Affinity based and molecularly imprinted cryogels: Applications in biomacromolecule purification.

The publications in macro-molecularly imprinted polymers have increased drastically in recent years with the development of water-based polymer systems. The macroporous structure of cryogels has allowed the use of these materials within different applications, particularly in affinity purification and molecular imprinting based methods. Due to their high selectivity, specificity, efficient mass transfer and good reproducibility, molecularly imprinted cryogels (MICs) have become attractive for researchers in the separation and purification of proteins.

Molecularly imprinted cryogels for human serum albumin depletion.

Molecularly imprinted polymers can be used for the selective capture of a target molecule from complex medium. Cryogels novel matrices, which characterized by their supermacropores that makes their use advantageous when studying with biological samples. By combining high selectivity of the molecular imprinting approach with using cryogel as a base polymer, in this protocol, preparation of the albumin-imprinted cryogels is described.

Molecularly imprinted poly(hydroxyethyl methacrylate) based cryogel for albumin depletion from human serum.

Macroporous cryogels imprinted with human serum albumin (HSA) have been prepared by copolymerization of 2-hydroxyethyl methacrylate with a functional co-monomer of N-methacryloyl-L-phenylalanine. The cryogels were used for the depletion of HSA from human serum. HSA-imprinted cryogels were prepared with gel fraction yields up to 90%, and their chemical structure, morphology and porosity were characterized by FTIR-spectroscopy, scanning electron microscopy, swelling studies and flow dynamics.

Enzyme-catalyzed crosslinking in a partly frozen state: a new way to produce supermacroporous protein structures.

In this study a new way to produce supermacroporous protein structures was investigated. Enzyme-mediated crosslinking of gelatin or casein was performed in a partly frozen state, which yielded stable, protein-based cryogels. The reaction kinetics for the formation of cryogels were found to be fairly slow, most likely due to the low temperature (-12 °C) used or due to an increased viscosity owing to the cryo-concentration taking place.

Dye attached poly(hydroxyethyl methacrylate) cryogel for albumin depletion from human serum.

Cibacron Blue F3GA was immobilized on poly(hydroxyethyl methacrylate) cryogel and it was used for selective and efficient depletion of albumin from human serum. The poly(hydroxyethyl methacrylate) was selected as the basic component because of its inertness, mechanical strength, chemical and biological stability, and biocompatibility. Cibacron Blue F3GA was covalently attached to the poly(hydroxyethyl methacrylate) cryogel to produce poly(hydroxyethyl methacrylate)-Cibacron Blue F3GA cryogel affinity column.

Formation of macroporous self-assembled hydrogels through cryogelation of Fmoc-Phe-Phe.

In this study, it was found that macroporous hydrogels were formed when self-assembly of fluorenyl-9-methoxycarbonyl (Fmoc)-diphenylalanine (Phe-Phe) peptides was induced using glucono-δ-lactone (GdL) in apparently frozen samples. Formed cryogels exhibited a heterogeneous structure with pore walls of densely packed fibres of assembled dipeptides and pores in the range 10-100 μm. Hydrogels formed from the same composition above the freezing point exhibited a homogenous structure without any apparent porosity.

An improved capillary model for describing the microstructure characteristics, fluid hydrodynamics and breakthrough performance of proteins in cryogel beds.

A capillary-based model modified for characterization of monolithic cryogels is presented with key parameters like the pore size distribution, the tortuosity and the skeleton thickness employed for describing the porous structure characteristics of a cryogel matrix. Laminar flow, liquid dispersion and mass transfer in each capillary are considered and the model is solved numerically by the finite difference method. As examples, two poly(hydroxyethyl methacrylate) (pHEMA) based cryogel beds have been prepared by radical cryo-copolymerization of monomers and used to test the model.

Apparent heterogeneity in the pIII-peptide fusion protein in single-phage clones isolated from peptide libraries.

Ligand homogeneity is an important issue in affinity chromatography. Using phages expressing peptides on the pIII protein, a heterogeneity in the binding of monoclonal phages was observed during affinity chromatography on supermacroporous cryogels. Fractions with different apparent binding affinities could be separated by stepwise elution.

Ultrafast Responsive Poly- (N-isopropylacrylamide) Gel Produced by Cryostructuring of Self-crosslinkable Polymer Microgels.

A macroporous material composed of closely aggregated particles was prepared by cryo-structuration of N-isopropylacrylamide-co-N-hydroxymethylacrylamide (NIPA-co-HMAm) particle suspensions. The formed structure was maintained by the formation of covalent bonds through self-crosslinking between the particles while the system was in a semi-frozen state thus avoiding the need to freeze-dry the sample. This resulted in macroporous structure composed of closely aggregated thermoresponsive particles which exhibit an ultrafast temperature response.

Modulating the porosity of cryogels by influencing the nonfrozen liquid phase through the addition of inert solutes.

The freezing of monomeric mixtures is known to concentrate solutes in a nonfrozen phase in the area surrounding the ice crystals. The concentration of such solutes is determined by the freezing temperature. Although salts or solvents do not directly react in the polymerization reaction, they do change the composition and properties of the nonfrozen phase.

Evaluation of selective composite cryogel for bromate removal from drinking water.

Bromate, which is a potential carcinogen, should be removed from drinking water to levels of less than 10 microg/L. A chitosan-based molecularly imprinted polymer (MIP) and a sol-gel ion-exchange double hydrous oxide (Fe(2)O(3) x Al(2)O(3) x xH(2)O) adsorbent (inorganic adsorbent) were prepared for this purpose. The sorption behavior of each adsorbent including sorption kinetics, isotherms, effect of pH and selective sorption were investigated in detail.

