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Affinity chromatography of fibroblast growth factors on substituted polystyrene.

The heparin-binding growth factors aFGF and bFGF (acidic and basic fibroblast growth factor) from crude bovine brain extract were co-eluted with purified [125I]aFGF and/or [125I]bFGF as tracers from heparin-Sepharose and from several insoluble substituted polystyrenes used as stationary phases in low-pressure affinity chromatography. The ability of the resins to isolate FGFs was determined by measuring the eluted radioactivity. It was demonstrated that the various substituted polystyrene resins retain [125I]aFGF and [125I]bFGF with different specificities according to the chemical nature of the substituted groups bound to the polystyrene support. Bifunctional resins substituted with sulphonate and phenylalanine sulphamide groups adsorbed both [125I]aFGF and [125I]bFGF whereas bifunctional resins substituted with sulphonate and sulphamide serine adsorbed only [125I]bFGF. These stationary phases could be adapted to high-performance affinity chromatography and used to isolate growth factors of the FGF family.

Animals↗

Surface modification and electrostatic charge of polystyrene particles.

Particle surface modification by poloxamer adsorption can significantly alter the electrostatic charge, adhesion behaviour and consequently handling properties of a material. The charge reduction on polystyrene spheres achieved by this modification technique is dependent on the concentration, molecular weight and conformation of poloxamer at the particle surface. Adsorption isotherms of poloxamers on polystyrene particles follow a Langmuir profile and there is an apparent correlation between the extent of adsorption and ability of poloxamer to reduce electrostatic charge. Surface analysis techniques, X-ray photoelectron spectroscopy and Time of Flight Secondary Ion Mass Spectrometry have generated data on the thickness of the adsorbed poloxamer layer and provided evidence to suggest that the polypropylene oxide component of the poloxamer adsorbs to the polystyrene surface and there is a polyethylene oxide rich outer surface which may influence the charge alteration.

Adsorption↗

Adsorption-desorption of antigen to polystyrene plates used in ELISA.

Since ELISA reliability depends to a great extent upon solid-phase reagent concentration and stability, we sought to analyse the influence of experimental conditions during ELISA performance on the adsorption/desorption of proteins to microplates. The effect upon desorption of several experimental parameters (antigen concentration, antibody concentration and affinity, washings, conjugate and inhibitor incubations) and quantitative treatment of protein-polystyrene adsorption were analysed. The adsorption to polystyrene microplates was studied with a hapten-conjugated protein (BSA-Ar36) in order to facilitate the analysis of the influence of antibody affinity on desorption during ELISA. Our results show that polystyrene plates adsorb BSA-Ar36 according to the Langmuir isotherm. The adsorption constant was 2.1 X 10(8) L/mol and maximal surface concentration of protein on solid phase was 1.8 X 10(-7) g/cm2. Although desorption was present, we observed that it did not affect the reliability of results of either direct or inhibition ELISA, because it was not dependent on the composition of the sample.

Adsorption↗

Separation of transfer ribonucleic acids on polystyrene anion exchangers.

The transfer RNA separation by chromatography on strong-base polystyrene exchange materials is examined and compared with the widely used reversed-phase chromatography. Results indicate important differences in some transfer RNA (tRNA) elution patterns by the anion-exchange chromatography, as compared with the reversed-phase chromatography. Transfer RNAs containing hydrophobic groups are adsorbed more strongly. The anion exchanger has twice the number of theoretical plates. Single peaks of tRNA2Glu and tRNA1Phe obtained from the reversed-phase column give multiple peaks only polystyrene anion-exchange chromatography. All six leucine tRNAs (Escherichia coli) and differences in tRNA populations synthesized during early and late stages of the dividing lymphocytes from normal human blood can be characterized by the anion-exchange chromatography. Different separation profiles are obtained by two separation systems for tyrosine tRNAs from mouse liver and mouse-plasma-cell tumor. The results indicate that, in contrast to the reversed-phase chromatography, strong-base-polystyrene anion-exchange chromatography is capable of separating tRNAs with minor structlral differences.

Animals↗

Amphiphilic block copolymers as bile acid sorbents: 1. Synthesis of polystyrene-b-poly(N,N,N-trimethylammoniumethylene acrylamide chloride).

