractive index obtained by using Beam Propagation Method (BPM) algorithm. Results provided by this model are in good correlation with experimental data, reaching a reduction in time from one day to 15 minutes, giving a fast but accurate tool to optimize the design and maximizing sensitivity, and allows analyzing the influence of different variables (diameter, height and lattice parameter). Sensitivity is obtained for a variety of configurations, reaching a limit of detection under 1 ng/ml. Optimum design is not only chosen because of its sensitivity but also its feasibility, both from fabrication (limited by aspect ratio and proximity of the pillars) and fluidic point of view. (© 2011 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

In Vitro Coralling of a Transmembrane Protein in a Double Cushioned Lipid Bilayer Assembly

In this talk, the characteristic motion associated with a large transmembrane protein (5HT3A∗) in a Double Cushion biomimetic assembly, is an assembly that is passivated with a Bovine Serum Albumin layer along with PEG(poly-ethylene glycol) as the cushion will be presented. These results will be compared to receptors in both planar POPC assemblies and Single cushion assemblies. The main conclusion of this work is that while FRAP recoveries (percent bulk mobility) of these large membrane receptors are quite low in these systems, Single Particle Tracking (SPT) has revealed reasonably large mobile fractions when viewed at much higher spatial resolution and slower time scales. This is especially true for receptors in the ‘Double cushion’ assemblies. These assemblies display very low FRAP recoveries, but SPT clearly reveal that nearly all receptors are actually quite mobile but are spatially confined to very small corrals. The high mobility of receptors in these corralling domains demonstrates that the strong attractive interactions between the transmembrane protein and solid support have been virtually eliminated and therefore it is reasonable to believe that these receptors will remain active.

Dynamics of protein aggregation at the oil–water interface characterized by single molecule TIRF microscopy

Single molecule total internal reflectance fluorescence microscopy was used to observe the dynamic mechanisms of bovine serum albumin layer formation at the silicone oil–water interface. After an initial induction period, the mean diffusion coefficient of protein objects was observed to decrease as a function of exposure time. Simultaneously, the total adsorption rate was observed to decrease; both observations are consistent with protein layer formation. Importantly, the distribution of diffusion coefficients broadened systematically with exposure time, suggesting the presence of surface objects with a wide range of hydrodynamic radii ranging from a few nm (consistent with monomers) to ∼180 nm. The spatial distribution of the adsorption rate, which was initially uniform, gradually became inhomogeneous, also consistent with the existence of large aggregates. The kinetics of protein layer formation was dependent on the bulk protein concentration in the aqueous phase, and a kinetic population balance model was used to explain the nonlinear concentration dependence.

Single Particle Tracking Reveals Corralling of a Transmembrane Protein in a Double-Cushioned Lipid Bilayer Assembly

A predominate question associated with supported bilayer assemblies containing proteins is whether or not the proteins remain active after incorporation. The major cause for concern is that strong interactions with solid supports can render the protein inactive. To address this question, a large transmembrane protein, the serotonin receptor, 5HT(3A), has been incorporated into several supported membrane bilayer assemblies of increasing complexity. The 5HT(3A) receptor has large extracellular domains on both sides of the membrane, which could cause strong interactions. The bilayer assemblies include a simple POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine) supported planar bilayer, a “single-cushion” POPC bilayer with a PEG (poly(ethylene glycol)) layer between membrane and support, and a “double-cushion” POPC bilayer with both a PEG layer and a layer of BSA (bovine serum albumin). Single-cushion systems are designed to lift the bilayer from the surface, and double-cushion systems are designed to both lift the membrane and passivate the solid support. As in previously reported work, protein mobilities measured by ensemble fluorescence recovery after photobleaching (FRAP) are quite low, especially in the double-cushion system. But single-particle tracking of fluorescent 5HT(3A) molecules shows that individual proteins in the double-cushion system have quite high local mobilities but are spatially confined within small corralling domains (<r(C)2> 450 nm). Comparisons with the simple POPC membrane and the single-cushion POPC−PEG membrane reveal that BSA both serves to minimize interactions with the solid support and creates the corrals that reduce the long-range (ensemble averaged) mobility of large transmembrane proteins. These results suggest that in double-cushion assemblies proteins with large extra-membrane domains may remain active and unperturbed despite low bulk diffusion constants.

Adsorption and bioactivity studies of albumin onto hydroxyapatite surface

Bovine serum albumin (BSA) may have an inhibitory or promoter effect on hydroxyapatite (HA) nucleation when apatite is precipitated in a medium containing the protein. In this study we evaluated the influence of BSA on the precipitation of calcium phosphate phases (CP) from simulated body fluid (SBF) when the protein was previously bounded to HA surface. The kinetics of BSA immobilization onto hydroxyapatite surface was performed in different buffers and protein concentrations in order to adjust experimental conditions in which BSA was tightly linked to HA surface for long periods in SBF solution. It was shown that for BSA concentration higher than 0.1mg/mL the adsorption to HA surface followed Langmuir–Freundlich mechanisms, which confirmed the existence of cooperative protein–protein interactions on HA surface. Fourier Transformed Infrared Attenuated Total Reflectance Microscopy (FTIRM-ATR) evidenced changes in BSA conformational state in favor of less-ordered structure. Analyses from high resolution grazing incident X-ray diffraction using synchrotron radiation (GIXRD), Scanning Electron Microscopy (SEM) and Atomic Force Microscopy (AFM) showed that a poorly crystalline calcium phosphate was precipitated on the surface of HA discs coated with BSA, after the immersion in SBF for 4 days. The new bioactive layer had morphological characteristics similar to the one formed on the HA surface without protein. It was identified as a carbonated apatite with preferential crystal growth along apatite 002 direction. The GIXRD results also revealed that BSA layer bound to the surface inhibited the HA dissolution leading to a reduction on the formation of new calcium phosphate phase.

Effect of pH on the Formation of a Bovine Serum Albumin Layer on a Poly(stryren-co-maleic Acid) Surface

The layer formation of bovine serum albumin (BSA) on a poly(styrene-co-maleic acid) (PSMA) surface was investigated by using quartz crystal microbalance (QCM) technique at various pH values. The formation of a BSA surface was examined by atomic force microscopy (AFM). To study the effect on the layer formation, the pH of solution was varied from 2 to 7.4 while the concentration of BSA was in the range of 0.01 to 5 mg/ml during the layer absorption. It was found that the BSA adsorption strongly depends on the pH of solution, and the concentration of BSA. The absorption layer occurred maximum at the pH value of 3.5 which resulted from the charge of PSMA and BSA molecules. The layer formation reached the saturate value at the concentration higher than 3 mg/ml. The molecular packing of the BSA layer at different pH values was determined by AFM and total mass change of QCM.