A Simple Construction Procedure for Eesolvable Incomplete Block Designs for any Number of Treatments

A simple, straightforward procedure, which requires no special tables or generators, is presented for constructing resolvable incomplete block designs for v=pk, v=p²k, …, treatments, for k≥p, in incomplete blocks of size k. Also, it is shown, how to obtain incomplete block designs for any v in blocks of size k and k+1. The procedure allows construction of balanced incomplete block designs for p = k a prime number. For p = n not a prime number, incomplete block designs can be obtained by the procedure, but are not balanced. However, for p_s being the smallest prime factor of n, p_s + 1 for v = n², p_s²+ p_s + 1 for v = n³, …, arrangements can be obtained for which the occurrence of any treatment pair in the blocks is either zero or one. This is called a zero-one concurrence design. Procedures are described for obtaining additional zero-one concurrence arrangements. It is shown that the efficiency of these designs is maximum. Both intra-block and inter-block analyses are described.     

Designs Suited for Varietal Trials with Multiple Basals

DAS (1960) gave a method of construction of confounded balanced asymmetrical factorial designs of the type v × 2² by using BIB designs. In the present paper a method has been given for construction of balanced asymmetrical factorial designs of the type (v–t) × 2² by using truncated balanced incomplete block designs obtainable by omitting t treatments. Likewise, partially balanced asymmetrical factorial designs can also be obtained by omitting any particular treatment alongwith its first or second associate treatments from the v treatments of a PBIB design. We can get a large number of new designs not available in literature through this technique. These designs are well suited for varietal trials with multiple basals.     

A statistical approach to improve compound screening in cell culture media

The Chinese hamster ovary (CHO) cell line is widely used for the production of recombinant proteins due to its high growing capacity and productivity, as well as other cell lines derived later than CHO. Adapting cell culture media for each specific cell line is a key to exploit these features for cost effective and fast product generation. Media supplementation is generally addressed by means of one‐factor‐at‐a‐time or classical design of experiments approaches but these techniques may not be efficient enough in preliminary screening phases. In this study, a novel strategy consisting in folding over the Plackett–Burman design was used to increase cell growth and trastuzumab production of different CHO cell lines through supplementation with nonanimal recombinant compounds. Synergies between compounds could be detected with a reduced number of experiments by using this methodology in comparison to more conventional fractional factorial designs. In the particular case reported here, the sequential use of this modified Plackett–Burman in combination with a Box‐Behnken design led to a 1.5‐fold increase in cell growth (10 × 10⁶ cells/mL) and a two‐fold in trastuzumab titer (122 mg/L) in suspension batch culture.     

Stirred tank bioreactor culture combined with serum‐/xenogeneic‐free culture medium enables an efficient expansion of umbilical cord‐derived mesenchymal stem/stromal cells

Mesenchymal stem/stromal cells (MSC) are being widely explored as promising candidates for cell‐based therapies. Among the different human MSC origins exploited, umbilical cord represents an attractive and readily available source of MSC that involves a non‐invasive collection procedure. In order to achieve relevant cell numbers of human MSC for clinical applications, it is crucial to develop scalable culture systems that allow bioprocess control and monitoring, combined with the use of serum/xenogeneic (xeno)‐free culture media. In the present study, we firstly established a spinner flask culture system combining gelatin‐based Cultispher^®S microcarriers and xeno‐free culture medium for the expansion of umbilical cord matrix (UCM)‐derived MSC. This system enabled the production of 2.4 (±1.1) x10⁵ cells/mL (n = 4) after 5 days of culture, corresponding to a 5.3 (±1.6)‐fold increase in cell number. The established protocol was then implemented in a stirred‐tank bioreactor (800 mL working volume) (n = 3) yielding 115 million cells after 4 days. Upon expansion under stirred conditions, cells retained their differentiation ability and immunomodulatory potential. The development of a scalable microcarrier‐based stirred culture system, using xeno‐free culture medium that suits the intrinsic features of UCM‐derived MSC represents an important step towards a GMP compliant large‐scale production platform for these promising cell therapy candidates.     

Statistical Analyses for Multistage Experiment Designs

Suppose that t experiments are conducted simultaneously on the same set of experimental units. For example, suppose that t mutually orthogonal latin square experiment designs are used for the t experiments on n² experimental units. Statistical literature is voluminous on construction of such designs, but contains relatively little and incomplete results on statistical analyses for such designs. Six statistical analyses are presented for a pair of orthogonal latin square experiment designs. Then, the methods are generalized for t mutually orthogonal experiment designs. The results are also extended to a set of t mutually balanced Youden experiment designs.     

