Miroslav Bleha

Purification of the specific immunoglobulin G1 by immobilized metal ion affinity chromatography using nickel complexes of chelating porous and nonporous polymeric sorbents based on poly(methacrylic esters). Effect of polymer structure

Ni2+ complexes of the chelating nonporous and porous bead sorbents based on methacrylic esters crosslinked with ethylene dimethacrylate were used in isolation of the horseradish peroxidase-specific immunoglobulin IgG1 from the crude mouse ascitic fluid by immobilized metal ion affinity chromatography (IMAC). Iminodiacetic and aspartic acids were attached to porous poly(glycidyl methacrylate) beads differing in size, morphology and chemical composition. Ethylenediaminetriacetic acid and quinolin-8-ol chelating groups were attached mainly to the surface hydroxyl groups in nonporous poly(diethylene glycol methacrylate) beads through spacers. The latter sorbents exhibited better kinetic characteristics than the former but a very low IgG1 sorption capacity. In a single-step IMAC procedure, the best efficiency in the specific IgG1 purification was obtained with porous sorbents (recovery 92%, purity 73%). Differences in IMAC separations are discussed from the point of view of morphology of polymer beads as well as of the type and concentration of chelating ligands.

Gas permeabilities of polymers with indan groups in the main chain.: 2: Polyimides

Abstract Permeability coefficients and ideal selectivities for H2, CO2, O2, and N2 were determined for two series of polyimides with indan groups in the main chain. The bulkiness of the indan group was varied by replacing methyl groups at the indan ring system by bulky cyclohexyl groups. The effects of the linking group in the dianhydride part of the polymer repeating unit could be explained on the basis of bulkiness and flexibility. However, the introduction of two cyclohexyl substituents at the indan group to increase the bulkiness of this structural element had no uniform effect on permeability coefficients and selectivities. Positive or negative effects were observed, depending on the linking group in the dianhydride moiety. This was tentatively attributed to the conformational flexibility of the cyclohexyl rings, which can result in different sterical requirements.

Effect of salt concentration gradient on separation of different types of specific immunoglobulins by ion-exchange chromatography on DEAE cellulose

A three-stage process, consisting of an ammonium sulfate precipitation step, dialysis desalination with microporous anion-exchange Neosepta membranes and anion-exchange chromatography on DEAE-cellulose DE-52 was used for the isolation of mouse monoclonal antibodies specific against different antigens. The ascites fluids contained monoclonal antibodies against human IgG, against horseradish peroxidase and against the heavy chain of human IgM. The effect of the salt concentration gradient in the elution buffer was examined with the aim of optimizing chromatographic conditions. The quality of separation of protein zones was determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis under non-reducing conditions. The immunoreactivity of purified monoclonal antibodies was determined by enzyme-linked immunosorbent assay using a solid-phase adsorbed antigens against which each monoclonal antibody type was directed.

Effect of inert components on the porous structure of 2-hydroxyethyl methacrylate-ethylene dimethacrylate copolymers

The products obtained by suspension copolymerization of 2-hydroxyethyl methacrylate with ethylene dimethacrylate in water in the presence of 1-dodecanol and cyclohexanol as inert diluents were characterized by scanning electron microscopy, mercury porosimetry, water and cyclohexane regain and volume swelling experiments. The porosity of the resulting poly(2-hydroxyethyl methacrylate) beads was readily adjusted by a change in the ratio of 1-dodecanol (non-solvating diluent) to cyclohexanol (solvating diluent). The morphological structure of the porous samples was also influenced by the choice of drying technique. Freeze-drying of samples swollen in water increased porosity compared to samples air-dried from ether.

Gas permeabilities of polymers with indan groups in the main chain.: 1: Poly(ether ketone)s

Abstract The gas permeabilities for hydrogen, carbon dioxide, oxygen and nitrogen of a series of poly(ether ketone)s containing indan groups were determined. The effects of structural variations on permeability coefficients and selectivities for the gas pairs H 2 /N 2 , CO 2 /N 2 , O 2 /N 2 , H 2 /O 2 , CO 2 /O 2 and H 2 /CO 2 are discussed. Based on the results, it is suggested that gas permeation and hence selectivity in these polymers is controlled by a specific chain segment, which consists of a sequence of flexibly linked phenyl rings. These segments are connected by the bulky and immobile indan groups.

