Bovine Serum Albumin Monolayer Research Articles

Effect of different valent ions (Na+, Ca2+ & Y3+) on structural and morphological features of protein (BSA) thin films adsorbed on hydrophobic silicon (H-Si) surface

Structure and morphology of bovine serum albumin (BSA) films adsorbed on hydrophobic silicon (H-Si) surface from BSA solution in presence of different cationic ions (Na+, Ca2+ and Y3+) are studied. Electron density profile as obtained from X-ray reflectivity analysis shows that ≈ 55 Å thick monolayer of pure BSA is adsorbed on H-Si surface, while atomic force microscopy image displays its globular morphology. A bilayer of globular BSA molecules of average thickness ≈ 85 Å is formed on the H-Si surface at 1–10 mM of Na+ and Ca2+ ions. However, Y3+ ions favor the immobilization of a tri-layer of BSA on H-Si surface except for 5 mM, at which a tetralayer is formed due to the effect of re-entrant condensation of BSA solution. Elemental mapping analysis as obtained from scanning electron microscopy confirms that the attachment of Y3+ ions is higher in comparison with Na+ and Ca2+ ions due to maximum charge density of hydrated Y3+ ions. The contact angle results display intermediate hydrophobicity for Na+ ions, whereas minimum and maximum hydrophobicity for Ca2+ and Y3+ ions interacted protein films, respectively. It is found that the adsorption of BSA is relatively higher in presence of all ions on hydrophobic surface than hydrophilic. All the adsorbed BSA films are found mostly stable with time.

Effects of Macromolecular Crowding on Nanoparticle Diffusion: New Insights from Mössbauer Spectroscopy.

In this work, we used Mössbauer spectroscopy as a new approach for experimental quantification of the self-diffusion coefficient (DMössbauer) and hydrodynamic (HD) size of iron-containing nanoparticles (NPs) in complex crowded solutions, mimicking cell cytoplasm. As a probe, we used 9 nm cobalt ferrite NPs (CFNs) dispersed in solutions of bovine serum albumin (BSA) with a volume fraction (φBSA) of 0-0.2. Our results show that the broadening of Mössbauer spectra is highly sensitive to the diffusion of CFNs, while when φBSA = 0.2, the CFN-normalized diffusivity is reduced by 86% compared to that of a protein-free solution. CFN colloids were also studied by dynamic light scattering (DLS). Comparison of the experimental data shows that DLS significantly underestimates the diffusion coefficient of CFNs and, consequently, overestimates the HD size of CFNs at φBSA > 0, which cannot be attributed to the formation of the BSA monolayer on the surface of CFNs.

Conductance Changes in Bovine Serum Albumin Caused by Drug-Binding Triggered Structural Transitions.

Proteins, due to their binding selectivity, are promising candidates for fabricating nanoscale bio-sensors. However, the influence of structural change on protein conductance caused by specific protein-ligand interactions and disease-induced degeneration still remains unknown. Here, we excavated the relationship between circular dichroism (CD) spectroscopy and conductive atomic force microscopy (CAFM) to reveal the effect of the protein secondary structures changes on conductance. The secondary structure of bovine serum albumin (BSA) was altered by the binding of drugs, like amoxicillin (Amox), cephalexin (Cefa), and azithromycin (Azit). The CD spectroscopy shows that the α-helical and β-sheet content of BSA, which varied according to the molar ratio between the drug and BSA, changed by up to 6%. The conductance of BSA monolayers in varying drug concentrations was further characterized via CAFM. We found that BSA conductance has a monotonic relation with α-helical content. Moreover, BSA conductance seems to be in connection with the binding ability of drugs and proteins. This work elucidates that protein conductance variations caused by secondary structure transitions are triggered by drug-binding and indicate that electrical methods are of potential application in protein secondary structure analysis.