Medium optimization for spore production of Coniothyrium minitans using statistically‐based experimental designs

Statistically‐based experimental designs were used to optimize a chemically defined solid medium for the spore production of Coniothyrium minitans. In the first optimization step the influence of starch, urea, phosphate, magnesium, calcium, thiamin and trace elements on spore production was evaluated using a fractional factorial design. Starch and trace elements influenced spore production positively while urea affected spore production negatively. The other components had no significant influence on spore production. In the second and third steps the concentrations of starch, urea and trace elements were further optimized using central composite designs and response surface analysis. This optimization strategy allowed the spore production to be increased by a factor 7 from 4 × 10⁹ to almost 3 × 10¹⁰ spores per Petri dish of 9 cm diameter. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 64: 92–100, 1999.     

On the Origin of the Ideality Factor in Perovskite Solar Cells

The measurement of the ideality factor (n_id) is a popular tool to infer the dominant recombination type in perovskite solar cells (PSC). However, the true meaning of its values is often misinterpreted in complex multilayered devices such as PSC. In this work, the effects of bulk and interface recombination on the n_id are investigated experimentally and theoretically. By coupling intensity‐dependent quasi‐Fermi level splitting measurements with drift diffusion simulations of complete devices and partial cell stacks, it is shown that interfacial recombination leads to a lower n_id compared to Shockley–Read–Hall (SRH) recombination in the bulk. As such, the strongest recombination channel determines the n_id of the complete cell. An analytical approach is used to rationalize that n_id values between 1 and 2 can originate exclusively from a single recombination process. By expanding the study over a wide range of the interfacial energy offsets and interfacial recombination velocities, it is shown that an ideality factor of nearly 1 is usually indicative of strong first‐order non‐radiative interface recombination and that it correlates with a lower device performance. It is only when interface recombination is largely suppressed and bulk SRH recombination dominates that a small n_id is again desirable.     

Transcultural Properties of the Composite Scale of Morningness: The Relevance of the “Morning Affect” Factor

Morningness scales have been translated into several languages, but a lack of normative data and methodological differences make cross‐cultural comparisons difficult. This study examines the psychometric properties and factor structure of the Composite Scale of Morningness (CSM) in samples from five countries: France (n=627), Italy (n=702), Spain (n=391), Thailand (n=503), and Australia (n=654). Strong national differences are identified. A quadratic relationship between age and CSM total score was apparent in the Australian data with a downward trend after age 35 yrs. There was no age effect in any sample in the range from 18 to 29 yrs. Factor analysis identified a three‐factor solution in all groups for both men and women. Tucker's congruence coefficients indicate that: (1) this solution is highly congruent between sexes in each culture, and (2) a morning affect factor is highly congruent between cultures. These results indicate there are national differences in factorial structure and that cut‐off scores used to categorize participants as morning‐ and evening‐types should be established for different cultural and age groups.     

Predicting leaf wax n‐alkane 2H/1H ratios: controlled water source and humidity experiments with hydroponically grown trees confirm predictions of Craig–Gordon model

The extent to which both water source and atmospheric humidity affect δ²H values of terrestrial plant leaf waxes will affect the interpretations of δ²H variation of leaf waxes as a proxy for hydrological conditions. To elucidate the effects of these parameters, we conducted a long‐term experiment in which we grew two tree species, Populus fremontii and Betula occidentalis, hydroponically under combinations of six isotopically distinct waters and two different atmospheric humidities. We observed that leaf n‐alkane δ²H values of both species were linearly related to source water δ²H values, but with slope differences associated with differing humidities. When a modified version of the Craig–Gordon model incorporating plant factors was used to predict the δ²H values of leaf water, all modelled leaf water values fit the same linear relationship with n‐alkane δ²H values. These observations suggested a relatively constant biosynthetic fractionation factor between leaf water and n‐alkanes. However, our calculations indicated a small difference in the biosynthetic fractionation factor between the two species, consistent with small differences calculated for species in other studies. At present, it remains unclear if these apparent interspecies differences in biosynthetic fractionation reflect species‐specific biochemistry or a common biosynthetic fractionation factor with insufficient model parameterization.     

