Removal Of Bovine Serum Albumin Research Articles

Microplasma jet treatment of bovine serum albumin coatings for controlling enzyme and cell attachment

We investigated a new approach to control protein and cell attachment inside 96-well polystyrene plates. The wells were first coated with bovine serum albumin (BSA) to inhibit cell and protein attachment. The BSA-coated wells were then treated with a helium microplasma jet for increasing times that resulted in gradual removal of BSA from the surface. It was found that the amount of enzyme and cell attachment could be controlled in the wells where BSA was only partially removed by the microplasma jet. In addition to the surface coverage of BSA, the new surface chemistry induced by the microplasma jet treatment also had an important role in the control of enzyme and cell attachment. In summary, microplasma jet treatment of BSA-coated polystyrene wells is a simple and effective method for controlling enzyme and cell attachment. This might find use for high-throughput screening of new cell culture platforms where control over the level protein, enzyme or cell adherence is needed in order to maintain a specific cell function.

Removal of bovine serum albumin from wastewater using fouling resistant ultrafiltration membranes based on the blends of cellulose acetate, and PVP-TiO2 nanoparticles

Fouling resistant ultrafiltration membranes based on the blends of polyvinylpyrrolidone (PVP), TiO2 nanoparticles and cellulose acetate, CA-PVP-TiO2 (CATP), for removal of bovine serum albumin (BSA) were prepared by using phase inversion process. The influences of PVP and TiO2 on the preparation of phase inverted cellulose acetate (CA) ultrafiltration membrane were explored in terms of morphology study, equilibrium water content (EWC), hydraulic resistance, permeability performance, hydrophilicity, and thermal stability. After the introduction of PVP and TiO2 to the ternary (polymer-solvent-non-solvent) system, the formations of finger-like structures and macro-voids were reduced significantly. An improvement in porosity, average pore size, and hydrophilic nature of the CA membranes were detected after the introduction of PVP and TiO2 into the polymer matrix. The interaction between TiO2 and CA was confirmed and the degradation temperature of the CA membrane was significantly improved. BSA protein removal efficiency, anti-fouling performance, and recycling potential of the UF membranes were investigated. The CATP membrane (10.5 wt % CA: 4 wt % PVP: 2 wt % TiO2) has displayed high BSA removal efficiency and flux recovery ratios (NFR) with enhanced anti-fouling performances for the three fouling/rinsing cycles.

Efficient separation of mannan–protein mixtures by ionic liquid aqueous two-phase system, comparison with lectin affinity purification

Analysis of carbohydrates from complex biological samples often requires their isolation from proteins and other contaminants to avoid interference. An effective separation of mannan–protein mixtures by 1-butyl-3-methylimidazolium bromide/K2HPO4 ionic liquid aqueous two-phase system (IL-APTS) is reported. Extraction efficiency of bovine serum albumin (BSA) ranged from 92% to 97% while extraction efficiency of mannan reached values from 95% to about 100% depending on phase and/or model sample composition. On the contrary, lower efficiency of BSA removal (73–84%) was recorded for lectin affinity purification with concanavalin A–triazine bead cellulose (Con A-TBC); the low mannan-binding capacity was limiting factor here. The size exclusion chromatography pattern of model mannan–BSA samples after both IL-APTS and Con A-TBC treatments were consistent with the spectrophotometric component analysis. In case of biological experiment, the ionic liquid separation technique was superior in pre-purification of 2-aminobenzamide-labelled mannan from cell culture medium prior to HPLC-FLD analysis.

In-situ monitoring of etching of bovine serum albumin using low-temperature atmospheric plasma jet

Bio-decontamination of surfaces by means of atmospheric pressure plasma is nowadays extensively studied as it represents promising alternative to commonly used sterilization/decontamination techniques. The non-equilibrium atmospheric pressure plasmas were already reported to be highly effective in removal of a wide range of biological residual from surfaces. Nevertheless the kinetics of removal of biological contamination from surfaces is still not well understood as the majority of performed studies were based on ex-situ evaluation of etching rates, which did not allow investigating details of plasma action on biomolecules. This study therefore presents a real-time, in-situ ellipsometric characterization of removal of bovine serum albumin (BSA) from surfaces by low-temperature atmospheric plasma jet operated in argon. Non-linear and at shorter distances between treated samples and nozzle of the plasma jet also non-monotonic dependence of the removal rate on the treatment duration was observed. According to additional measurements focused on the determination of chemical changes of treated BSA as well as temperature measurements, the observed behavior is most likely connected with two opposing effects: the formation of a thin layer on the top of BSA deposit enriched in inorganic compounds, whose presence causes a gradual decrease of removal efficiency, and slight heating of BSA that facilitates its degradation and volatilization induced by chemically active radicals produced by the plasma.