The Cooperative Effect of BSA Langmuir Monolayers and Magnesium Ions on Calcium Carbonate Crystallization

The crystallization of calcium carbonate at interface of Langmuir monolayers of bovine serum albumin and subphase containing magnesium ions was studied in this paper. The results were characterized by using powder X-ray diffraction (XRD) and scanning electron microscopy (SEM). The effect rules were obtained by the cooperation of bovine serum albumin Langmuir monolayers and magnesium ions. BSA Langmuir monolayers controlled calcium carbonate to magnesium ions in solution. The experiment results showed that in the presence of both BSA Langmuir monolayers at interface and magnesium ions in solutions, an orientation aragonite with regular spherical morphology was precipitated. It is indicated that BSA Langmuir monolayers and magnesium ions have a cooperative effect on controlling the polymorph and orientation of calcium carbonate crystal. The experiments suggested that BSA Langmuir monolayer acts in combination with magnesium ions to inhibit calcite crystal growth, while favoring the formation of aragonite crystals.

Fabrication of long range surface plasmon waveguide biosensors in a low-index fluoropolymer

The authors report the fabrication of long-range surface plasmon polariton biosensors consisting of thin narrow Au stripes embedded in a low refractive index fluoropolymer with etched fluidic channels. The fabrication process incorporates a sacrificial SiO2 channel etch stop layer to avoid waveguide damage and prevent channel over-etching, and an Al etch mask to minimize thermal cracking of the fluoropolymer during channel etching. Process details are reported along with fabrication results, and the optical and biosensing performance of the fabricated devices are demonstrated. The biosensors were tested for bulk sensitivity and bovine serum albumin (BSA) protein adsorption producing notable changes for solutions with different refractive indices (increment of 2 × 10−3) and a change in insertion loss of 0.1 dB as a monolayer of BSA forms on the device.

Morphology Effect on Charge Transport in Doped Bovine Serum Albumin Self-Assembled Monolayers

Direct exploration of the mutually interfering morphological and charge-transport characteristics in self-assembled monolayers (SAMs) is reported. These strongly coupled properties are addressed by means of surface spectroscopy techniques, combined so as to consistently account at high sensitivity for a broad range of surface properties without any use of a top contact. Applied to doped bovine serum albumin (BSA) SAMs, we show how the BSA conformation, its dehydration, and monolayer assembly are all correlated. Moreover, the electrical properties, transport, and charge trapping are highly affected by the SAM compositional and structural state, with a specific role of water molecules. Our results reveal and further demonstrate a useful approach to the complex challenges presented by (bio) molecular electronics.

Facile synthesis and performance studies of BSA and PDA@Ag hollow microcapsules using SiO2 microspheres as the templates

This paper describes a general strategy that uses dopamine as a stable anchor to present protein molecules on the surface of silicon microspheres via self-assembled method (SAM), and after the removal of templates, the bovine serum albumin (BSA) microcapsules were successfully prepared. Two self assembling methods are described here, one is using glutaraldehyde (GA) as crosslinking agent to modify multilayer protein on the surface of silicon microspheres; In another way, dopamine is used as a anchor layer to graft the BSA monolayers onto silicon templates. The BSA coated silicon microspheres and BSA hollow microcapsules were characterized by transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM). The biocompatibility of the BSA coated silicon microspheres was evaluated in vitro by MTT method. Then the polydopamine (PDA) hollow microcapsules was prepared using SiO2 nanoparticles as the templates, and Ag namoparticles was then modified on the PDA capsules. The catalytic performance of SiO2@ PDA@Ag microspheres for methylene blue (MB) was studied. And the Escherichia coli and Staphylococcus aureus was used as the indicator bacteria to investigate the antimicrobial activity of the obtained SiO2@ PDA@Ag microspheres.