Immobilized-metal affinity sorbents based on hydrophilic methacrylate polymers and their interaction with immunoglobulins

Abstract Hydrophilic homogeneous poly[2-(2-hydroxyethoxy)ethyl methacrylate- co -ethylene dimethacrylate] poly(DEGMA- co -EDMA) beads have been prepared as precursors for introducing the chelating groupings of ethylenediaminetriacetic acid, quinolin-8-ol, or N -(2-pyridylmethyl)glycine. The effect of the chelating ligand structure on coordination with Ni 2+ ions and that of the metal complexes on binding the model protein (horseradish-peroxidase-specific immunoglobulin IgG 1 ) was analyzed. It was found that the protein binding affinity and selectivity was the highest for IMA sorbents with quinolin-8-ol groups.

Electrical resistance and diffusion permeability of microporous polyethylene membranes modified with polypyrrole and polyaniline in solutions of electrolytes

Composite systems based on a microporous polyethylene (PE) membrane modified with conducting polymers such as polypyrrole (PPy) and polyaniline (PANI) have been elaborated. Conducting polymers were deposited onto PE membranes in situ during the oxidative polymerization of pyrrole from gas phase or by the polymerization of aniline in aqueous medium. Composite membranes have a low resistance in electrolyte solutions owing to the coating with conducting polymers inside the pores. The dependence of electrical resistance of modified membranes on concentration of hydrochloric acid or sodium hydroxide solutions, in which the conducting polymers are in protonated or non-protonated states, respectively, was studied using alternating and direct electric currents. Diffusion permeability of composite membranes to acid, salt and alkali solutions has been also studied.

Microporous membranes from polyolefin-polyamide blend materials☆

An alternative method using established polymer processing and membrane formation processes to prepare hydrophilic membranes based on acid modified polyolefins is introduced. This process includes blend formation by reactive extrusion of functionalized polyolefins (polypropylene; PP) and a hydrophilic polyamide (PA). In a first step the pore formation is realized by extracting non-covalently bound polyamide. The resulting membranes are tested by pure water as well as protein filtration. The properties are discussed in terms of permeate flux, retention capability and fouling behavior.

Ultrafiltration and microfiltration membranes in latex purification by diafiltration with suction

Operation conditions of diafiltration with suction in purification of poly(glycidyl) methacrylate latex from sodium tetraborate and emulsifier were studied in a batch process using ultrafiltration blend polysulfone/poly(vinylpyrrolidone) and microfiltration Synpor® membranes. Intensity of permeate suction was controlled by changing the pumping rate at fixed cross-sections of the inlet tubes in both the retentate and permeate lines. An optimum value of flow rate was determined for each membrane type to ensure the best purification efficiency. Operating at this flow rate prevented not only undesirable dilution of the latex with osmotic water but also ensured the highest membrane permeability to solutes without cake formation on the membrane surface. It was shown that 92% degree of latex purification could be obtained by 8-h suction diafiltration with Synpor membrane having the pore entrance sizes close to nanoparticle dimensions. The possibility of complete purification of GMA nanoparticles from impurities using the hybrid membrane process combining dialysis followed by suction diafiltration with microporous membranes, and ultrafiltration with an appropriate membrane is discussed.

Purification of polymer nanoparticles by diafiltration with polysulfone/hydrophilic polymer blend membranes

Abstract The separation ability of new UF membranes based on blends of polysulfone (PS) with hydrophilic polymers (HP) was investigated in purification of polymer nanoparticle latices prepared by seeded emulsion polymerization. Low molecular weight components (emulsifier, salts, initiator) used in the synthesis of both glycidyl methacrylate (GMA) and sulfonated (SSS) nanoparticles as well as monomer residues were separated from nanoparticles of colloidal size (70–100 nm) by diafiltration using a three-compartment through-flow cell equipped with two membranes. In contrast to traditional pressure-driven diafiltration, this process is carried out under vacuum. The efficiency of new membranes and some commercial membranes were compared in the purification of nanoparticles. The influence of the nature of HP and structural characteristics of the skin layer of UF PS/HP membranes on their permeability to components of latex emulsions was estimated. The structure parameters of the skin membrane layer varied in the intervals: 1–7 nm (average diameter of pores), 0.6–49.2×10 10 (pore density) and 0.1–0.88% (relative surface porosity). For each type of membrane, an optimum correlation between the structure membrane parameters was found for ensuring the maximum permeate flux and purification degree of nanoparticle emulsions. It was shown that 95–98% and 85–89% purification degrees of GMA and SSS nanoparticle emulsions can be achieved by 6 h diafiltration with some UF blend PS/PVP membranes.