Magnon excitation by spin injection in thin Fe/Cr/Fe films

The technique of Brillouin light scattering is used to observe strong excitation of magnons in antiferromagnetically coupled trilayers of Fe/Cr/Fe at room temperature. The magnons are driven out of equilibrium by a microwave current applied in the trilayer through point contacts. The magnitude of the scattering intensity is investigated as a function of the magnon wave number and applied magnetic field. Confirming recent theoretical predictions, the observations provide strong evidence of electronic spin injection in the rf driving field.     

Macroporous molecularly imprinted polymer/cryogel composite systems for the removal of endocrine disrupting trace contaminants

A new concept for the preparation of selective sorbents with high flow path properties is presented by embedding molecularly imprinted polymers (MIPs) into various macroporous gels (MGs). A MIP was first synthetized with 17beta-estradiol (E2) as template for the selective adsorption of this endocrine disrupter. The composite macroporous gel/MIP (MG/MIP) monoliths were then prepared at subzero temperatures. Complete recovery of E2 from a 2 microg/L aqueous solution was achieved using the polyvinyl alcohol (PVA) MG/MIP monoliths whereas only 49-74% was removed with non-imprinted polymers (when no template was used). The PVA MG/MIP monolith columns were operated at almost 10 times higher flow rate (50 mL/min) compared to the MIP columns with operation flow rate of 1-5 mL/min. The possibility for processing the particulate containing wastewater effluents at high flow rates with selectivity on E2 removal, as well as the easy preparation of the monoliths, make the macroporous MG/MIP systems attractive and robust sorbents for the clean up of water from endocrine disrupting trace contaminants.     

Effect of matrix elasticity on affinity binding and release of bioparticles. Elution of bound cells by temperature-induced shrinkage of the smart macroporous hydrogel

The first step of bacterial or viral invasion is affinity and presumably multisite binding of bioparticles to an elastic matrix like a living tissue. We have demonstrated that model bioparticles such as inclusion bodies (spheres of about 1 microm in size) Escherichia coli cells (rods 1 x 3 microm), yeast cells (8 microm spheres), and synthetic microgel particles (0.4 microm spheres) are binding via different affinity interactions (IgG antibody-protein A, sugar-lectin, and metal ion-chelate) to a macroporous hydrogel (MH) matrix bearing appropriate ligands. The elastic deformation of the MH results in the detachment of affinity bound bioparticles. The particle detachment on elastic deformation is believed to be due to multipoint attachment of the particles to affinity matrix and the disturbance of the distance between affinity ligands when the matrix is deformed. No release of affinity bound protein occurred on elastic deformation. The efficiency of the particle release by the elastic deformation depends on the density of the ligands at the particle surface as well as on the elasticity of the matrix for relatively large particles. The release of the particles occurred irrespectively of whether the deformation was caused by external forces (mechanical deformation) or internal forces (the shrinkage of thermosensitive macroporous poly-N-isopropylacrylamide hydrogel on increase in temperature).     

Immobilised peptide displaying phages as affinity ligands. Purification of lactoferrin from defatted milk

An affinity purification procedure for the direct purification of lactoferrin from defatted (skimmed) milk has been developed. The procedure is based on using selected phage clones expressing a peptide with high binding affinity for lactoferrin which were covalently coupled to macroporous poly(dimethylacrylamide) monolithic column. Large pore size (10-100 microm) of macroporous poly(dimethylacrylamide) makes it possible to couple long (1 microm) phage particles as ligands without any risk of blocking the monolithic column. Bound lactoferrin was eluted using 1M NaCl with a purity of >95%. The technique presents a good alternative to conventional immunoaffinity chromatography for purification of a protein of interest from complex samples due to (i) the robustness of the system in terms of recovery and ligand leakage and (ii) economical aspect in terms of low ligand cost.     

Characterization of polyacrylamide based monolithic columns

Supermacroporous monolithic polyacrylamide (pAAm)-based columns have been prepared by radical cryo-copolymerization (copolymerization in the moderately frozen system) of acrylamide with functional co-monomer, allyl glycidyl ether (AGE), and cross-linker N,N'-methylene-bis-acrylamide (MBAAm) directly in glass columns (ID 10 mm). The monolithic columns have uniform supermacroporous sponge-like structure with interconnected supermacropores of pore size 5-100 microm. The monoliths can be dried and stored in the dry state. High mechanical stability of the monoliths allowed sterilization by autoclaving. Column-to-column reproducibility of pAAm-monoliths was demonstrated on 5 monolithic columns from different batches prepared under the same cryostructuration conditions.     

High throughput processing of particulate-containing samples using supermacroporous elastic monoliths in microtiter (multiwell) plate format

Two steps in parallel processing of multiple biosamples, namely, sample clarification and capture of the target protein, were integrated and combined with the direct assay of captured protein using a newly developed microtiter (96-well) plate system based on the monoliths of hydrophilic elastic supermacroporous material, cryogel. Cryogel monoliths have pore size large enough for microbial and mammalian cells to pass through unretained. Moreover, cryogel monoliths are elastic allowing them to be slightly compressed and easily introduced into the wells. When expanded, cryogel monoliths fill the well tightly with no risk of leakage in between the monolith and the walls of the well. The capillary forces keep the liquid inside the pores of the cryogel monolith making the monolith columns drainage protected. The application of a certain volume of liquid on top of a cryogel monolith column results in the displacement of exactly the same volume of liquid from the column. The concept of using supermacroporous gels in 96-well plate format offers new possibilities to the biotechnologist allowing separation of particulate matter, capturing of soluble material from particle containing media, and parallel assay of large number of non-clarified samples.     

Direct chromatographic capture of enzyme from crude homogenate using immobilized metal affinity chromatography on a continuous supermacroporous adsorbent

A continuous supermacroporous matrix has been developed allowing direct capture of enzyme from non-clarified crude cell homogenate at high flow-rates. The continuous supermacroporous matrix has been produced by radical co-polymerization of acrylamide, allyl glycidyl ether and N,N'-methylene-bis(acrylamide) which proceeds in aqueous solution of monomers frozen inside a column (cryo-polymerization). After thawing, the column contains a continuous matrix having interconnected pores of 10-100 microm size. Iminodiacetic acid covalently coupled to the cryogel is a rendering possibility for immobilized metal affinity chromatographic purification of recombinant His-tagged lactate dehydrogenase, (His)6-LDH, originating from thermophilic bacterium Bacillus stearothermophilus, but expressed in Escherichia coli. The large pore size of the adsorbent makes it possible to process particulate-containing material without blocking the column. No preliminary filtration or centrifugation is needed before application of crude extract on the supermacroporous column. A total of 210 ml crude homogenate, 75 ml of it non-clarified, was processed on a single 5.0 ml supermacroporous column at flow speeds up to 12.5 ml/min without noticeable impairment of the column properties. Mechanically the cryogel adsorbent is very stable. The continuous matrix could easily be removed from the column, dried at 70 degrees C and kept in a dry state. After rehydration and reinsertion of the matrix into an empty column, (His)6-LDH was purified as efficiently as on the newly prepared column. The procedure of manufacturing the supermacroporous continuous cryogel is technically simple. Starting materials and initiators are cheap and available and are simply mixed and frozen under specified conditions. Altogether these qualities reveal that the supermacroporous continuous cryogels is a very interesting alternative to existing methods of protein purification from particulate-containing crude extracts.