Utilization of Electrospun Polystyrene Membranes as a Preliminary Step for Rapid Diagnosis

Recent advances in clinical practice drive deoxyribonucleic acid (DNA) as an important class of biomarker. Monitoring the change in their concentration suggests the initiation and/or progression of various disorders. However, low quantity of DNA biomarkers in body fluids requires a delicate isolation methodology that provides efficient separation and easy handling. This study describes a newer‐generation separation technology relying on electrospun fibers of sub‐micrometer diameter of a commodity polymer for DNA biomarkers in simulative serum. Fibrous polystyrene membranes are prepared by electrospinning and they are subjected to post‐modification with Au. The composite membranes may provide a convenient environment for the removal of bovine serum albumin (BSA) from BSA and DNA mixtures. The eluent can be used as an efficient tool for detection of DNA biomarkers associated with diagnosis of numerous life‐threatening diseases. image

Short- and long-term outcomes of the absence of protein during bovine blastocyst formation in vitro.

In cattle, individual in vitro embryo culture after Day 6 benefits development, allowing non-invasive analysis of culture medium. However, undefined supplements in culture reduce analytical reliability. In this study we assayed the short- and long-term performance of embryos after bovine serum albumin removal over a 24-h period in individual culture. The absence of protein decreased embryo development and cell counts in the inner cell mass without affecting blastocyst sex ratio. However, the absence of protein produced embryos with an improved tendency to survive vitrification after 24h in culture (P=0.07). After transfer to recipients, birth rates of embryos that had been cultured with protein tended to decrease (P<0.06) mostly as a result of a higher number of miscarriages (P<0.013), reflecting lower viability. Birthweight, gestation length, height and thorax circumference did not differ between embryos cultured with or without protein. In fresh blastocysts cultured without protein, gene expression analysis showed higher abundance (P<0.05) of insulin-like growth factor 2 receptor (IGF2R; imprinting) and activating transcription factor 4 (ATF4) and DNA-damage-inducible transcript 3 (DDIT3; endoplasmic reticulum stress) transcripts, with DNA methyltransferase 3A (DNMT3A; imprinting) tending to increase (P=0.062). However, in hatched blastocysts that survived cryopreservation, glucose-6-phosphate dehydrogenase (G6PD) was overexpressed in embryos cultured without protein (P<0.01). The absence of protein results in fewer blastocysts but improved long-term viability after cryopreservation.

Enhancement and Mitigation Mechanisms of Protein Fouling of Ultrafiltration Membranes under Different Ionic Strengths.

To determine further the enhancement and mitigation mechanisms of protein fouling, filtration experiments were carried out with polyvinylidene fluoride (PVDF) ultrafiltration (UF) membranes and bovine serum albumin (BSA) over a range of ionic strengths. The interaction forces, the adsorption behavior of BSA on the membrane surface, and the structure of the BSA adsorbed layers at corresponding ionic strengths were investigated. Results indicate that when the ionic strength increased from 0 to 1 mM, there was a decrease in the PVDF-BSA and BSA-BSA electrostatic repulsion forces, resulting in a higher deposition rate of BSA onto the membrane surface, and the formation of a denser BSA layer; consequently, membrane fouling was enhanced. However, at ionic strengths of 10 and 100 mM, membrane fouling and the BSA removal rate decreased significantly. This was mainly due to the increased hydration repulsion forces, which caused a decrease in the PVDF-BSA and BSA-BSA interaction forces accompanied by a decreased hydrodynamic radius and increased diffusion coefficient of BSA. Consequently, BSA passed more easily through the membrane and into permeate. There was less accumulation of BSA on the membrane surface. A more nonrigid and open structure BSA layer was formed on the membrane surface.

