Chromatographic behaviour of monoclonal antibodies against wild-type amidase from Pseudomonasaeruginosa on immobilized metal chelates

The aim of this work was to devise a one-step purification procedure for monoclonal antibodies (MAbs) of IgG class by immobilized metal affinity chromatography (IMAC). Therefore, several stationary phases were prepared containing immobilized metal chelates in order to study the chromatographic behaviour of MAbs against wild-type amidase from Pseudomonas aeruginosa. Such MAbs adsorbed to Cu(II), Ni(II), Zn(II) and Co(II)-IDA agarose columns. The increase in ligand concentration and the use of longer spacer arms and higher pH values resulted in higher adsorption of MAbs into immobilized metal chelates. The dynamic binding capacity and the maximum binding capacity were 1.33 +/- 0.015 and 3.214 +/- 0.021 mg IgG/mL of sedimented commercial matrix, respectively. A K(D) of 4.53 x 10(-7) M was obtained from batch isotherm measurements. The combination of tailor-made stationary phases of IMAC and the correct selection of adsorption conditions permitted a one-step purification procedure to be devised for MAbs of IgG class. Culture supernatants containing MAbs were purified by IMAC on commercial-Zn(II) and EPI-30-IDA-Zn(II) Sepharose 6B columns and by affinity chromatography on Protein A-Sepharose CL-4B. This MAb preparation revealed on SDS-PAGE two protein bands with M(r) of 50 and 22 kDa corresponding to the heavy and light chains, respectively. Copyright (C) 2011 John Wiley & Sons, Ltd.

Multi-scaled femtosecond laser structuring of stationary titanium surfaces

The evolution of the topography of titanium surfaces treated with femtosecond laser radiation in stationary conditions as a function of radiation fluence and number of laser pulses is investigated. Depending on the processing parameters, ripples, microcolumns, wavy or smooth surfaces can be obtained. The ripples predominate for fluences near the damage threshold of titanium (0.2+/-0.1) J/cm(2), while microcolumns form during the first 200 pulses for fluences between (0.6+/-0.2) and (1.7+/-0.2) J/cm(2). A wavy topography develops for fluences and number of pulses higher than (1.7+/-0.2) J/cm(2) and 300, respectively. A bimodal surface topography consisting of surface ripples overlapping a microcolumnar topography can be obtained if the surfaces are firstly treated to create microcolumns followed by laser treatment with a lower fluence near the ablation threshold of the material, in order to generate periodic ripple

Laser-assisted deposition of thin films from photoexcited vapour phases

Laser-assisted chemical vapour deposition (LCVD) has been extensively studied in the last two decades. A vast range of applications encompass various areas such as microelectronics, micromechanics, microelectromechanics and integrated optics, and a variety of metals, semiconductors and insulators have been grown by LCVD. In this article, we review briefly the LCVD process and present two case studies of thin film deposition related to laser thermal excitation (e.g., boron carbide) and non-thermal excitation (e.g., CrO(2)) of the gas phase.

Regulation of antioxidant enzymes gene expression in the yeast Saccharomyces cerevisiae during stationary phase

Gene expression of three antioxidant enzymes, Mn superoxide dismutase (MnSOD), Cu,Zn superoxide dismutase (Cu,ZnSOD), and glutathione reductase (GR) was investigated in stationary phase Saccharomyces cerevisiae during menadione-induced oxidative stress. Both GR and Cu,ZnSOD mRNA steady state levels increased, reaching a plateau at about 90 min exposure to menadione. GR mRNA induction was higher than that of Cu,ZnSOD (about 14-fold and 9-fold after 90 min, respectively). A different pattern of response was obtained for MnSOD mRNA, with a peak at about 15 min (about 8-fold higher) followed by a decrease to a plateau approximately 4-fold higher than the control value. However, these increased mRNA levels did not result in increased protein levels and activities of these enzymes. Furthermore, exposure to menadione decreased MnSOD activity to half its value, indicating that the enzyme is partially inactivated due to oxidative damage. Cu,ZnSOD protein levels were increased 2-fold, but MnSOD protein levels were unchanged after exposure to menadione in the presence of the proteolysis inhibitor phenylmethylsulfonyl fluoride. These results indicate that the rates of Cu,ZnSOD synthesis and proteolysis are increased, while the rates of MnSOD synthesis and proteolysis are unchanged by exposure to menadione. Also, the translational efficiency for both enzymes is probably decreased, since increases in protein levels when proteolysis is inhibited do not reflect the increases in mRNA levels. Our results indicate that oxidative stress modifies MnSOD, Cu,ZnSOD, and GR gene expression in a complex way, not only at the transcription level but also at the post-transcriptional, translational, and post-translational levels.

Longitudinal versus polar wrinkling of core-shell fibers with anisotropic size mismatches

We consider a fiber made of a soft elastic material, encased in a stiff elastic shell (core-shell geometry). If the core and shell dimensions are mismatched, e.g., because the core shrinks while the shell does not, but the two remain attached, then an elastic instability is triggered whereby wrinkles may appear on the shell. The wrinkle orientation may be longitudinal (along the fiber axis), polar (along the fiber perimeter), or a mixture of both, depending on the fiber's geometrical and material parameters. Here we investigate under what conditions longitudinal or polar wrinkling will occur.