Design,Synthesis, Lipophilic Properties,and Binding Affinities of Potential Ligands in Positron Emission Tomography (PET) for Visualization of Brain Dopamine D4 Receptors

We report the synthesis of compounds structurally related to the high‐affinity dopamine D₄ receptor ligand N‐{2‐[4‐(3‐cyanopyridin‐2‐yl)piperazin‐1‐yl]ethyl}‐3‐methoxybenzamide ( 1e ). All compounds were specifically designed as potential PET radioligands for brain D₄ receptor visualization, having lipophilicity within a range for brain uptake and weak non‐specific binding (0.75<cLogP<3.15) and bearing a substituent for easy access to labeling with the positron emitter isotope ¹¹C or ¹⁸F. The best compound of the series, N‐{2‐[4‐(4‐chlorophenyl)piperazin‐1‐yl]ethyl}‐6‐fluoropyridine‐3‐carboxamide ( 7a ), displayed excellent selectivity over D₂ and D₃ receptors (>100‐fold), but its D₄ receptor affinity was suboptimal for imaging of brain D₄ receptors (K_i=30 nM ).     

Nitrile‐Substituted QA Derivatives: New Acceptor Materials for Solution‐Processable Organic Bulk Heterojunction Solar Cells

The development of non‐fullerene‐based electron acceptors (especially organic molecules with sufficient absorption property within the solar spectrum region) for bulk‐heterojunction (BHJ) organic solar cells (OSCs) is an important issue for the achievement of high photoconversion efficiency. In this contribution, a new class of organic acceptors di‐cyan substituted quinacridone derivatives (DCN‐nCQA, n = 4, 6 and 8) for BHJ solar cells was designed and synthesized. DCN‐nCQA molecules possess facile synthesis, solution processability, visible and near‐IR light absorption and relatively stable characteristics. The DCN‐8CQA molecule exhibited a proper LUMO energy level (–4.1 eV), small bandgap (1.8 eV) and moderate electron mobility (10^?4 cm² V^?1 S^?1), suggesting that this molecule is an ideal acceptor material for the classical donor material regio‐regular poly (3‐hexylthiophene) (P3HT). A photovoltaic device with a structure of [ITO/PEDOT:PSS/P3HT:DCN‐8CQA/LiF/Al] displayed a power conversion efficiency of 1.57% and a fill factor of 57% under 100 mW cm^?2 AM 1.5G simulated solar illumination. The DCN‐nCQA molecules showed remarkable absorption in the region from 650 to 700 nm, where P3HT has a weak absorption promoting overlap with the solar spectrum and potentially improving the performance of the solar cell.     

Four Factor Balanced Optimal Designs of Resolution V in Terms of a Modified Trace Criterion

Balanced optimal two level fractional factorial designs of resolution V have been reported in series of papers by SRIVASTAVA and CHOPRA (1971a, 1971b, 1974) and CHOPRA and SRIVASTAVA (1973a, 1973b). The optimality criterion considered by them is the trace optimality which implies minimisation of the average variance of the estimated effects which include the general mean μ, main effects, A_i, and first order interactions, A_ij. In the present work some new optimal designs have been obtained on the basis of a modified criterion of trace optimality. The criterion requires minimisation of the average variance of each of the main effects, two factor interactions, etc. separately instead of minimising average variance of all the estimated effects.     

Statistical optimization of culture media for production of phycobiliprotein by Synechocystis sp. PCC 6701

Synechocystis sp. PCC 6701 has a brilliantly colored pigment, phycobiliprotein containing phycoerythrin. Culture medium was optimized by sequential designs in order to maximize phycobiliprotein production. The observed fresh weights after 6 days were 0.58 g/L in BG-11, 0.83 g/L in medium for Scenedesmus sp. and 0.03∼0.52 g/L in the other tested media. Medium for Scenedesmus sp. was selected to be optimized by fractional factorial design and central composite design since the medium maintained a more stable pH within a desirable range due to higher contents of phosphate. The fractional factorial design had seven factors with two levels: KNO₃, NaNO₃, NaH₂PO₄, Na₂HPO₄, Ca(NO₃)₂, FeEDTA, and MgSO₄. From the result of fractional factorial design, nitrate and phosphate were identified as significant factors. A central composite design was then applied with four variables at five levels each: nitrate, phosphate, pH, and light intensity. Parameters such as fresh weight and phycobiliprotein contents were used to determine the optimum value of the four variables. The proposed optimum media contains 0.88 g/L of nitrate, 0.32 g/L of phosphate under 25 μE·m⁻²·s⁻¹ of light intensity. The maximum phycobiliprotein contents have been increased over 400%, from 4.9 to 25.9 mg/L after optimization.     