The systematic investigation of the synthesis of polystyrene-b-poly(N,N,N-trimethylammoniumethylene acrylamide chloride) was accomplished by employing both polystyrene-b-poly(tert-butyl acrylate) and its hydrolyzed derivative, polystyrene-b-poly(acrylic acid) (PS-b-PAA) as starting materials, and coupling them with N,N-dimethylethylenediamine (DMED). The various reactions and intermediates we examined include aluminum amides, acid chlorides, and imides derived from carbodiimides, all in a variety of solvents. We present below our investigation of several synthetic routes and conclude that the carbodiimide coupling of PS-b-PAA with DMED followed by quaternization and counterion exchange is the most effective method of achieving the target. A brief discussion of the merits of each procedure in the context of block copolymers is given, and IR spectroscopic evidence for the postpolymerization synthesis of the poly(acrylamide) block is provided.

Acrylic Resins↗

Synthesis and characterization of new micrometer-sized radiopaque polymeric particles of narrow size distribution by a single-step swelling of uniform polystyrene template microspheres for X-ray imaging applications.

Radiopaque micron-sized non-cross-linked polystyrene/poly(2-methacryloyloxyethyl(2,3,5-triiodobenzoate)) particles of narrow size distribution were prepared by a single-step swelling of uniform polystyrene template microspheres with emulsion droplets of methylene chloride containing the initiator benzoyl peroxide and the iodinated monomer 2-methacryloyloxyethyl(2,3,5-triiodobenzoate), followed by the polymerization of the monomer within the swollen template particles at 73 degrees C. Radiopaque micron-sized uniform cross-linked polystyrene/poly(2-methacryloyloxyethyl(2,3,5-triiodobenzoate)-divinylbenzene) composite particles were prepared similarly with emulsion droplets of methylene chloride containing divinylbenzene, in addition to the initiator and the iodinated monomer. Radiopaque micron-sized uniform cross-linked poly(2-methacryloyloxyethyl(2,3,5-triiodobenzoate)-divinylbenzene) particles were formed by dissolving the template polystyrene polymer belonging to the former cross-linked composite particles. Characterization of these novel radiopaque polymeric particles was performed by methods such as FTIR, TGA, DSC, SEM, XPS, elemental analysis, and light microscopy. The influence of the weight ratio [2-methacryloyloxyethyl(2,3,5-triiodobenzoate)]/[polystyrene] and [2-methacryloyloxyethyl(2,3,5-triiodobenzoate)]/[divinylbenzene] on the bulk and surface properties of the non-cross-linked and cross-linked particles, respectively was elucidated. The radiopacity of these iodinated particles was demonstrated by an imaging technique based on X-ray absorption usually used in hospitals. These novel radiopaque particles may be used for different X-ray imaging needs, e.g., blood pool, body organs, embolization, dental composition, implants, protheses, and nanocomposites.

Absorptiometry, Photon↗

Synthesis and high performance liquid chromatography/electrospray mass spectrometry single-bead decoding of split-pool structural libraries of polyamines supported on polystyrene and polystyrene/ethylene glycol resins.

Natural polyamines are ubiquitous biomolecules present in all living cells. These cationic compounds play essential roles in both cell growth and differentiation and are known to interact in complex ways with polyanionic biomolecules. Consequently, there is significant interest in expanding nature's polyamine diversity using combinatorial synthesis and screening strategies. This article describes an efficient split-pool solid-phase synthetic strategy toward the generation of encoded libraries of polyamines via the exhaustive borane-promoted reduction of trityl-linked, resin-bound polyamides. Model structural libraries of tetra- and pentaamines were designed from a set of geometrically diverse amino acid building blocks. To encode the libraries, a partial termination synthesis approach was employed at the polyamide stage, allowing each library to be analyzed from single beads by HPLC/ESMS under two sets of conditions featuring both pH extremes. Determination of the sequence of polyamine residues was simply achieved by the mass differences observed between the full oligomers and the terminated ones. Both polystyrene- and TentaGel-supported libraries, including a library of 4913 pentaamines, were prepared and successfully decoded. For the TentaGel-supported libraries, suitable for on-bead aqueous screening of biomolecules, a novel trityl-derivatized resin was prepared in which the trityl group is anchored to the poly(ethylene glycol) chains via a methylene group. The resulting resin is much more resistant than other commercially available polystyrene-poly(ethylene glycol) trityl resins to the harsh borane reduction conditions required. Two workup conditions for the cleavage of the resultant borane-amine adducts were evaluated on the TentaGel bound polyamide 14. Although the two methods showed a comparable efficiency when using the polystyrene support, with 14 it was found that the piperidine-exchange method afforded polyamines of higher purity than the iodine-based oxidative method previously developed in our laboratory.

Chromatography, High Pressure Liquid↗

Uptake of polychlorinated biphenyls (PCBs) from an aqueous medium by polyethylene, polyvinyl chloride, and polystyrene films.

