Sequencing of parts and robot moves in a robotic cell

In this paper, we deal with the problem of sequencing parts and robot moves in a robotic cell where the robot is used to feed machines in the cell. The robotic cell, which produces a set of parts of the same or different types, is a flow-line manufacturing system. Our objective is to maximize the long-run average throughput of the system subject to the constraint that the parts are to be produced in proportion of their demand. The cycle time formulas are developed and analyzed for this purpose for cells producing a single part type using two or three machines. A state space approach is used to address the problem. Both necessary and sufficient conditions are obtained for various cycles to be optimal. Finally, in the case of many part types, the problem of scheduling parts for a specific sequence of robot moves in a two machine cell is formulated as a solvable case of the traveling salesman problem.     

Single cell deposition and patterning with a robotic system

Integrating single-cell manipulation techniques in traditional and emerging biological culture systems is challenging. Microfabricated devices for single cell studies in particular often require cells to be spatially positioned at specific culture sites on the device surface. This paper presents a robotic micromanipulation system for pick-and-place positioning of single cells. By integrating computer vision and motion control algorithms, the system visually tracks a cell in real time and controls multiple positioning devices simultaneously to accurately pick up a single cell, transfer it to a desired substrate, and deposit it at a specified location. A traditional glass micropipette is used, and whole- and partial-cell aspiration techniques are investigated to manipulate single cells. Partially aspirating cells resulted in an operation speed of 15 seconds per cell and a 95% success rate. In contrast, the whole-cell aspiration method required 30 seconds per cell and achieved a success rate of 80%. The broad applicability of this robotic manipulation technique is demonstrated using multiple cell types on traditional substrates and on open-top microfabricated devices, without requiring modifications to device designs. Furthermore, we used this serial deposition process in conjunction with an established parallel cell manipulation technique to improve the efficiency of single cell capture from ～80% to 100%. Using a robotic micromanipulation system to position single cells on a substrate is demonstrated as an effective stand-alone or bolstering technology for single-cell studies, eliminating some of the drawbacks associated with standard single-cell handling and manipulation techniques.     

Exact methods for the robotic cell problem

This paper investigates an exact method for the Robotic Cell Problem. We present a branch-and-bound algorithm which is the first exact procedure specifically designed with regard to this complex flow shop scheduling variant. Also, we propose a new mathematical programming model as well as new lower bounds. Furthermore, we describe an effective genetic algorithm that includes, as a mutation operator, a local search procedure. We report the results of a computational study that provides evidence that medium-sized instances, with up to 176 operations, can be optimally solved. Also, we found that the new proposed lower bounds outperform lower bounds from the literature. Finally, we show, that the genetic algorithm delivers good solutions while requiring short CPU times.     

Metabolite production and growth efficiency

The capacity to sustain the large fluxes of carbon and energy required for rapid metabolite production appears to be inversely related to the growth efficiency of micro-organisms. From an overall energetic point of view three main classes of metabolite may be distinguished. These are not discrete categories, as the energetics of biosynthesis will depend on the precise biochemical pathways used and the nature of the starting feed stock(s). (1) for metabolites like exopolysaccharides both the oxidation state and the specific rate of production appear to be inversely related to the growth efficiency of the producing organism. Maximum rates of production are favored when carbon and energy flux are integrated, and alteration of this balance may negatively effect production rates. (2) The production of metabolites like organic acids and some secondary metabolites results in the net production of reducing equivalents and/or ATP. It is thought that the capacity of the organism to dissipate this product associated energy limits its capacity for rapid production. (3) For metabolites like biosurfactants and certain secondary metabolites that are composed of moieties of significantly different oxidation states production from a single carbon source is unfavorable and considerable improvements in specific production rate and final broth concentration may be achieved if mixed carbon sources are used. By careful selection of production organism and starting feedstock(s) it may be possible to tailor the production, such that the adverse physiological consequences of metabolite overproduction on the production organism are minimized.     

