Many successful, recent therapies for life-threatening diseases such as cancer and rheumatoid arthritis are based on the recognition between native or genetically engineered antibodies and cell-surface receptors. Although naturally produced by the immune system, the need for antibodies with unique specificities and designed for single application, has encouraged the search for novel antibody purification strategies. The availability of these products to the end-consumer is strictly related to manufacture costs, particularly those attributed to downstream processing. Over the last decades, academia and industry have developed different types of interactions and separation techniques for antibody purification, affinity-based strategies being the most common and efficient methodologies. The affinity ligands utilized range from biological to synthetic designed molecules with enhanced resistance and stability. Despite the successes achieved, the purification “paradigm” still moves interests and efforts in the continuous demand for improved separation performances. This review will focus on recent advances and perspectives in antibody purification by affinity interactions using different techniques, with particular emphasis on affinity chromatography.

Most of the discussion and controversy on organisation of work concepts has been referenced to the manufacturing industry along the 20th century: it started with the concept of “scientific management” from Taylor, and continued with the new ideas on the importance of human factors as Mayo pointed out in the 1930s. Immediately after the 2nd World War Friedmann studied the human problems related to new manufacturing technologies and automation. And the late 1950 and 1960s were decades of strong debate on the socio-technics with the research at Tavistock Institute of London and the emergence of national programmes on new forms of work organisation. At the end of the last century the concept of collaborative work was developed together with the definition(s) of information systems and organisational design. However, the interest came from other production activities, like the services. This article analyses the approaches developed on these debates on the collaborative work and information system and its application to the manufacturing industry.

The simultaneous separation and quantification of two casein peptides {(IPP}, {VPP)} presenting potent inhibitory activity of angiotensin-converting-enzyme {(ACE)} and casein in fermented milks was developed. Gradient elution was carried out at a flow-rate of 1 {mL/min}, using a mixture of two solvents. Solvent A was 0.1% {TFA} in water and solvent B was acetonitrile-water-trifluoracetic acid 95:5:0.1. The effluent was monitored by {UV} detector at 214 nm. Calibration curves were constructed in the interval of 0.01-1.0 {mg/mL} for {VPP}, 0.005-1.0 {mg/mL} for {IPP}, and 0.05-3.0 {mg/mL} for casein. R2 invariably exceeded 0.999. The detection limits were 0.004 for {VPP}, 0.002 {mg/mL} for {IPP}, and 0.02 {mg/mL} for casein. Repeatability of the method was evaluated by six consecutive injections of two standard solutions containing {VPP}, {IPP}, and casein. The {RSD} values for concentration were all below 5.08%. Recovery studies were carried out to determine the accuracy of the method. Recoveries ranged between 88 and 98.2%. The methodology was applied, not only, for the monitorization of {VPP}, {IPP}, and casein in commercial fermented milks labeled as presenting antihypertensive properties, but also, in milk with different degrees of fermentation by L. Helveticus, and in other commercial functional fermented milks, such as, those presenting cholesterol lowering properties.

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The time-varying microstructure of sleep EEG spindles may have clinical significance in dementia studies. In this work, the sleep spindle is modeled as an AM-FM signal and parameterized in terms of six parameters, three quantifying the instantaneous envelope (IE) and three quantifying the instantaneous frequency (IF) of the spindle model. The IE and IF waveforms of sleep spindles from patients with dementia and normal controls were estimated using the time-frequency technique of complex demodulation (CD). Sinusoidal curve-fitting using a matching pursuit (MP) approach was applied to the IE and IF waveforms for the estimation of the six model parameters. Specific differences were found in sleep spindle instantaneous frequency dynamics between spindles from dementia subjects and spindles from controls.

Synthetic affinity ligands can circumvent the drawbacks of natural immunoglobulin (Ig)-binding proteins by imparting resistance to chemical and biochemical degradation and to in situ sterilization, as well as ease and low cost of production. Protein L (PpL), isolated from Peptostreptococcus magnus strains, interacts with the Fab (antigen-binding fragment) portion of Igs, specifically with kappa light chains, and represents an almost universal ligand for the purification of antibodies. The concepts of rational design and solid-phase combinatorial chemistry were used for the discovery of a synthetic PpL mimic affinity ligand. The procedure presented in this chapter represents a general approach with the potential to be applied to different systems and target proteins.

Multiheme c-type cytochromes from members of the Desulfovibrionacea and Geobactereacea families play crucial roles in the bioenergetics of these microorganisms. Thermodynamic studies using NMR and visible spectroscopic techniques on tetraheme cytochromes c3 isolated from Desulfovibrio spp. and more recently on a triheme cytochrome from Geobacter sulfurreducens showed that the properties of each redox centre are modulated by the neighbouring redox centres enabling these proteins to perform energy transduction and thus contributing to cellular energy conservation. Electron/proton transfer coupling relies on redox-linked conformational changes that were addressed for some multiheme cytochromes from the comparison of protein structure of fully reduced and fully oxidised forms. In this work, we identify for the first time in a multiheme cytochrome the simultaneous presence of two different conformations in solution. This was achieved by probing the different oxidation stages of a triheme cytochrome isolated from G. sulfurreducens using 2D-NMR techniques. The results presented here will be the foundations to evaluate the modulation of the redox centres properties by conformational changes that occur during the reoxidation of a multiheme protein.

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Generalised fractional centred differences and derivatives are studied in this Chapter. These generalise to real orders the existing ones valid for even and odd positive integer orders. For each one, suitable integral formulations are presented. The limit computation inside the integrals leads to generalisations of the Cauchy derivative. Their computations using a special path lead to the well known Riesz potentials. A study for coherence is done by applying the definitions to functions with Fourier transform. The existence of inverse Riesz potentials is also studied.

Thermal perturbation of the dicluster ferredoxin from Acidianus ambivalens was investigated employing a toolbox of spectroscopic methods. FTIR and visible CD were used for assessing changes of the secondary structure and coarse alterations of the [3Fe4S] and [4Fe4S] cluster moieties, respectively. Fine details of the disassembly of the metal centers were revealed by paramagnetic NMR and resonance Raman spectroscopy. Overall, thermally induced unfolding of AaFd is initiated with the loss of α-helical content at relatively low temperatures (Tapp (m) ~ 44 °C), followed by the disruption of both iron−sulfur clusters (Tapp (m) ~ 53−60 °C). The degradation of the metal centers triggers major structural changes on the protein matrix, including the loss of tertiary contacts (Tapp (m) ~ 58 °C) and a change, rather than a significant net loss, of secondary structure (Tapp (m) ~ 60 °C). This latter process triggers a secondary structure reorganization that is consistent with the formation of a molten globule state. The combined spectroscopic approach here reported illustrates how changes in the metalloprotein organization are intertwined with disassembly of the iron−sulfur centers, denoting the conformational interplay of the protein backbone with cofactors.