The sorption of selected polychlorinated biphenyl (PCB) congeners (from tri to deca chlorinated) by three food-packaging plastic films [polyethylene, polyvinyl chloride (PVC), and polystyrene] from an aqueous solution was investigated. From the data generated, PCB uptake, partition, and diffusion coefficients were calculated for the various films. Polyethylene exhibited the highest PCB uptake, diffusion, and partition coefficients when compared to the other materials. Although PVC indicated larger sorption diffusion and partition coefficients for the lower chlorinated congeners than polystyrene, a reversal of this trend was observed for the higher congeners. For polyethylene and PVC, the PCB uptake decreased as the chlorine numbers in the congeners increased, confirming the correlation between increasing chlorination and increasing cohesive density within the PCB molecules. For polystyrene, the uptake decreased from tri to penta congeners, but showed an increase for the hexa, and then a decreased uptake until the deca chlorination. A comparison of the molecular sizes of the PCB congeners showed that the partition (Ke) and sorption diffusion (Ds) coefficients generally decreased with their increasing molar volumes. The resulting Ke values were used to determine the extent of sorption because these values indicate the affinity of PCBs for the plastic films. Results from this study can be of practical importance for cases of product quality related to the transfer of contaminants from the product to the packaging materials.

Food Packaging↗

Fabrication of self-organized chemically and topologically heterogeneous patterns on the surface of polystyrene-b-oligothiophene block copolymer films.

A novel fabrication of the chemically and topologically heterogeneous patterns on the surface of polymeric films over an area of more than 1 square centimeter in a single step was demonstrated by using the self-organizing character of polystyrene-b-oligothiophene block copolymers. Hexagonally arranged open pores of a size of approximately 2 mum are spontaneously formed by casting the polymer solution under a moist air flow. The amphiphilic character of the polystyrene-b-oligothiophene block copolymers played the crucial role as a surfactant to stabilize the inverse emulsion of water droplets in the organic solvent, and subsequently the structure of the arranged hydrophilic oligothiophene segments remained on the interiors of the micropores. The chemical composition of the surface of the microporous films was characterized by time-of-flight secondary ion mass spectrometry (ToF-SIMS) to prove the chemical heterogeneity. The ToF-SIMS imaging clearly indicated that the oligothiophene forms the aggregated structure on the interior of the open micropores on the surface while the flat area on the surface was covered with the polystyrene.

Microscopy, Electron, Scanning↗

Fabrication of polystyrene latex nanostructures by nanomanipulation and thermal processing.

The capability to fabricate nanoscale structures is a fundamental step toward realizing the promise and potential of nanotechnology. We report on precise manipulation and thermal processing using 100-nm polystyrene latex nanoparticles. This approach is illustrated by fabricating a three-dimensional nanostructure by using an AFM tip to position nanoparticles and then thermally processing to "sinter" the particles to form a contiguous, stable structure. We suggest that this is a general approach, but the use of polystyrene latex particles offers an advantage of low-temperature processing. Use of polystyrene latex also extends the range of materials for which we have demonstrated manipulation and suggests applications including fluorescent doping and electrically conducting polymers.

Crystallization↗

Influence of fluorescent labelling of polystyrene particles on phagocytic uptake, surface hydrophobicity, and plasma protein adsorption.

PURPOSE: To investigate the influence of fluorescent labelling of polystyrene particles on phagocytic uptake, surface hydrophobicity and protein adsorption. METHODS: Phagocytic uptake was analysed using chemiluminescence. Hydrophobicity was quantified by adsorption measurements of a hydrophobic dye. Protein adsorption was evaluated by two-dimensional electrophoresis. RESULTS: Commercially available fluorescently labelled particles showed marked differences when compared to unlabelled particles: phagocytic uptake and surface hydrophobicity of labelled particles were diminished. Also the plasma protein adsorption pattern was found to be different from the unlabelled particles: for example, the amount of fibrinogen adsorbed was strongly reduced on the labelled particles. On the other hand, some unknown proteins could be detected on the fluorescently marked particles. In contrast, plain polystyrene particles and labelled ones could be successfully synthesised by Paulke which did not show any considerable differences in phagocytic uptake, surface hydrophobicity and protein adsorption. Polysorbate 20 added as stabilizer to particle suspensions led to completely different behaviour of the particles: the particles showed altered protein adsorption patterns, dominated by immunoglobulins and especially by apolipoproteins. Furthermore, these particles were not phagocytized at all. CONCLUSIONS: Surface hydrophobicity and phagocytic uptake in vitro as well as the interactions with plasma proteins of commercially available polystyrene particles were strongly affected by fluorescent labelling. Particles synthesised by Paulke remained unchanged after labelling. The results show the importance of thorough surface characterization for using particles in test systems in vitro and in vivo.