Measuring the efficiency of sound production

Sound production efficiency is a complex phenotypic trait that incorporates biochemical and mechanical events beginning with substrate oxidation and ending with the radiation of sound. Its accurate measurement is significant in understanding the mechanisms and energetics underlying acoustic signaling and sexual selection. I show that in the short-tailed cricket Anurogryllus arboreus Walker, acoustic performance is apparently the same in acoustic free fields and in the reverberant conditions of a respirometry chamber. I present three methods for simultaneous and nearly simultaneous determination of calling metabolic rate and acoustic power output. The new methods yielded metabolic rates 3%-6% lower than matched controls using traditional flow-through respirometry (mean=8.1 mW); however, none of theses differences were statistically significant. I also evaluate four methods for determining the efficiency of sound production. The means of an individual's efficiencies calculated using these methods vary between 0.50% and 0.60%, with no statistically significant differences between the methods. I conclude with a critical evaluation of these techniques.     

The RNA polymerase factory: a robotic in vitro assembly platform for high-throughput production of recombinant protein complexes

The in-depth structure/function analysis of large protein complexes, such as RNA polymerases (RNAPs), requires an experimental platform capable of assembling variants of such enzymes in large numbers in a reproducible manner under defined in vitro conditions. Here we describe a streamlined and integrated protocol for assembling recombinant archaeal RNAPs in a high-throughput 96-well format. All aspects of the procedure including construction of redesigned expression plasmids, development of automated protein extraction/in vitro assembly methods and activity assays were specifically adapted for implementation on robotic platforms. The optimized strategy allows the parallel assembly and activity assay of 96 recombinant RNAPs (including wild-type and mutant variants) with little or no human intervention within 24 h. We demonstrate the high-throughput potential of this system by evaluating the side-chain requirements of a single amino acid position of the RNAP Bridge Helix using saturation mutagenesis.     

Cytokine production by a megakaryocytic cell line

Summary The regulation of megakaryopoeisis by cytokines is not yet well understood. It is possible that autocrine loops are established during megakaryocyte growth and differentiation, aiding in the maturation of these cells. The CHRF-288-11 human megakaryoblastic cell line has been examined for cytokine production in growing cells and cells stimulated to differentiate by the addition of phorbol esters. It has been demonstrated that these cells produce RNA corresponding to the interleukins IL-1α, 1β, 3, 7, 8, and 11, granulocyte-macrophage colony stimulating factor (GM-CSF), stem cell factor (SCF), transforming growth factor-β (TGF-β), tumor necrosis factor-α (TNF-α), interferon-α (INF-α), and basic fibroblast growth factor (bFGF). Additionaly, RNA corresponding to the receptors for IL-6, GM-CSF, SCF, INF-α,β, bFGF, and monocyte colony stimulating factor (M-CSF) were also expressed by the cells. The receptor for TNF-α was detected immunologically. Analysis at the protein level demonstrated that significant amounts of INF-α, TNF-α, GM-CSF, SCF, IL-1α, and a soluble form of the IL-6 receptor were produced by the cells. Addition of phorbol esters to CHRF-288-11 cells enhances their megakaryocytic phenotype; such treatment also results in increased secretion of INF-α, TNF-α, and GM-CSF. These results suggest that potential autocrine loops are established during the differentiation of CHRF-288-11 cells, which may alter the capability of the cell to differentiate. These findings are similar to those recently obtained for marrow-derived megakaryocytes (Jiang et al.) suggesting that CHRF-288-11 cells provide a useful model system for the study of cytokine release during megakaryocyte differentiation.     

Wheat production efficiency from Soviet virgin lands

The direction of the time trend in wheat production efficiency from Soviet arid virgin lands is analyzed with the use of a developed agrohydrologic model. It is based on the concepts of water resource input (soil moisture) and corresponding agro output (spring wheat). The model evaluates each growing season and assigns a decimal value (from 0.0 to 1.0) which is then multiplied by maximum spring wheat yield. This derived product is that growing season's estimated yield. A nonparametric statistical method by Kendall is applied to examine the study period wheat production efficiency time trend. It is suggested that an increasing trend occurred over the study period.     