Characterization of adsorption and lubrication of synovial fluid proteins and HA on DLC joint bearings surface

Adsorption and competitive adsorption behavior of bovine serum albumin (BSA), human γ-globulin (HGG), and hyaluronic acid (HA) molecules on a diamond-like carbon (DLC) film surface were investigated by the means of a quartz crystal microbalance with dissipation (QCM-D), and antigen-antibody reactions. The results showed that a BSA monolayer could be formed on the DLC film surface with side-on orientation and that an HGG monolayer could be formed on the DLC film surface with end-on orientation by using pure BSA or HGG solutions, respectively. HA molecules were hardly adsorbed onto the surface of DLC film or HGG or BSA biomolecular layer. The results of competitive adsorption showed that HGG molecules could displace BSA molecules and that HGG was the dominantly adsorbed molecule on the DLC film surface in a mixture of BSA, HGG, and HA. The frictional behavior of a DLC film sliding against Al2O3 in a mixture of BSA, HGG, and HA, was investigated by means of the friction coefficient and an open circuit potential, which suggested that the HGG molecule was the main component that formed the lubricating film.

Albumin-coated SPIONs: an experimental and theoretical evaluation of protein conformation, binding affinity and competition with serum proteins.

The variety of nanoparticles (NPs) used in biological applications is increasing and the study of their interaction with biological media is becoming more important. Proteins are commonly the first biomolecules that NPs encounter when they interact with biological systems either in vitro or in vivo. Among NPs, super-paramagnetic iron oxide nanoparticles (SPIONs) show great promise for medicine. In this work, we study in detail the formation, composition, and structure of a monolayer of bovine serum albumin (BSA) on SPIONs. We determine, both by molecular simulations and experimentally, that ten molecules of BSA form a monolayer around the outside of the SPIONs and their binding strength to the SPIONs is about 3.5 × 10(-4) M, ten times higher than the adsorption of fetal bovine serum (FBS) on the same SPIONs. We elucidate a strong electrostatic interaction between BSA and the SPIONs, although the secondary structure of the protein is not affected. We present data that supports the strong binding of the BSA monolayer on SPIONs and the properties of the BSA layer as a protein-resistant coating. We believe that a complete understanding of the behavior and morphology of BSA-SPIONs and how the protein interacts with SPIONs is crucial for improving NP surface design and expanding the potential applications of SPIONs in nanomedicine.

Structure of bovine serum albumin adsorbed on silica investigated by quartz crystal microbalance

Abstract Adsorption of bovine serum albumin (BSA) on the surface of SiO 2 was investigated mainly using quartz crystal microbalance (QCM) with energy dissipation mode. BSA adsorption kinetics and coverage were determined as a function of pH of the solution with the ionic strength of 1 × 10 −2 M. The maximum coverage was obtained in the pH range 4.5–5.4, near the protein isoelectric point. For pH larger than 5.4 and smaller than 4.5 a monotonic decrease of the adsorbed mass was observed. The images obtained using atomic force microscopy (AFM) method revealed various BSA structures depending on the pH at which they were adsorbed. The identified conformations were assigned to the compact triangular form with an effective length L ef = 8.3 nm and the elongated one with length L ef = 26.7 nm. The dependence between the structure of the protein layers formed on the surface and BSA conformations in solution was additionally followed by contact angle (CA) measurements. It turned out that CA values are very sensitive to the structure of BSA monolayers formed on the silica surface. Comparison of QCM and AFM data with those obtained by other indirect methods (CA) additionally enabled interpretation of the dependence of BSA adsorption on the pH of the solution. The obtained results confirmed the significant role of the highly anisotropic surface charge distribution of BSA molecules on the process of their adsorption.

Coat formation of surface-active proteins on aqueous surfaces during drying

Segregation of the protein bovine serum albumin (BSA) and lactose in thin aqueous films during drying was investigated by examining the composition of the dried films using inverse micro Raman spectroscopy (IMRS) and X-ray photoelectron spectroscopy (XPS) sputter-depth profiling. The composition was uniform through the thickness of the dried films except within a 10nm region at the exposed surface where BSA had accumulated, most likely due to its surface activity. The thickness of the BSA layer was similar to the diameter of a BSA molecule, which suggests that a single monolayer of BSA adsorbed at the exposed surface. The BSA surface concentration of the dried films was constant over a wide range of BSA bulk concentrations, indicating that the aqueous surface became saturated with BSA during drying. The BSA surface layer of order 10nm was significantly thinner than the film thickness of order 10μm, which implies that BSA formed a surface coating rather than a shell, and thus lent no structural rigidity to the film.

