Optimization and mathematical modeling of the transtubular bioreactor for the production of monoclonal antibodies from a hybridoma cell line

This report describes the use of a transtubular bioreactor to study the relative effects of diffusion versus perfusion of medium on antibody production by a hybridoma cell line. The study was performed with a high-density cell culture maintained in a serum-free, low-protein medium for 77 days. It was determined that the reactor possessed a macro-mixing pattern residence time distribution similar to a continuous stirred tank reactor (CSTR). However, due to the arrangement of the medium lines in the reactor, the flow patterns for nutrient distribution consist of largely independent medium path lengths ranging from short to long. When operated with cyclic, reversing, transtubular medium flow, some regions of the reactor (with short residence times) are more accessible to medium than others (with long residence times). From this standpoint, the reactor can be divided into three regions: a captive volume, which consists of medium primarily delivered via diffusion; a lapped volume, which provides nutrients through unilateral convection; and a swept volume, which operates through bilateral convection. The relative sizes of these three volumes were modified experimentally by changing the period over which the direction of medium flow was reversed from 15 min (larger captive volume) to 9 h (larger swept volume). The results suggest that antibody concentration increases as the size of the diffusion-limited (captive) volume is increased to a maximum at around 30 min with a sharp decrease thereafter. As reflected by changes in measured consumption of glucose and production of lactate, no significant difference in cellular metabolism occurred as the reactor was moved between these different states. These results indicate that the mode of operation of the transtubular bioreactor may influence antibody productivity under serum-free, low-protein conditions with minimal effects on cellular metabolism.     

Inherent and apparent optical measurements in the Hudson/Raritan Estuary

Duringan August, 1999 field campaign, measurements were made to establish hydrologicoptical properties of the Hudson/Raritan Estuary (New York-New Jersey): 1)concurrent above-and below-surface spectral irradiance; 2) sampling forlaboratory determination of inherent optical properties; and 3) concentrationsof optically-important water quality parameters. We used a bio optical modelbased on to predict thesubsurface irradiance reflectance from optically important water constituents.This model was then validated with the measured reflectance spectra from thefield spectroradiometers. Modeling of reflectance is a prerequisite forprocessing remote sensing data to desired thematic maps. These are key input tothe geographic information system (GIS) used to manage the water qualitycondition of the estuary.     

Browsing gene banks for Fe2S2 ferredoxins and structural modeling of 88 plant-type sequences: an analysis of fold and function.

One-hundred-and-seventy-nine sequences of Fe2S2 ferredoxins and ferredoxin precursors were identified in and retrieved from currently available protein and cDNA databases. On the basis of their cluster-binding patterns, these sequences were divided into three groups: those containing the CX4CX2CXnC pattern (plant-type ferredoxins), those with the CX5CX2CXnC pattern (adrenodoxins), and those with a different pattern. These three groups contain, respectively, 139, 36, and 4 sequences. After excluding ferredoxin precursors in the first group, two subgroups were identified, again based on their cluster-binding patterns: 88 sequences had the CX4CX2CX29C pattern, and 29 had the CX4CX2CXmC (m not equal 29) pattern. The structures of the 88 ferredoxins with the CX4CX2CX29C pattern were modeled based on the available experimental structures of nine proteins within this same group. The modeling procedure was tested by building structural models for the ferredoxins with known structures. The models resulted, on average, in being within 1 A of the backbone root-mean-square deviation from the corresponding experimental structures. In addition, these structural models were shown to be of high quality by using assessment procedures based on energetic and stereochemical parameters. Thus, these models formed a reliable structural database for this group of ferredoxins, which is meaningful within the framework of current structural genomics efforts. From the analysis of the structural database generated it was observed that the secondary structural elements and the overall three-dimensional structures are maintained throughout the superfamily. In particular, the residues in the hydrophobic core of the protein were found to be either absolutely conserved or conservatively substituted. In addition, certain solvent-accessible charged groups, as well as hydrophobic groups, were found to be conserved to the same degree as the core residues. The patterns of conservation of exposed residues identified the regions of the protein that are critical for its function in electron transfer. An extensive analysis of protein-protein interactions is now possible. Some conserved interactions between residues have been identified and related to structural and/or functional features. All this information could not be obtained from the analyses of the primary sequences alone. Finally, the analysis of the sequences of the related subgroup featuring the CX4CX2CXmC (m not equal 29) cluster-binding pattern in the light of the structural and functional insights provided by the inspection of the mentioned structural database affords some hints on the functional features of ferredoxins belonging to this subgroup.     

Effects of age and distance on the composition of mixed deciduous forest fragments in an agricultural landscape

Abstract. Forest patches in central Belgium were inventoried twice for the presence or absence of forest plant species to study the effects of age and distance on species composition. All forests in the study area were subdivided based on their land use history. To avoid effects of autocorrelated environmental characteristics on species composition, habitat homogeneity was indirectly investigated using a TWINSPAN classification of the vegetation data. Two major habitats (alluvial and non‐alluvial forests) were distinguished and analysed separately. Patterns of species composition were investigated at the landscape level using Mantel tests. Species composition similarity measures were calculated between all pairs of fragments based on the floristic data. A highly significant correlation was found between species composition similarity and inter‐patch distance. Difference in age, which we used as a measure for habitat quality, was less important in explaining species composition patterns. The effects of spatial configuration became significant when difference in age was accounted for, and a partial correlation was calculated between inter‐patch distance and species composition similarity. Different results were found for alluvial and non‐alluvial forest types. Alluvial forests were more influenced by the spatial configuration than the non‐alluvial. For the non‐alluvial forest type effects measured with the difference in age between forest fragments obscured the effects of inter‐patch distance. Based on our findings we suggest that species composition is not only internally generated, but external processes such as differential colonization caused by varying degrees of isolation may be of overriding importance.     