Bioelectricity production using a new electrode in a microbial fuel cell

Electrode materials play a key role in enhancing the electricity generation in the microbial fuel cell (MFC). In this study, a new material (Ti-TiO(2)) was used as an anode electrode and compared with a graphite electrode for electricity generation. Current densities were 476.6 and 31 mA/m(2) for Ti-TiO(2) and graphite electrodes, respectively. The PCR-DGGE analysis of enriched microbial communities from estuary revealed that MFC reactors were dominated by Shewanella haliotis, Enterococcus sp., and Enterobacter sp. Bioelectrochemical kinetic works in the MFC with Ti-TiO(2) electrode revealed that the parameters by non-linear curve fitting with the confidence bounds of 95% gave good fit with the kinetic constants of η (difference between the anode potential and anode potential giving one-half of the maximum current density) = 0.35 V, K (s) (Half-saturation constant) = 2.93 mM and J (max) = 0.39 A/m(2) for T = 298 K and F = 96.485 C/mol-e(-). From the results observed, it is clear that Ti-TiO(2) electrode is a promising candidate for electricity generation in MFC.     

Size-dependent scattering efficiency in dye-sensitized solar cell

Effect of scattering particle size on light scattering efficiency in dye-sensitized solar cell has been investigated with a FTO/semitransparent nano-TiO₂ layer (main-layer)/scattering layer (overlayer) structure, where two different rutile TiO₂ particles of 0.3 μm (G1) and 0.5 μm (G2) were used for a scattering overlayer and 20-nm anatase particle for nano-TiO₂ main-layer. The conversion efficiency of 7.55% for the 7 μm-thick main-layer film is improved to 8.94% and 8.78% when G1 and G2 particulate overlayers are introduced, respectively, corresponding to 18.4% and 16.3% increments. While the conversion efficiency of the 14 μm-thick main-layer is slightly improved from 8.60% to 9.09% and 9.15% upon depositing G1 and G2 particulate overlayers, respectively. Significant improvement and strong size-dependence upon deposition of scattering overlayer on the relatively thinner TiO₂ main-layer film are associated with the quantity and wavelength of transmitted light and the difference in reflectivity of G1 and G2 scattering particles.     

High efficiency bioethanol production from barley straw using a continuous pretreatment reactor

We developed a new pretreatment process for producing high-efficiency bioethanol from a lignocellulosic biomass. Barley straw was pretreated with sodium hydroxide in a twin-screw extruder for continuous pretreatment. The biomass to ethanol ratio (BTER) for optimal pretreatment conditions was evaluated by response surface methodology. Simultaneous saccharification and fermentation (SSF) was conducted to investigate the BTER with 30 FPU/g cellulose of enzyme and 7% (v/v) yeast (Saccharomyces cerevisiae CHY 1011) using 10% (w/v) pretreated biomass under various pretreatment conditions. The maximum BTER was 73.00% under optimal pretreatment conditions (86.61 °C, 0.58 M, and 84.79 mL/min for temperature, sodium hydroxide concentration, and solution flow rate, respectively) and the experimental BTER was 70.01 ± 0.59%. SSF was performed to investigate the optimal enzyme and biomass dosage. As a result, maximum ethanol concentration and ethanol yield were 46.00 g/L and 77.36% at a loading pretreated biomass of 20% with 30 FPU/g cellulose of the enzyme dosage for barley straw to bioethanol. These results are a significant contribution to the production of bioethanol from barley straw.     

Economics of technical efficiency in white honey production: Using stochastic frontier production function

This study estimates and compares technical efficiency levels among beekeeping projects with various production capacities using a sample drawn from Fayoum Governorate in Egypt. The study also compares among beekeeping’s most important economic indicators, estimates the optimum production amount, and investigates why technical efficiency in Egyptian beekeeping is declining. We conclude that the average cost to produce one ton of honey decreases when production capacity increases. Furthermore, the average total, net return, and return on investment of one Egyptian pound increases when production capacity increases. Moreover, the results of a stochastic frontier production function indicate an increased return to scale. Finally, the results of maximum likelihood estimation show that technical inefficiency helps explain the deviation of actual from optimal production amounts.     