Factorial design and response surface optimization of crude violacein for Chromobacterium violaceum production

Initially an eleven variable, sixteen assay 2^15–11 fractional factorial design, was used to determine the key factors in the production of violacein produced by Chromobacterium violaceum, CCT 3496. Subsequently five and three factor central composite designs were executed to determine response surfaces with the aim of optimizing cellular mass and crude violacein production. The 7.5 g l^–1 dry cell mass and 0.17 g l^–1 crude violacein productions obtained with our initial culture medium were increased to 21 g l^–1 and 0.43 g l^–1, respectively, for a medium investigated in the three factor design.     

A Simple Procedure for Constructing Experiment Designs with Incomplete Blocks of Sizes 2 and 3

In order to maximize control of heterogeneity within complete blocks, an experimenter could use incomplete blocks of size k = 2 or 3. In certain situations, incomplete blocks of this nature would eliminate the need for such spatial types of analyses as nearest neighbor. The intrablock efficiency factors for such designs are relatively low. However, with recovery of interblock information, FEDERER and SPEED (1987) have presented measures of design efficiency factors which demonstrate that efficiency factors approach unity for certain ratios of the intrablock and interblock variance components. Hence with recovery of interblock information, even incomplete block designs with k = 2 or 3 have relatively high efficiency factors. The reduction in the intrablock error variance over the complete block error variance in many situations will provide designs with high efficiency. A simple procedure for constructing incomplete blocks of sizes 2 and 3 is presented. It is shown how to obtain additional zero-one association confounding arrangements when v = 4 t, t an integer, and for v = pk, k ≤ p. It is indicated how to do the statistical analysis for these designs.     

Lipoprotein Oxidation and Plasma Vitamin E in Nondiabetic Normotensive Obese Patients

Objective: To correlate the susceptibility of low‐(LDL) and very‐low‐density lipoprotein to oxidation in vitro and the concentrations of serum antibodies against malondialdehyde‐modified LDL and plasma vitamin E with the anthropometric and laboratory characteristics of obesity. Research Methods and Procedures: A total of 75 nondiabetic, normotensive obese patients were assigned to one of four groups according to their body mass index (BMI): moderately obese (30 ≤ BMI ≤ 34.9 kg/m², n = 11), severely obese (35 ≤ BMI ≤ 39.9 kg/m², n = 20), morbidly obese (40 ≤ BMI ≤ 50 kg/m², n = 29), and very severely obese (BMI > 50 kg/m², n = 15). Results: The oxidation lag time for LDL from patients with a BMI ≥35 kg/m² was shorter than that for LDL from non‐obese controls (n = 13), whereas very‐low‐density lipoprotein oxidation lag times were not significantly different. The serum antibodies against modified LDL were similar in all groups, whereas the plasma vitamin E concentrations of obese patients were decreased (p ≤ 0.01). There was a negative correlation between LDL oxidation lag time and BMI (r = ?0.35, p = 0.0008), and between plasma vitamin E and BMI (r = ?0.53, p < 0.0001) and waist‐to‐hip ratio (r = ?0.40, p = 0.0003). Discussion: The LDL of nondiabetic, normotensive obese patients is more readily oxidized, and plasma vitamin E concentrations are low. These are both risk factors for coronary heart disease.     

Low level laser irradiation precondition to create friendly milieu of infarcted myocardium and enhance early survival of transplanted bone marrow cells

