Adsorption bovine serum albumin protein

Duracher, D., Veyret, R., Elaissari, A., and Pichot, C. 2004. Adsorption of bovine serum albumin protein onto amino-containing thermosensitive core-shell latexes. Polymer International 53 618-26. 

We used an anti-DNA antibody as an exploratory model system. The antibody was monoelonal from mouse sourees and its subelass was IgM. Mouse IgG (MW 1.5 x 10 Da) and IgM (MW 9 X 10 Da) antibodies from normal plasma, and bovine serum albumin were used for the eontrol measurements. To prevent the nonspeeilie adsorption of proteins to the uneovered, bare Au site in the modified eleetrode surfaee, the DNA-modified eleetrode prepared by the standard proeedure was further treated with aqueous 2-mercaptoethanol solution and was used for the measurements. 

Membranes offer a format for interaction of an analyte with a stationary phase alternative to the familiar column. For certain kinds of separations, particularly preparative separations involving strong adsorption, the membrane format is extremely useful. A 5 x 4 mm hollow-fiber membrane layered with the protein bovine serum albumin was used for the chiral separation of the amino acid tryptophan, with a separation factor of up to 6.6.62 Diethey-laminoethyl-derivatized membrane disks were used for high-speed ion exchange separations of oligonucleotides.63 Sulfonated membranes were used for peptide separations, and reversed-phase separations of peptides, steroids, and aromatic hydrocarbons were accomplished on C18-derivatized membranes. 

Proteins may be covalently attached to the latex particle by a reaction of the chloromethyl group with a-amino groups of lysine residues. We studied this process (17) using bovine serum albumin as a model protein - the reaction is of considerable interest because latex-bound antigens or antibodies may be used for highly sensitive immunoassays. The temperature dependence of the rate of protein attachment to the latex particle was unusually small - this rate increased only by 27% when the temperature was raised from 25°C to 35°C. This suggests that non-covalent protein adsorption on the polymer is rate determining. On the other hand. the rate of chloride release increases in this temperature interval by a factor of 17 and while the protein is bound to the latex particle by only 2 bonds at 25°C, 22 bonds are formed at 35°C. 

Fig. 6. Plateau-values, I"P1 /mg m 2, of adsorption isotherms of lysozyme (LSZ), ribonuclease (RNase), a -lactalbumin (aLA), calcium-depleted (X -lactalbumin (aLA(-Ca )) and bovine serum albumin (BSA) on hydrophobic polystyrene (PS) and hydrophilic hematite (a — Fe203) and silica (Si02) surfaces. An indication of the charge density of the surface is given by the zeta-potential, C, and of the proteins by + and signs. Ionic strength 0.05 M T = 25°C. (Derived from Currie et al. 2003).

Any detectable effect on the reaction or behavior of a particular system by the interior wall of the container or reaction vessel. Because proteins can form high-affinity complexes with glass and plastic surfaces, one must exercise caution in the choice of reaction kinetic conditions. Wall effects can be discerned if one determines catalytic activity under different conditions that minimize or maximize contact of the solution with the container. In principle, an enzyme-catalyzed reaction should proceed at the same rate if placed in a capillary or a culture tube however, contact with the wall is maximized in a capillary, and wall effects should be more prominent. Some investigators add bovine serum albumin to prevent adsorption of their enzyme onto the container s walls. 

From the experiments it is clear that poly electrolyte is adsorbed on the surface of the black lipid film. This applies both to the experiments with gelatin and bovine serum albumin, which gave no decrease of film resistance, and to the experiments with bovine erythrocyte ghost protein and polyphosphate. The adsorption of protein on the phospholipid-water interface may be controlled independently by investigating the electrophoretic behavior of emulsion droplets, stabilized by phospholipid, in a protein solution, as a function of pH. In this way Haydon (3) established protein adsorption on the phospholipid-water interface. If the high resistance (107 ohms per sq. cm.) of black lipid films is to be ascribed to the continuous layer of hydrocarbon chains in the interior of the film, as is generally done, an increase in film conductivity is not expected from adsorption without penetration. 