Effects of Protein Concentration and pH on Oil Droplet Size in O/W Emulsions Stabilized by Bovine Serum Albumin

Here, we report the effect of protein concentration on oil droplet size and adsorbed protein density in oil-in-water emulsions stabilized by bovine serum albumin (BSA) under various pH conditions. The droplet size changed with BSA concentration in a different way depending on pH although it eventually converged on the same value (ca. 15 µm) irrespective of pH. It was suggested that the adsorbed BSA monolayer at low and high pH could not effectively protect oil droplets against coalescence during homogenization.

Interaction and diffusion of gold nanoparticles in bovine serum albumin solutions

We studied the interactions of small sized, radius of 2.5–10 nm, gold nanoparticles (AuNPs) with bovine serum albumin (BSA). Fluctuation correlation spectroscopy (FCS) was used to monitor the changes in the Brownian diffusion of these NPs. Our results indicate a BSA monolayer formation at the NP surface with a thickness of 3.8 nm. The thickness of the adsorbed layer was independent of NP size. Best fit was obtained by the anticooperative binding model with the Hill coefficient of n = 0.63. Dissociation constant (KD) increases with particle size indicating stronger interaction of BSA with smaller sized NPs.

Biosensing using straight long-range surface plasmon waveguides

Straight long-range surface plasmon waveguides are demonstrated as biosensors for the detection of cells, proteins and changes in the bulk refractive index of solutions. The sensors consist of 5 μm wide 22 nm thick Au stripes embedded in polymer (CYTOP™) with microfluidic channels etched into the top cladding. Bulk sensing is demonstrated by sequentially injecting six solutions of different refractive indices in 2 × 10(-3) RIU increments; such index steps were detected with a signal-to-noise ratio of ~1000. Selective capture of cells is demonstrated using Au waveguides functionalized with antibodies against blood group A, and red blood cells of group A and O in buffer as positive and negative analyte. Bovine serum albumin in buffer was used to demonstrate protein sensing. A monolayer of bovine serum albumin physisorbed on a carboxyl-terminated self-assembled monolayer on Au was detected with a signal-to-noise ratio of ~300. Overall, the biosensor demonstrated a good capability for detecting bulk changes in solution and for sensing analyte over a very wide range of mass (from cells to proteins). The biosensors are compact, inexpensive to fabricate, and may find use over a wide range of cost-sensitive sensing and detection applications.

In Situ Measurement of Bovine Serum Albumin Interaction with Gold Nanospheres

We present in situ observations of adsorption of bovine serum albumin (BSA) on citrate-stabilized gold nanospheres. We implemented scattering correlation spectroscopy as a tool to quantify changes in the nanoparticle brownian motion resulting from BSA adsorption onto the nanoparticle surface. Protein binding was observed as an increase in the nanoparticle hydrodynamic radius. Our results indicate the formation of a protein monolayer at similar albumin concentrations as those found in human blood. Additionally, by monitoring the frequency and intensity of individual scattering events caused by single gold nanoparticles passing the observation volume, we found that BSA did not induce colloidal aggregation, a relevant result from the toxicological viewpoint. Moreover, to elucidate the thermodynamics of the gold nanoparticle-BSA association, we measured an adsorption isotherm which was best described by an anticooperative binding model. The number of binding sites based on this model was consistent with a BSA monolayer in its native state. In contrast, experiments using poly(ethylene glycol)-capped gold nanoparticles revealed no evidence for adsorption of BSA.

Ultrathin Chitin Films for Nanocomposites and Biosensors

Chitin is the second most abundant biopolymer and insight into its natural synthesis, enzymatic degradation, and chemical interactions with other biopolymers is important for bioengineering with this renewable resource. This work is the first report of smooth, homogeneous, ultrathin chitin films, opening the door to surface studies of binding interactions, adsorption kinetics, and enzymatic degradation. The chitin films were formed by spincoating trimethylsilyl chitin onto gold or silica substrates, followed by regeneration to a chitin film. Infrared and X-ray photoelectron spectroscopy, X-ray diffraction, ellipsometry, and atomic force microscopy were used to confirm the formation of smooth, homogeneous, and amorphous chitin thin films. Quartz crystal microbalance with dissipation monitoring (QCM-D) solvent exchange experiments showed these films swelled with 49% water by mass. The utility of these chitin films as biosensors was evident from QCM-D and surface plasmon resonance studies that revealed the adsorption of a bovine serum albumin monolayer.