Protein-templated biomimetic silica nanoparticles.

Biomimetic silica particles can be synthesized as a nanosized material within minutes in a process mimicked from living organisms such as diatoms and sponges. In this work, we have studied the effect of bovine serum albumin (BSA) as a template to direct the synthesis of silica nanoparticles (NPs) with the potential to associate proteins on its surface. Our approach enables the formation of spheres with different physicochemical properties. Particles using BSA as a protein template were smaller (∼250-380 nm) and were more monodisperse than those lacking the proteic core (∼700-1000 nm) as seen by dynamic light scattering (DLS), scanning electron microscopy (SEM), and environmental scanning electron microscopy (ESEM) analysis. The absence of BSA during synthesis produced silica nanoparticles without any porosity that was detectable by nitrogen adsorption, whereas particles containing BSA developed porosity in the range of 4 to 5 nm which collapsed on the removal of BSA, thus producing smaller pores. These results were in accordance with the pore size calculated by high-resolution transmission electron microscopy (HTEM). The reproducibility of the BSA-templated nanoparticle properties was determined by analyzing four batches of independent synthesizing experiments that maintained their properties. The high positive superficial charge of the nanoparticles facilitated adsorption under mild conditions of a range of proteins from an E. coli extract and a commercial preparation of laccase from Trametes versicolor. All of the proteins were quantitatively desorbed. Experiments conducted showed the reusability of the particles as supports for the ionic adsorption of the biomolecules. The protein loading capacity of the BSA-based biomimetic particles was determined using laccase as 98.7 ± 6.6 mg·g(-1) of particles.

Biosorption of bovine serum albumin by Ulva lactuca biomass from industrial wastewater: Equilibrium, kinetic and thermodynamic study

Batch biosorption experiments have been carried out for the removal of bovine serum albumin (BSA) from simulated industrial wastewater onto Ulva lactuca seaweed. Various vital parameters influencing the biosorption process such as initial concentration of BSA, pH of the solution, adsorbent dosage and temperature have been determined. The biosorption kinetics follows a pseudo-second order kinetic model. Equilibrium isotherm studies demonstrate that the biosorption followed the Freundlich isotherm model, which implies a heterogeneous sorption phenomenon. Various thermodynamic parameters such as changes in enthalpy, free energy and entropy have been calculated. The positive value of ΔH° and the negative value of ΔG° show that the sorption process is endothermic and spontaneous. The positive value of change in entropy ΔS° shows increased randomness at the solid–liquid interface during the biosorption of BSA onto U. lactuca seaweed.

Selective precipitation‐assisted recovery of immunoglobulins from bovine serum using controlled‐fouling crossflow membrane microfiltration

Efficient and economic recovery of immunoglobulins (Igs) from complex biological fluids such as serum, cell culture supernatant or fermentation cell lysate or supernatant, represents a substantial challenge in biotechnology. Methods such as protein A affinity chromatography and anion exchange chromatography are limited by cost and selectivity, respectively, while membrane chromatography is limited by low adsorptive area, flow distribution problems and scale-up difficulties. By combining the traditional salt-assisted precipitation process for selective removal of Igs from serum followed by constant-permeate flux membrane microfiltration for low fouling, we demonstrate an exciting new, efficient and economic hybrid method. The high selectivity of an ammonium sulfate-induced precipitation step was used to precipitate the Igs leaving the major undesirable impurity, the bovine serum albumin (BSA), in solution. Crossflow membrane microfiltration in diafiltration mode was then employed to retain the precipitate, while using axial flow rates to optimize removal of residual soluble BSA to the permeate. The selectivity between immunoglobulin G (IgG) and BSA obtained from the precipitation step was approximately 36, with 97% removal of the BSA with diafiltration in 5 diavolumes with resulting purity of the IgG of approximately 93% after the membrane microfiltration step. Complete resolubilization of the IgG was obtained without any aggregation at the concentrations of ammonium sulfate employed in this work. Further, membrane pore size and axial Reynolds number (recirculation rate) were shown to be important for minimizing fouling and loss of protein precipitate.