Antifreeze protein from shorthorn sculpin: Identification of the ice-binding surface

Shorthorn sculpins, Myoxocephalus scorpius, are protected from freezing in icy seawater by alanine-rich, alpha-helical antifreeze proteins (AFPs). The major serum isoform (SS-8) has been reisolated and analyzed to establish its correct sequence. Over most of its length, this 42 amino acid protein is predicted to be an amphipathic alpha-helix with one face entirely composed of Ala residues. The other side of the helix, which is more heterogeneous and hydrophilic, contains several Lys. Computer simulations had suggested previously that these Lys residues were involved in binding of the peptide to the [11-20] plane of ice in the <-1102> direction. To test this hypothesis, a series of SS-8 variants were generated with single Ala to Lys substitutions at various points around the helix. All of the peptides retained significant alpha-helicity and remained as monomers in solution. Substitutions on the hydrophilic helix face at position 16, 19, or 22 had no obvious effect, but those on the adjacent Ala-rich surface at positions 17, 21, and 25 abolished antifreeze activity. These results, with support from our own modeling and docking studies, show that the helix interacts with the ice surface via the conserved alanine face, and lend support to the emerging idea that the interaction of fish AFPs with ice involves appreciable hydrophobic interactions. Furthermore, our modeling suggests a new N terminus cap structure, which helps to stabilize the helix, whereas the role of the lysines on the hydrophilic face may be to enhance solubility of the protein.     

Synthesis and structural study of a fully protected α-C-galactosyl model dipeptide

Summary The synthesis of three N-alkyl-6,7-dideoxy-1,2:3,4-di-O-isopropylidene-7-[(alkyl-carbonyl)amino]-L-glycero-α-D-galacto-octopyranuronamides6a-c, analogous model dipeptides containing two amide groups connected to the α-carbon bearing the fully protected galactose as a side chain, has been realized with the aim of determining the conformational influence of the galactosyl moiety on the peptide backbone. Molecular modeing of6a, X-ray crystallography of6c and IR and NMR experiments on6a-c in organic solvents show that the carbohydrate ring assumes a twist boat conformation. In non-polar organic solvents, the NH of the left amide group interacts with one ketal oxygen of the galactosyl group.     

Migration of Naphthalene through Low Organic Content Sandy Soil Columns: Comparison of Unsaturated and Saturated Conditions

The transient transport of naphthalene through low organic matter content soil columns was investigated in different water-saturation and flow conditions. Some parameters were tested as flow rate, column height, and water saturation conditions. The soil was a clayed sandy soil from the Algerian coast near Boumerdes. The organic carbon content was 0.13% and the main mineral components were quartz (88%), clays minerals (7%) and calcite (3%). The height of the packing of the soil column (5.1 cm in diameter) varied from 15 to 40 cm. Simultaneous step injections of inert tracer (calcium chloride) and naphthalene at 10 mg L^?1 were performed. Tracer and naphthalene breakthrough curves (BTCs) were measured continuously by conductimetry and UV – 220 nm, respectively. The BTCs were simulated using the classical mixing cells in series with exchange model (MCE). In unsaturated conditions the comparison of the mean residence time of tracer BTCs with the geometrical pore volume gave us access to average water saturation along the column as a function of height. The higher the soil bed was, the higher the mean water saturation. The comparison of naphthalene distribution coefficients (K_d) in different flow conditions with the theoretical value from the Karickhoff law showed that in saturated conditions the obtained value was close to the theoretical one. In unsaturated conditions, the measured naphthalene K_d's were much lower than the theoretical value and correlated to the water saturation.     

Process-oriented ecological modeling approach and scientific workflow system

A scientific workflow system is designed specifically to organize, manage and execute a series of research steps, or a workflow, in a given runtime environment. The vision for scientific workflow systems is that the scientists around the world can collaborate on designing global-scaled experiments, sharing the data sets, experimental processes, and results on an easy-to-use platform. Each scientist can create and execute their own workflows and view results in real-time, and then subsequently share and reuse workflows among other scientists. Two case studies, using the Kepler system and BioVeL, are introduced in this paper. Ecological niche modeling process, which is a specialized form of scientific workflow system included in both Kepler system and BioVeL, was used to describe and discuss the features, developmental trends, and problems of scientific workflows.     

State-space modeling clarifies productivity regime shifts of Japanese flying squid

Regime shifts of climatic and environmental conditions potentially affect the productivity of fishery resources, posing challenges in stock management. The stocks of the Japanese flying squid (Todarodes pacificus) are suspected to suffer from regime shifts, but detecting the occurrence of regime shifts in this species is generally difficult and unreliable because the short-lived nature of this species inherently confounds the effect of regime shifts with observation and process errors. Here we developed a new state-space assessment model to evaluate the influence of regime shifts on the spawner-recruit relationship of the Japanese flying squid. The model simultaneously estimates the population dynamics of multiple stocks that could share some life history parameters, thereby stabilizing parameter inference. We demonstrate that two regime shifts in productivity around 1991 and 2015 caused two- to threefold changes of maximum sustainable yields. The model with regime shifts clarifies the relationship between fishing pressure and spawner abundance that is difficult to detect in a model with no regime shift. The state-space approach is a promising tool for accurately assessing stock status by separating the recruitment process from observation errors and is expected to contribute to the effective management of marine biological resources sensitive to regime shifts.