Tartrazine Dye and Bovine Serum Albumin: the Influence of pH on Adsorption Process

Multilayers from Tartrazine dye (Tart) alternated with bovine serum albumin (BSA) were successfully prepared and a uniform growth has been observed. The influence of two values of Tart solution pH (3 and 7.5) on adsorption process of this molecule on BSA monolayer at physiological pH was investigated using kinetics and isotherms of adsorption, as well as thermostimulated adsorption experiments. All effects found by the changing of the pH were explicated mainly taking into account the ionic attraction (Tart at pH 3) and ionic repulsion (Tart at pH 7.5) that occurs during the adsorption process of Tart onto BSA monolayer.

Savinase action on bovine serum albumin (BSA) monolayers demonstrated with measurements at the air–water interface and liquid Atomic Force Microscopy (AFM) imaging

We studied the enzymatic action of Savinase on bovine serum albumin (BSA) organized in a monolayer spread at the air/water interface or adsorbed at the mica surface. We carried out two types of experiments. In the first one we followed the degradation of the protein monolayer by measuring the surface pressure and surface area decrease versus time. In the second approach we applied AFM imaging of the supported BSA monolayers adsorbed on mica solid supports and extracted information for the enzyme action by analyzing the obtained images of the surface topography in the course of enzyme action. In both cases we obtained an estimate for the turnover number (TON) of the enzyme reaction.

Crystallization and self-assembly of calcium carbonate under albumin Langmuir monolayers

It is an interesting phenomenon for natural organisms to have control over the shape and size of inorganic nanocrystals and to arrange them into polycrystalline structures. This phenomenon has been driving many attempts to mimic the biomineralization process for synthesizing novel materials. In the present work, novel polycrystalline spindle-like calcite crystals consisting of nanocrystals have been synthesized via a phase transformation process under a Langmuir monolayer of bovine serum albumin (BSA) at room temperature. The polycrystals are composed of hundreds of well assembled calcite nanoneedles consisting of orderly oriented and aggregated nanocrystals transformed from amorphous phase. The evidence for the phase transformation process has been observed in detail by structural characterization of the products at different growth stages. It has been found that the products are evolved from amorphous particles to spindle-like polycrystalline particles. The mineralization process and the interaction between the inorganic and bioorganic components are discussed in relation to relevant protein-mediated nucleation models of biomineralization. Hopefully, the present research is to help understanding the protein-directed formation process of complex structures as well as biomineralization mechanism.

Nanogel surface coatings for improved single-molecule imaging substrates

Surfaces that resist protein adsorption are important for many bioanalytical applications. Bovine serum albumin (BSA) coatings and multi-arm poly(ethylene glycol) (PEG) coatings display low levels of non-specific protein adsorption and have enabled highly quantitative single-molecule (SM) protein studies. Recently, a method was developed for coating a glass with PEG-BSA nanogels, a promising hybrid of these two low-background coatings. We characterized the nanogel coating to determine its suitability for SM protein experiments. SM adsorption counting revealed that nanogel-coated surfaces exhibit lower protein adsorption than covalently coupled BSA surfaces and monolayers of multi-arm PEG, so this surface displays one of the lowest degrees of protein adsorption yet observed. Additionally, the nanogel coating was resistant to DNA adsorption, underscoring the utility of the coating across a variety of SM experiments. The nanogel coating was found to be compatible with surfactants, whereas the BSA coating was not. Finally, applying the coating to a real-world study, we found that single ligand molecules could be tethered to this surface and detected with high sensitivity and specificity by a digital immunoassay. These results suggest that PEG-BSA nanogel coatings will be highly useful for the SM analysis of proteins.