Removal of bovine serum albumin using solid-phase extraction with in-situ polymerized stationary phase in a microfluidic device

Serum albumin, one of the most abundant serum proteins, blocks the expression of other important biomarkers. The objective of this study is to remove serum albumin effectively by using solid-phase extraction (SPE) in microfluidic devices. Photo-polymerized adsorbent as a stationary phase of SPE was used to remove bovine serum albumin (BSA). The adsorption capacity was examined with the effect of pH and concentration in BSA solution, and adjustment of monomer concentration such as hydrophilic 2-acrylamido-2-methyl-1-propanesulfonic acid and acrylamide in the adsorbent. The effect of hydrophobic butyl methacylate on BSA adsorption was also studied. Selective removal in a bicomponent with BSA and bovine γ-globulin was performed by adjusting the pH as required.

Cleaning of albumin‐contaminated Ti and Cr surfaces: an XPS and QCM study

AbstractThe cleaning of protein‐contaminated metallic surfaces is a major challenge for non‐disposable devices used for biomedical applications and in the food industry. The objective of this work is to estimate the cleaning efficiency of sodium dodecyl sulfate (SDS) buffered at pH 7.4, as a function of temperature, for an immersion time of 15 h. Bovine serum albumin (BSA) is pre‐adsorbed on pure Ti‐ and Cr‐deposited quartz crystals. BSA adsorption and removal were investigated by X‐ray photoelectron spectroscopy (XPS) and quartz crystal microbalance (QCM). Before cleaning, the amount of adsorbed BSA corresponds to one monolayer, and the oxide thickness is about 5 nm for the Cr quartz (Cr3+ in oxide and hydroxide). For Ti quartz, the metal is not detected (10 nm or more of Ti4+ in TiO2). The C 1s and N 1s peaks indicate that the cleaning with SDS is achieved at 37 °C for Ti. For Cr, there are no significant changes from 37 to 60 °C. At 80 °C, the surface is partially cleaned (70% efficiency); at 90 °C, the surface cleaning with SDS is achieved (90% efficiency). The oxide layer of the chromium sample has not changed in composition and thickness.QCM measurements were performed at 37 °C. Switches between BSA and SDS solutions indicate that for Ti, SDS induces partial BSA removal (40% efficiency) after ∼20 min, while for Cr it seems that SDS adsorbs reversibly on the BSA layer, without any cleaning effect. Thus, QCM results are in agreement with XPS data obtained at the same temperature. Copyright © 2008 John Wiley &amp; Sons, Ltd.

Improvement of the Cleanability of Pseudo-Sensitized Stainless Steel Surfaces by Solution Treatment

Type 316L stainless steel particles were pseudo-sensitized by being heated at 700 degrees C for 100 h under a reduced pressure of 1.3 x 10(-3) Pa. Pseudo-sensitization treatment resulted in the formation of chromium-rich precipitates at the outermost surfaces of the stainless steel particles. The curve for the apparent surface charge density (sigma app) of the pseudo-sensitized particles showed a more basic character compared with the curve for the sigma app of the original particles. There was no significant difference between the amounts of bovine serum albumin (BSA) adsorbed on the original and the pseudo-sensitized particles. On the contrary, the efficiency of BSA removal from the pseudo-sensitized particles during batch-wise and continuous cleaning operations with 0.1M NaOH solution were significantly smaller than that for the original particles. It was indicated that the extremely lowered cleanability of the pseudo-sensitized particles was caused by a marked increase in the positive sigma app values after the pseudo-sensitization. Solution treatment of the pseudo-sensitized particles at 1,050 degrees C for 1 h under a reduced pressure of 1.3 x 10(-3) Pa could put again the chromium precipitates into a solid solution, thereby returning the surface chemical composition and the sigma app nearly to the original states. In addition, the solution-treated particles showed a good cleanability comparable to that of the original particles. These results demonstrated that solution treatment could improve the lowered cleanability of the pseudo-sensitized stainless steel surfaces.