Morrissey 53) used transmission infrared spectroscopy to study protein adsorption onto silica particles in a heavy water (DzO) buffer. By observing the shift in the amide I absorption band, he could deduce the fraction of protein carbonyl groups involved in bonding to the silica surface. He found that bovine IgG had a bound fraction of 0.20 at low bulk solution concentrations, but only about 0.02 at high solution concentrations. However, neither prothrombin nor bovine serum albumin exhibited a change in bound fraction with concentration. Parallel experiments with flat silica plates using ellipsometry showed that the IgG-adsorbed layers had an optical thickness of 140 A and a surface concentration of 1.7 mg/m2 at low bulk solution concentration — in concentrated solutions the surface amount was 3.4 mg/m2 with a thickness of 320 A (Fig. 17). 

There are several recent examples of the switching of nonspecific protein binding on polymer surfaces by application of an external stimulus. Alexander and coworkers demonstrated that protein adhesion can be controlled on PNIPAM surface brushes [14, 181]. For instance, it was reported that the adsorption of FITC-labeled bovine serum albumin (FITC-BSA) on PNIPAM/hexadecanethiol micropatterned surfaces could be tuned by LCST. However, this effect was found to be less pronounced after prolonged incubation times or repeated heating/cooling cycles. The authors suggested that this behavior could be due to unspecific PNIPAM-protein interactions [14],... 

Whitesides and coworkers describe the use of an elastomeric membrane to pattern proteins and cells on bacteriological polystyrene (PS), glass, and poly(dimethyl-siloxane) (PDMS) substrates [92], A patterned PDMS membrane was casted from lithographically structured photoresists and brought into close contact with the substrates (Fig. 6). When incubated with a solution of fibronectin (FN), adsorption of the cell-adhesion-mediating protein to the surface was restricted to the exposed areas. The membrane was peeled off and cells were seeded from a serum-free medium. Passivation to cell attachment of the untreated portions of the surface was achieved by adding 1% bovine serum albumin (BSA) to the cell-seeding medium, which... 

An example of the immobilization of antibodies on channel surfaces was presented by Eteshola and Leckband [395]. A microfluidic sensor chip was developed to quantify a model analyte (sheep IgM) with sensitivities down to 17 nM. This was achieved by first immobilizing a layer of bovine serum albumine (BSA) onto the channel wall, followed by specific adsorption of protein A to which the primary antibody for IgM was coupled covalently. This antibody could capture IgM, which was detected with the secondary antibody, labeled with horseradish peroxidase (Scheme 4.91). This enzyme catalyzes the conversion of the fluorogenic substrate 3-(p-hydroxyphenyl)propioni c acid into a fluorophore, which was quantified off-chip with a spectrofluorometer. The measured fluorescence signal was proportional to the analyte concentration in the test sample. 

The effects of conditioning layers of two important blood serum proteins, albumin and fibrinogen were investigated. Protein adsorption was studied using bovine serum albumin (BSA) and fibrinogen (F) from Sigma. The samples were incubated for 3 h at 37°C in solutions of albumin (1 mg/mL) and fibrinogen (0.2 mg/mL) prepared in phosphate buffered saline (PBS, 0.01 M phosphate buffer, 0.0027 M KC1, 0.137 MNaCl, pH 7.4). After the incubation period, the samples were rinsed 3 times with PBS and analyzed by the various surface characterization techniques. 

Comparison of equilibrium adsorption (Figures 4 and 5) and minute protein adsorption/flocculation as a function of protein concentration, Cp, demonstrates strong but variable effects of pH and salinity.4 The equilibrium adsorption of proteins is as large as 1 mg/m2 (or 300 mg/g) at pH 3.5 (i.e. between pH(IEP) of silica and proteins) for bovine serum albumin (BSA) with 0.9 wt.% NaCl and gelatin without NaCl, or at pH(IEP) of protein for ovalbumin without NaCl. The lowest equilibrium adsorption (0.1-0.2 mg/m2) is typically observed at pH = 2, which is close to pH(IEPS o2) 2.2, and without NaCl (Figure 4). It should be... 

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