Dexamethasone stimulates thyrotropin-releasing hormone production in a C cell line

The hypothalamic peptide hormone TRH is also found in other tissues, including the thyroid. While TRH may be regulated by T3 in the hypothalamus, other regulators of TRH have not been identified and the regulation of TRH in nonhypothalamic tissues is unknown. We recently demonstrated the biosynthesis of TRH in the CA77 neoplastic thyroidal C cell line. We studied the regulation of TRH by dexamethasone in this cell line because glucocorticoids have been postulated to inhibit TSH secretion by decreasing TRH in the hypothalamus. Furthermore, TRH in the thyroid inhibits thyroid hormone release. Thus by regulating thyroidal TRH, glucocorticoids could also directly affect thyroid hormone secretion. Treatment of CA77 cells for 4 days with dexamethasone produced dose-dependent increases in both TRH mRNA and cellular and secreted TRH. Increases in TRH mRNA and peptide levels could be seen with 10(-9) M dexamethasone. A 4.8-fold increase in TRH mRNA and a 4-fold increase in secreted peptide were seen with 10(-7) M dexamethasone. Dexamethasone treatment did not increase beta-actin mRNA levels or cell growth. These results suggest that glucocorticoids may be physiological regulators of TRH in normal C cells. In addition to their inhibitory effects on TSH, glucocorticoids may decrease thyroid hormone levels by increasing thyroidal TRH. Since the glucocorticoid effects on C cell TRH are the converse of what is expected for hypothalamic TRH, glucocorticoid effects in these two tissues may be mediated by different regulators.     

Activation of collagenase production in a rat macrophage-like cell line

Collagenase activity from a rat macrophage line, AK-5, has been studied. The enzyme is present in these cells and its release is stimulated by Con A, LPS and a lymphokine preparation from rat spleen cells. Collagenase activity from these cells is inhibited by EDTA, cysteine or dithiothreitol, thereby suggesting it to be similar to other mammalian collagenases.     

Immunoglobulin production by a human-mouse somatic cell hybrid

The fusion of human lymphocytes and TEPC-15 mouse myeloma cells, which had not been adapted to culture, resulted in the establishment of in vitro hybrid cell cultures. Ten clones of this somatic cell hybrid were examined. There was preferential exclusion of human chromosomes: between two and five human chromosomes were identified in the hybrid clones by Giemsa banding. All of the clones had the mouse parental histocompatibility antigens, but only four clones also retained the human parental histocompatibility antigens. Secretion of parental immunoglobulin was determined by SDS-gel electrophoresis of species-specific immune precipitates. Synthesis of parental immunoglobulin by individual hybrid cells was determined by double label fluorescent antibody staining. Individual cells from six of the clones secreted and synthesized both human and mouse parental immunoglobulins. Three clones secreted only one parental immunoglobulin. Cells from one of these clones secreted and synthesized only human immunoglobulin. Cells from the remaining two clones secreted only one parental species of immunoglobulin but synthesized both human and mouse immunoglobulins. Finally, one clone did not secrete immunoglobulin, yet the individual cells synthesized both human and mouse parental species of immunoglobulin.     

Ecdysteroid production by a continuous insect cell line

The spent medium from ten established cell lines was extracted and tested for ecdysteroids by radioimmunoassay. Of the seven lepidopteran lines tested, only IAL-TNDI and MRRL-CH showed evidence of ecdysteroid production. However, the results were erratic and difficult to evaluate and these lines were dropped from further consideration. However, of the three cockroach cell lines tested, one, UMBGE 4, produces ecdysteroid and consistently releases virtually all of it into the medium. The main ecdysteroid was identified as ecdysone and the increase was logarithmic during the first 11 days of the subculture, with a decrease from day 11 to day 14. UMBGE 4 is a vesicle cell line which also tested positive for chitin synthesis. When the pH of the medium was lowered from pH 7.4 to pH 6.3, both the chitin synthesis and the ecdysone synthesis dropped by roughly 50%.     