Role of a Surfactant in Caustic Alkali Cleaning of Polyethylene Terephthalate Surfaces Fouled with Organic Substances

The role of polyoxyethylene lauryl ether (POELE), a nonionic surfactant, in removing bovine serum albumin (BSA) and catechin from polyethylene terephthalate (PET) particles was quantitatively studied. Relatively large fractions of adsorbed BSA and catechin were removed from PET particles under weak alkaline conditions below pH 9, whereas cleaning with high concentrations of NaOH was not so effective, especially on the removal of BSA. This was attributed to the lower susceptibility of PET particles to water-based alkali (OH-) cleaning because of the lower degree of the polarity of PET surfaces. The combined use of NaOH solutions of moderate pHs and POELE could successfully improve the efficiency of cleaning of PET particles without a significant dissolution of PET materials. There was a good correlation between the lowered surface tension (7) of the NaOH solution with POELE and the removal efficiency of BSA or catechin. No effect of POELE was observed on the NaOH cleaning of hydrophilic alumina (A12O3) particles fouled with BSA. It could be concluded that the role of POELE in NaOH cleaning of PET particles was to lower the gamma of the NaOH solution and hence to facilitate penetration of OH- ions into the PET-BSA and PET-catechin interfaces.

The Mode of Action of Sodium Hypochlorite in the Cleaning Process

The action of sodium hypochlorite (NaOCl) solution in the cleaning of alumina (Al2O3) particles fouled with bovine serum albumin (BSA) or pectin was studied as a function of pH. The efficacy of NaOCl in the cleaning process was evaluated in the pH region where OH- alone exerted no significant action on BSA or pectin removal. The efficiency and the rate of the removal of BSA or pectin increased with increasing available chlorine concentrations and pH. The pH dependence of the efficacy of NaOCl could be explained on the basis of the equilibrium between undissociated hypochiorous acid (HOCl) and dissociated hypochlorite ion (OCl-). The efficacy of NaOCl in the removal of BSA or pectin depended on the concentration of OCl-, but not on HOCl. Size-exclusion chromatographic analysis showed that BSA molecules were partially decomposed by the action of NaOCl, whereas no significant variation in the molecular weight of pectin molecules took place. The difference between the actions of OCl- in the removal of BSA and pectin could probably be ascribed to the reactivity of OCl- with their molecules. It could be concluded that the concentration of OCl- was a major factor determining the actual NaOCI concentration required for an efficient cleaning treatment.

Detergency Effect of Trisodium Phosphate on the Removal of Bovine Serum Albumin from Alumina Surfaces during Caustic Alkali Cleaning.

The detergency effect of trisodium phosphate (Na3PO4) on the removal of irreversibly adsorbed bovine serum albumin (BSA) from alumina (Al2O3) particles during cleaning with 0.05 M NaOH solution was kinetically studied. The desorption curve of BSA, obtained by plotting the logarithm of the amount of residual BSA against cleaning time, during cleaning with the NaOH solution with and without Na3PO4 could be apparently reduced to the sum of two straight lines with different slopes. This fact suggested that two species of BSA, i.e., a fasterdesorbing species (BSAf) and a slower-desorbing species (BSAs), were present on Al2O3 surfaces and desorbed according to a first-order process. The first-order desorption rate constant (ks) of BSAs obtained from cleaning with the NaOH solution alone was approximately 56-fold lower than that (Kf) of BSAf. The addition of Na3PO4 to the NaOH solution had no influence on the desorption of BSAf, whereas it accelerated the desorption of BSAs, with an approximately 7-fold increase in the ks. Potentiometric titration showed that phosphate ions were adsorbed on Al2O3 surfaces after cleaning with the NaOH solution containing Na3PO4. The results indicated that hydroxide ion and phosphate ions could displace the slower-desorbing BSAs species at Al2O3-water interfaces in a synergistic manner.

Surface modification of polypropylene membrane by low‐temperature plasma treatment