Evaluation of boronate-containing polymer brushes and gels as substrates for carbohydrate-mediated adhesion and cultivation of animal cells.

Boronate-containing thin polyacrylamide gels (B-Gel), polymer brushes (B-Brush) and chemisorbed organosilane layers (B-COSL) were prepared on the surface of glass slides and studied as substrates for carbohydrate-mediated cell adhesion. B-COSL- and B-Brush-modified glass samples exhibited multiple submicron structures densely and irregularly distributed on the glass surface, as found by scanning electron microscopy and atomic force microscopy. B-Gel was ca.

Gelatin-fibrinogen cryogel dermal matrices for wound repair: preparation, optimisation and in vitro study.

Macroporous sponge-like gelatin-fibrinogen (Gl-Fg) scaffolds cross-linked with different concentrations (0.05-0.5%) of glutaraldehyde (GA) were produced using cryogelation technology, which allows for the preparation of highly porous scaffolds without compromising their mechanical properties, and is a more cost-efficient process than freeze-drying.

Biomimetic macroporous hydrogels: protein ligand distribution and cell response to the ligand architecture in the scaffold.

Macroporous hydrogels (MHs), cryogels, are a new type of biomaterials for tissue engineering that can be produced from any natural or synthetic polymer that forms a gel. Synthetic MHs are rendered bioactive by surface or bulk modifications with extracellular matrix components. In this study, cell response to the architecture of protein ligands, bovine type-I collagen (CG) and human fibrinogen (Fg), immobilised using different methods on poly(2-hydroxyethyl methacrylate) (pHEMA) macroporous hydrogels (MHs) was analysed.

Modeling of protein breakthrough performance in cryogel columns by taking into account the overall axial dispersion.

A model considering the overall axial dispersion for describing protein adsorption and breakthrough in monolithic cryogel beds has been developed. The microstructure of cryogels was characterized by tortuous capillaries with a normal diameter distribution but a constant pore wall thickness. The axial dispersion within cryogel columns was described by using the overall axial dispersion coefficient, which can be easily obtained by matching the experimental breakthrough curves without adsorption or measuring residence time distributions (RTDs).

Dried-reswollen immobilized biocatalysts for detoxification of organophosphorous compounds in the flow systems.

New immobilized biocatalysts based on polypeptides containing N- or C-terminal polyhistidine sequences and possessing organophosphorus hydrolase activity were investigated for detoxification of organophosphorous neurotoxic compounds in the flow systems. The biocatalysts were revealed to have a high catalytic activity within wide pH and temperature ranges 7.5-12.

Building macroporous materials from microgels and microbes via one-step cryogelation.

Macroporous materials are prepared from microgels or microbes by one-step chemical cross-linking under semifrozen conditions. This avoids the use of freeze drying of the sample because a chemically stable structure is prepared under semifrozen conditions. Cryostructuration results in a material with pore walls composed of closely packed particles.

Chromato-panning: an efficient new mode of identifying suitable ligands from phage display libraries.

Background: Phage Display technology is a well established technique for high throughput screening of affinity ligands. Here we describe a new compact chromato-panning procedure for selection of suitable binders from a phage peptide display library.

Results: Both phages and E.

Biomimetic macroporous hydrogel scaffolds in a high-throughput screening format for cell-based assays.

Macroporous hydrogels (MHs) hold great promise as scaffolds in tissue engineering and cell-based assays. In this study, the possibility of combination of three-dimensional (3D) cell culture with a miniaturized screening format was demonstrated on human colon cancer HCT116, human acute myeloid leukemia KG-1 cells, and embryonic fibroblasts cultured on MHs (12.5 mm x 7.

Bilirubin recognition via molecularly imprinted supermacroporous cryogels.

Recent years molecular imprinting has received considerable attention as an excellent and simple approach to recognize small molecules and bioactive substances. The aim of this study is to prepare the bilirubin-imprinted supermacroporous cryogels which can be used for the adsorption of bilirubin from human plasma. N-methacryloyl-(L)-tyrosinemethylester (MAT) was chosen as the pre-organization monomer.

Cryogel applications in microbiology.

There is a great demand for improved technologies with regard to rapid processing of nano- and microparticles. The handling of viruses in addition to microbial and mammalian cells requires the availability of appropriate adsorbents. Recent developments in macroporous gels produced at subzero temperatures (known as cryogels) have demonstrated an efficiency for processing cell and virus suspensions, cell separation and cell culture applications.

Affinity processing of cell-containing feeds using monolithic macroporous hydrogels, cryogels.

Monolithic macroporous hydrogels, "cryogels," are produced by polymerization in a partially frozen state when the ice crystals perform as a porogen. Cryogels have a unique combination of properties: (i) large (10-100 microm) pores; (ii) minimal non-specific interactions due to the hydrophilic nature of the polymers; (iii) porosities exceeding 80-90%; (iv) good mechanical stability. These properties of cryogels allow for their application for direct capture of extracellularly expressed histidine-tagged protein from the fermentation broth and separation of different cell types.

Affinity precipitation of proteins using metal chelates.

Metal affinity precipitation has been successfully developed as a simple purification process for the proteins that have affinity for the metal ions. The copolymers of vinylimidazole with N-isopropylacrylamide are easily synthesized by radical polymerization. When loaded with Cu(II) and Ni(II) ions, these copolymers are capable of selectively precipitating proteins with natural metal-binding groups or histidine-tagged recombinant proteins.