We suggested that low‐level laser irradiation (LLLI) precondition prior to cell transplantation might remodel the hostile milieu of infarcted myocardium and subsequently enhance early survival and therapeutic potential of implanted bone marrow mesenchymal stem cells (BMSCs). Therefore, in this study we wanted to address: (1) whether LLLI pre‐treatment change the local cardiac micro‐environment after myocardial infarction (MI) and (2) whether the LLLI preconditions enhance early cell survival and thus improve therapeutic angiogenesis and heart function. MI was induced by left anterior descending artery ligation in female rats. A 635 nm, 5 mW diode laser was performed with energy density of 0.96 J/cm² for 150 sec. for the purpose of myocardial precondition. Three weeks later, qualified rats were randomly received with LLLI precondition (n= 26) or without LLLI precondition (n= 27) for LLLI precondition study. Rats that received thoracotomy without coronary ligation were served as sham group (n= 24). In the cell survival study, rats were randomly divided into 4 groups: serum‐free culture media injection (n= 8), LLLI precondition and culture media injection (n= 8), 2 million male BMSCs transplantation without LLLI pre‐treatment (n= 26) and 2 million male BMSCs transplantation with LLLI precondition (n= 25) group, respectively. Vascular endothelial growth factor (VEGF), glucose‐regulated protein 78 (GRP78), superoxide dismutase (SOD) and malondialdehyde (MDA) in the infarcted myocardium were evaluated by Western blotting, real‐time PCR and colorimetry, respectively, at 1 hr, 1 day and 1 week after laser irradiation. Cell survival was assayed with quantitative real‐time PCR to identify Y chromosome gene and apoptosis was assayed with transferase‐mediated dUTP end labelling staining. Capillary density, myogenic differentiation and left ventricular function were tested by immunohistochemistry and echocardiography, respectively, at 1 week. After LLLI precondition, increased VEGF and GRP78 expression, as well as the enhanced SOD activity and inhibited MDA production, was observed. Compared with BMSC transplantation and culture media injection group, although there was no difference in the improved heart function and myogenic differentiation, LLLI precondition significantly enhanced early cell survival rate by 2‐fold, decreased the apoptotic percentage of implanted BMSCs in infarcted myocardium and thus increased the number of newly formed capillaries. Taken together, LLLI precondition could be a novel non‐invasive approach for intraoperative cell transplantation to enhance cell early survival and therapeutic potential.     

Production and characterization of a low‐molecular‐weight bacteriocin from Bacillus licheniformis MKU3

Aims: Enhancing production and characterization of a low‐molecular‐weight bacteriocin from Bacillus licheniformis MKU3. Methods and Results: The culture supernatant of B. licheniformis MKU3 exhibited bacteriocin‐like activity against Gram‐positive and ‐negative bacteria and different fungi and yeast. SDS–PAGE analysis of the extracellular proteins of B. licheniformis MKU3 revealed a bacteriocin‐like protein with a molecular mass of 1·5 kDa. This bacteriocin activity was found to be stable under a pH range of 3·0–10·0 and at temperatures up to 100°C for 60 min, but inactivated by proteinase K, trypsin or pronase E. An experimental fractional factorial design for optimization of production medium resulted in a maximum activity of bacteriocin (11 000 AU ml^?1) by B. licheniformis MKU3. Conclusions: A low‐molecular‐weight bacteriocin‐like protein from B. licheniformis MKU3 exhibited a wide spectrum of antimicrobial activity against several Gram‐positive bacteria, several fungi and yeast. A 3·6‐fold increase in the production of bacteriocin was achieved using the culture medium optimized through a fractional factorial design. Significance and Impact of the Study: A bacteriocin with wide spectrum of activity against Gram‐positive bacterial pathogens, filamentous fungi and yeast suggested its potential clinical use. Statistical method facilitated optimization of cultural medium for the improved production of bacteriocin.     

Crystal structure of phenylacetic acid degradation protein PaaG from Thermus thermophilus HB8

Microbial degradation of phenylacetic acid proceeds via the hybrid pathway that includes formation of a coenzyme A thioester, ring hydroxylation, non‐oxygenolytic ring opening, and β‐oxidation‐like reactions. A phenylacetic acid degradation protein PaaG is a member of the crotonase superfamily, and is a candidate non‐oxygenolytic ring‐opening enzyme. The crystal structure of PaaG from Thermus thermophilus HB8 was determined at a resolution of 1.85 Å. PaaG consists of three identical subunits related by local three‐fold symmetry. The monomer is comprised of a spiral and a helical domain with a fold characteristic of the crotonase superfamily. A putative active site residue, Asp136, is situated in an active site cavity and surrounded by several hydrophobic and hydrophilic residues. The active site cavity is sufficiently large to accommodate a ring substrate. Two conformations are observed for helix H2 located adjacent to the active site. Helix H2 is kinked at Asn81 in two subunits, whereas it is kinked at Leu77 in the other subunit, and the side chain of Tyr80 is closer to Asp136. This indicates that catalytic reaction of PaaG may proceed with large conformational changes at the active site. Asp136 is the only conserved polar residue in the active site. It is located at the same position as those of 4‐chlorobenzoyl‐CoA dehalogenase and peroxisomal Δ³,Δ²‐enoyl‐CoA isomerase, indicating that PaaG may undergo isomerization or a ring‐opening reaction via a Δ³,Δ²‐enoyl‐CoA isomerase‐like mechanism. Proteins 2009. © 2009 Wiley‐Liss, Inc.