AbstractMicroporous polypropylene membranes were low temperature plasma treated with acrylic acid and allylamine. Parameters of plasma treatment were examined and optimized for the enhancement of membrane performance properties. Excess power damaged the membrane surface and excess monomer flow rate increased the reactor pressure to interfere with the glow discharge. Longer plasma treatment time resulted in even more plasma coating and micropore blocking. The contact angle with water decreased and wettabilities increased with the increase of plasma treatment time. Deposition of the plasma polymer on the membrane surface was confirmed by FTIR/ATR spectra of the treated surface. In determining the flux, the hydrophilicity of the surface played a role as important as that of the micropore size. Adequate plasma treatment could enhance both water flux and solute removal efficiency. Results from the BSA (bovine serum albumin) solution test confirmed that fouling was greatly reduced after the plasma treatment. The BSA solution flux through the plasma‐treated membranes depended on pH, whereas pH variation had no serious effects on the untreated membrane. Modification of the surface charge by the plasma treatment should exert a substantial influence on the adsorption and removal of BSA. © 2001 John Wiley &amp; Sons, Inc. J Appl Polym Sci 81: 1555–1566, 2001

Facilitation of Cleaning of Alumina Surfaces Fouled with Heat-Treated Bovine Serum Albumin by Ozone Treatment

Facilitation of cleaning of alumina (Al2O3) particles fouled with heat-treated bovine serum albumin (BSA), which contains sulfhydryl groups on the molecule, by gaseous ozone was studied. With increasing temperature of heat treatment, the amount of adsorbed BSA onto Al2O3 surfaces increased, whereas the rate of BSA desorption during alkali cleaning decreased markedly, resulting in the larger amounts of BSA remaining on Al2O3 surfaces. No significant amounts of BSA were removed from Al2O3 surfaces by alkali cleaning alone when treated at temperatures above 120°C. Before alkali cleaning, the heat-treated, BSA-fouled Al2O3 at 150°C were treated with 0.05 to 0.30% (vol/vol) gaseous ozone at room temperature. Ozone pretreatment markedly accelerated the rate of BSA desorption during subsequent alkali cleaning. The effect of ozone pretreatment on BSA removal depended on the concentration of ozone and treatment time and hence on the total amount of ozone supplied. The molecular weight (MW) of desorbed BSA during alkali cleaning without ozone pretreatment coincided with the MW of the native BSA, whereas the MW of desorbed BSA during the combined ozone-alkali cleaning was lower than the MW of the native BSA. This indicated that the heat-treated BSA molecules adsorbed on Al2O3 were partially decomposed into some fragments by ozone pretreatment, resulting in the facilitation of the removal of BSA during alkali cleaning.

Effect of ozone pretreatment on alkali cleaning of alumina fouled with bovine serum albumin

Pretreatment of alumina particles fouled with bovine serum albumin (BSA) by 0.3% (v/v) gaseous ozone markedly accelerated the removal of BSA during alkali cleaning through partial decomposition of the BSA molecule. The results suggest that ozone pretreatment can reduce the NaOH concentration required for adequate alkali cleaning by at least one order of magnitude.

Effects of bovine serum albumin on function of cryopreserved stallion spermatozoa during medium culture and uterine tube epithelial cell coculture

Abstract Objective To compare function of cultured cryopreserved stallion spermatozoa in a modified Tyrode’s medium (TM), with or without bovine serum albumin (BSA), or in uterine tube (oviduct) epithelial cell (OEC) coculture in TM, with or without BSA. Sample Population Cryopreserved spermatozoa from 6 proven stallions and OEC from bovine reproductive tracts in follicular phase. Procedure Thawed spermatozoa were cultured in TM, with or without BSA, or cocultured with OEC monolayers in TM, with or without BSA. Percentages of capacitated and acrosome-reacted spermatozoa were measured at 5 hours for TM cultures. Spermatozoal survival and motility characteristics were observed over time for all culture methods. Number of spermatozoa attaching to OEC were compared for cocultures. Results Use of TM without BSA altered spermatozoal function in cell-free medium culture and OEC coculture. A higher percentage of spermatozoa were acrosome reacted in TM with BSA, although percentages of capacitated spermatozoa did not differ. Spermatozoa survived longer and maintained superior motion in TM culture without BSA and in OEC cocultures. More spermatozoa were able to attach to OEC in TM without BSA. Conclusions Incubation of cryopreserved spermatozoa in media with BSA resulted in rapid decrease in percentage of intact, motile spermatozoa and limited their ability to interact with OEC. Clinical Relevance Current culture media used for assisted reproduction techniques in horses do not provide functionally capacitated spermatozoa. Removal of BSA from such media improves spermatozoal quality and survival. (Am J Vet Res 1999;60: 363–367)