Overall efficiency of in vitro embryo production and vitrification in cattle

In 5 replicates a total of 719 immature oocytes recovered from 94 slaughterhouse-derived bovine ovaries were matured and fertilized in vitro, then cultured for 7 to 9 d on a granulosa cell monolayer in TCM 199 supplemented with calf serum. Of 338 blastocysts (47% of oocytes cultured), 301 were vitrified in Hepes/bicarbonate buffered TCM-199 medium, 20% calf serum and dimethylsulfoxide and ethylene glycol as the cryoprotectants. After thawing in 1 M sucrose and subsequent culture in vitro, 237 (79%) of the blastocysts re-expanded and 177 (59%) hatched. Re-expansion and hatching rates differed between the blastocysts vitrified on Day 7 and Day 8 (84 and 69% vs 70 and 41%, respectively). We conclude that the applied methods are relatively simple and inexpensive to use, with an overall efficiency of the in vitro production/vitrification procedure being 1.9 hatched blastocyst/ovary. Therefore, this system seems suitable for large-scale production of cryopreserved bovine embryos for various purposes.     

L-Sorbose production in a continuous fermenter with and without cell recycle

The kinetics of continuous l-sorbose fermentation using Acetobacter suboxydans with and without cell recycle (100%) were investigated at dilution rates (D) of 0.05, 0.10, 0.15 and 0.3 h^–1. The biomass and sorbose concentrations for continuous fermentation without recycle increased as the dilution rate was increased from 0.05 to 0.10 h^–1. A maximum biomass concentration of 8.44 g l^–1 and sorbose concentration of 176.90 g l^–1 were obtained at D=0.10 h^–1. The specific rate of sorbose production and volumetric sorbose productivity at this dilution rate were 2.09 g g^–1 h^–1 and 17.69 g l^–1 h^–1. However, on further increasing the dilution rate to 0.3 h^–1, both biomass and sorbose concentrations decreased to 2.93 and 73.20 g l^–1 respectively, mainly due to washout of the reactor contents. However, the specific rate of sorbose formation and volumetric sorbose productivity at this dilution rate increased to 7.49 g g^–1 h^–1 and 21.96 g l^–1 h^–1 respectively. Continuous fermentation with 100% cell recycle served to further enhance the concentration of biomass and sorbose to 28.27 and 184.32 g l^–1 respectively (in the reactor at a dilution rate of 0.05 h^–1). Even though, there was a decline in the biomass and sorbose concentrations to 6.8 and 83.40 g l^–1 at a dilution rate of 0.3 h^–1, the specific rates of sorbose formation and volumetric sorbose productivity increased to 3.67 g g^–1h^–1 and 25.02 g l^–1 h^–1.     

Fluctuating water table affects gross ecosystem production and gross radiation use efficiency in a sedge-grass marsh

The eddy covariance method was used for continuous measurement of the seasonal courses of the following parameters of the carbon cycle in a sedge-grass marsh type of wetland ecosystem (49°01′29″N, 14°46′13″E, South Bohemia, Czech Republic, Central Europe): gross ecosystem production (GEP), net ecosystem production (NEP) and ecosystem respiration. During a 3-year series of measurements, we recorded marked fluctuations of the water table, which affected the overall water regime of the wetland studied. Between-year differences in the water regime strongly influenced the total annual carbon sequestration. The lowest annual GEP and NEP of 996 and 152 g m⁻² of carbon, respectively, were recorded in 2006, a year with two large floods, one in the spring, the other in the summer. By contrast, in the dry year of 2007, with no flood, the highest annual GEP and NEP were recorded: 1,328 and 274 g m⁻², respectively. Significant differences were found in the efficiency of solar energy use for GEP [gross radiation use efficiency, GRUE = GEP/PhAR (photosynthetically active radiation), i.e., amount of carbon gained per energy unit]. The highest GRUE was recorded immediately after the 2006 summer flood. In 2007, the GRUE decreased linearly with rising water table. A variable water regime thus markedly affects the processes of carbon accumulation and the efficiency of solar energy use for organic matter production in freshwater wetlands of the sedge-grass marsh type.