Adsorption↗

Transfer of lipophilic markers from PLGA and polystyrene nanoparticles to caco-2 monolayers mimics particle uptake.

PURPOSE: The objective of this study was to evaluate nanoparticle uptake by the Caco-2 monolayer model in vitro. Special emphasis was placed on the localization and the quantification of the uptake of fluorescently labeled polystyrene and poly(lactic-co-glycolic acid) (PLGA) nanoparticles. METHODS: Intracellular fluorescence was localized by fluorescence and confocal laser scanning microscopy. Particle uptake was quantified either directly, by counting internalized nanoparticles after separation from the Caco-2 monolayers, or indirectly, by extraction of the lipophilic fluorescence marker. In vitro release studies of lipophilic markers from nanoparticles were performed in standard buffer systems and buffer systems supplemented with liposomes. RESULTS: Instead of uptake of polystyrene and PLGA nanoparticles by Caco-2 monolayers an efficient transfer of lipophilic fluorescence markers from nanoparticles into Caco-2 cells with subsequent staining of intracellular lipophilic compartments was observed. Whereas in standard buffer no release of fluorescent marker from polystyrene and PLGA nanoparticles was observed, the release studies using liposome dispersions as receiver revealed an efficient transfer of fluorescent marker into the liposome dispersion. CONCLUSIONS: The results suggest that the deceptive particle uptake is caused by a collision-induced process facilitating the transfer of lipophilic fluorescent marker by formation of a complex between the nanoparticles and the biomembranes. Diffusion of the marker within this complex into lipophilic compartments of the cell strongly affects quantitative evaluation of particle uptake.

Biological Transport↗

Three dimensional registration of mechanical bladder activity using polystyrene fluorescent spheres: A technical note.

Optical marker tracing methods have been applied successfully in recent years to quantify local material deformation of heart tissue, skin and striated muscles. In this study, polystyrene fluorescent spheres (d = 0.6 mm) are glued to the ventral serosal bladder wall in the rabbit. Three dimensional video registration of the polystyrene spheres is used to calculate two directions of principal strain (epsilon (1), epsilon (2) ) on the bladder surface in vivo. The aim is to investigate the feasibility of the technique for this new application in two experimental circumstances: during spontaneous bladder wall activity and after electrical stimulation of bladder innervating nerve fibers. During spontaneous activity, random contraction and relaxation occurred simultaneously and separately across the bladder wall for the two principal strains epsilon (1) and epsilon (2). After extradural electrical stimulation of sacral nerve root S2, the principal strains epsilon ( 1) and epsilon (2) synchronized in time in such a way that epsilon ( 1) and epsilon (2) both represented contraction or both represented relaxation. One and the same bladder wall area passed through phases of contraction followed by relaxation and vice versa. After multiple stimulation periods, the coordination between the two principal strains during stimulation was reduced. This technique allows to identify local areas of contraction and relaxation in the intact bladder wall in vivo. Three dimensional video registration of polystyrene fluorescent spheres to study bladder wall contraction and its relaxation proved to be a feasible technique, with which electrical stimulation effects and spontaneous activity could be measured.

Animals↗

Coated tube enzyme immunoassay: factors affecting sensitivity and effects of reversible protein binding to polystyrene.

Coated tube enzyme immunoassay using alkaline phosphatase conjugated to rabbit (anti-human IgG) antiserum was studied to determine conditions of maximum sensitivity. The competitive binding assay utilized showed a large increase in sensitivity with immobilized antigen levels below the levels giving rise to the maximum in the coating-antigen dilution series. The effects of reversible antigen binding to the solid phase were investigated by comparison of untreated polystyrene tubes, polystyrene tubes treated with glutaraldehyde and glass tubes activated with an aminosilane. The use of glutaraldehyde treated tubes reduced, and the use of activated glass tubes prevented the time dependent release of immobilized antigen seen with the untreated polystyrene tubes. By comparison of these solid phases, it is shown that reversible antigen immobilized in a competitive binding assay gives rise to poorer conjugate binding (three-fold), and poorer sensitivity (six-fold). A noncompetitive response was found to occur at high free antibody levels and low competing antigen concentrations. This binding behavior is moderated by the minimization of the reversible antigen immobilization.

Animals↗

Identification of cytoskeletal components involved in attachment of L929 cells and macrophages to polystyrene.

We have previously shown that lactoperoxidase (LPO) covalently coupled to polystyrene tissue culture flasks can be used to radioiodinate monolayer cell proteins that come into intimate contact with the LPO-polystyrene surface. These studies have now been extended to include a detailed examination of the class of iodinated polypeptides migrating with apparent molecular weights of 50,000 and 55,000 in SDS polyacrylamide gels. Whereas in cultured L929 cells the 55,000 band is predominantly iodinated, in thioglycollate-activated murine peritoneal macrophages the 55,000 and 50,000 bands are of equal intensity. It is possible that the marked degree of exposure of the 50,000 mol wt polypeptide to immobilized LPO is related to the unique strength of macrophages attachment. After labeling of both L929 cells and macrophages with immobilized LPO, all polypeptides in this molecular weight region were subjected to peptide mapping by simultaneous limited proteolysis and electrophoresis in a second SDS polyacrylamide slab gel. The results clearly show that the two major polypeptides in this region are identical within the limits of resolution of this technique. The 55,000 mol wt polypeptide can also be identified in Triton X-100 cytoskeletons from L929 cells after labeling with soluble LPO either before or after detergent lysis. We conclude that this cell surface polypeptide is in continuity with the cytoskeleton and is preferentially exposed to the substratum during attachment to polystyrene.

Animals↗

Adhesion of cells to polystyrene surfaces.

The surface treatment of polystyrene, which is required to make polystyrene suitable for cell adhesion and spreading, was investigated. Examination of surfaces treated with sulfuric acid or various oxidizing agents using (a) x-ray photoelectron and attenuated total reflection spectroscopy and (b) measurement of surface carboxyl-, hydroxyl-, and sulfur-containing groups by various radiochemical methods showed that sulfuric acid produces an insignificant number of sulfonic acid groups on polystyrene. This technique together with various oxidation techniques that render surfaces suitable for cell culture generated high surface densities of hydroxyl groups. The importance of surface hydroxyl groups for the adhesion of baby hamster kidney cells or leukocytes was demonstrated by the inhibition of adhesion when these groups were blocked: blocking of carboxyl groups did not inhibit adhesion and may raise the adhesion of a surface. These results applied to cell adhesion in the presence and absence of serum. The relative unimportance of fibronectin for the adhesion and spreading of baby hamster kidney cells to hydroxyl-rich surfaces was concluded when cells spread on such surfaces after protein synthesis was inhibited with cycloheximide, fibronectin was removed by trypsinization, and trypsin activity was stopped with leupeptin.

Animals↗

Effects of carbon dioxide and sub-lethal levels of antibiotics on adherence of coagulase-negative staphylococci to polystyrene and silicone rubber.

Fifty coagulase-negative staphylococci (CNS) strains were investigated for adherence to both silicone rubber and polystyrene using a microtitre tray system. Culture in an atmosphere containing a physiological level of carbon dioxide (5% CO2) profoundly affected adherent growth to both surfaces. Most strains adhered less well in this atmosphere compared to in air alone, with mean reductions in adherence of 84% and 86% to silicone rubber and polystyrene respectively. Occasional strains adhered better in 5% CO2. The effects of antibiotic concentrations equivalent to 1/4 MIC of cefuroxime, ciprofloxacin, vancomycin and teicoplanin on the adherence of 10 CNS strains were also studied. Vancomycin and teicoplanin frequently increased adherence to silicone rubber and polystyrene compared to controls. The effects of antibiotics on adherence were not only strain dependent but also sometimes atmosphere or surface specific. Antibiotic-induced changes in adherence did not appear to correlate with changes in strain protein profiles or surface hydrophobicities.

Anti-Bacterial Agents↗

Relative electron beam measurements: scaling depths in clear polystyrene to equivalent depths in water.

For purposes of mean incident energy determination and the accrual of consistent treatment planning data, measurements of relative ionization or dose made in clear polystyrene must be scaled in depth to produce depth-ionization or depth-dose curves equivalent to what would have been measured in water. Recommendations from various protocols for clear polystyrene to water depth scaling factors differ by as much as 5%. Here, central axis measurements of relative ionization as a function of depth have been made with parallel-plate chambers both in a popular clear polystyrene phantom of density 1.045 g/cm3 and in water. Perturbation and displacement corrections were thus minimized. Both depth-ionization and depth-dose curves were formed for electron beams with nominal incident energies between 6 and 20 MeV and field sizes from 6 cm X 6 cm to 15 cm X 15 cm. Comparisons of the depths for 50% relative reading and practical range between the two phantoms yield average empirical scaling factors of 0.990 and 1.002, respectively.

Electrons↗