A novel method for simultaneous high resolution identification of HLA-A, HLA-B, and HLA-Cw alleles.

We describe a novel high resolution DNA based typing approach for HLA class I alleles, which identifies the recombinational motifs present in exons 2 and 3 of the HLA class I genes. Unique identification patterns for 201 known HLA-A, HLA-B, and HLA-Cw alleles were generated by the use of only 40 probes, which were targeted at these common motifs. The unambiguous identification of the alleles was achieved by the development of a new and powerful allelic separation technique that allows isolation of single alleles after amplification. To validate the method, we have used locus-specific primers to amplify exons 2 and 3 of HLA-A, HLA-B, and HLA-Cw loci from 22 heterozygous and 41 homozygous cell lines. After amplification, the allelic fragments from each locus were separated, blotted, and hybridized with the 40 probes. In all cases, the allelic products could be separated and 81 different class I alleles, 33 HLA-A, 30 HLA-B, and 18 HLA-Cw, were identified according to the predicted probe hybridization patterns.

Simultaneous fluorescence-activated cell sorter analysis of two distinct transcriptional elements within a single cell using engineered green fluorescent proteins.

Green fluorescent protein (GFP) is widely used as a reporter gene in both prokaryotes and eukaryotes. However, the fluorescence levels of wild-type GFP (wtGFP) are not bright enough for fluorescence-activated cell sorting or flow cytometry. Several GFP variants were generated that are brighter or have altered excitation spectra when expressed in prokaryotic cells. We engineered two GFP genes with different combinations of these mutations, GFP(S65T,V163A) termed GFP-Bex1, and GFP(S202F,T203I,V163A) termed GFP-Vex1. Both show enhanced brightness and improved signal-to-noise ratios when expressed in mammalian cells and appropriately excited, compared with wtGFP. Each mutant retains only one of the two excitation peaks of the wild-type protein. GFP-Bex1 excites at 488 nm (blue) and GFP-Vex1 excites at 406 nm (violet), both of which are available laser lines. Excitation at these wavelengths allows for the independent analyses of these mutants by fluorescence-activated cell sorting, permitting simultaneous, quantitative detection of expression from two different genes within single mammalian cells.

Simultaneous assessment of loss of heterozygosity at multiple microsatellite loci using semi-automated fluorescence-based detection: subregional mapping of chromosome 4 in cervical carcinoma.

Detection of loss of heterozygosity (LOH) by comparison of normal and tumor genotypes using PCR-based microsatellite loci provides considerable advantages over traditional Southern blotting-based approaches. However, current methodologies are limited by several factors, including the numbers of loci that can be evaluated for LOH in a single experiment, the discrimination of true alleles versus "stutter bands," and the use of radionucleotides in detecting PCR products. Here we describe methods for high throughput simultaneous assessment of LOH at multiple loci in human tumors; these methods rely on the detection of amplified microsatellite loci by fluorescence-based DNA sequencing technology. Data generated by this approach are processed by several computer software programs that enable the automated linear quantitation and calculation of allelic ratios, allowing rapid ascertainment of LOH. As a test of this approach, genotypes at a series of loci on chromosome 4 were determined for 58 carcinomas of the uterine cervix. The results underscore the efficacy, sensitivity, and remarkable reproducibility of this approach to LOH detection and provide subchromosomal localization of two regions of chromosome 4 commonly altered in cervical tumors.

Single cell Ca2+/cAMP cross-talk monitored by simultaneous Ca2+/cAMP fluorescence ratio imaging.

The spatial and temporal dynamics of two intracellular second messengers, cAMP and Ca2+, were simultaneously monitored in living cells by digital fluorescence ratio imaging using FlCRhR, a single-excitation dual-emission cAMP indicator, and fura-2, a dual-excitation single-emission Ca2+ probe. In single C6-2B glioma cells, isoproterenol- or forskolin-evoked cAMP accumulation (measured in vivo as an increased FlCRhR emission ratio) was reduced when cytosolic free Ca2+ concentration was elevated before, simultaneously with, or after cAMP activation. However, in REF-52 fibroblasts, Ca2+ neither prevented nor reduced forskolin-stimulated cAMP production. These results provide novel in vivo evidence for the Ca2+ modulation of the cAMP transduction pathway in C6-2B cells. The simultaneous microscopic measurement of cAMP and Ca2+ kinetics in single cells makes it now possible to study the regulatory interactions between these second messengers at the cellular and even the subcellular level.

Human cooperation in the simultaneous and the alternating Prisoner's Dilemma: Pavlov versus Generous Tit-for-Tat.

The iterated Prisoner's Dilemma has become the paradigm for the evolution of cooperation among egoists. Since Axelrod's classic computer tournaments and Nowak and Sigmund's extensive simulations of evolution, we know that natural selection can favor cooperative strategies in the Prisoner's Dilemma. According to recent developments of theory the last champion strategy of "win--stay, lose--shift" ("Pavlov") is the winner only if the players act simultaneously. In the more natural situation of players alternating the roles of donor and recipient a strategy of "Generous Tit-for-Tat" wins computer simulations of short-term memory strategies. We show here by experiments with humans that cooperation dominated in both the simultaneous and the alternating Prisoner's Dilemma. Subjects were consistent in their strategies: 30% adopted a Generous Tit-for-Tat-like strategy, whereas 70% used a Pavlovian strategy in both the alternating and the simultaneous game. As predicted for unconditional strategies, Pavlovian players appeared to be more successful in the simultaneous game whereas Generous Tit-for-Tat-like players achieved higher payoffs in the alternating game. However, the Pavlovian players were smarter than predicted: they suffered less from defectors and exploited cooperators more readily. Humans appear to cooperate either with a Generous Tit-for-Tat-like strategy or with a strategy that appreciates Pavlov's advantages but minimizes its handicaps.

Simultaneous optical and X-ray observations of flares and rotational modulation on the RS CVn binary HR 1099 (V711 Tau) from the MUSICOS 1998 campaign

We present simultaneous and continuous observations of the Hα, Hβ, He I D_3, Na I D_1,D_2 doublet and the Ca II H&K lines for the RS CVn system HR 1099. The spectroscopic observations were obtained during the MUSICOS 1998 campaign involving several observatories and instruments, both echelle and long-slit spectrographs. During this campaign, HR 1099 was observed almost continuously for more than 8 orbits of 2^d.8. Two large optical flares were observed, both showing an increase in the emission of Hα, Ca II H K, Hβ and He I D_3 and a strong filling-in of the Na I D_1, D_2 doublet. Contemporary photometric observations were carried out with the robotic telescopes APT-80 of Catania and Phoenix-25 of Fairborn Observatories. Maps of the distribution of the spotted regions on the photosphere of the binary components were derived using the Maximum Entropy and Tikhonov photometric regularization criteria. Rotational modulation was observed in Hα and He I D_3 in anti-correlation with the photometric light curves. Both flares occurred at the same binary phase (0.85), suggesting that these events took place in the same active region. Simultaneous X-ray observations, performed by ASM on board RXTE, show several flare-like events, some of which correlate well with the observed optical flares. Rotational modulation in the X-ray light curve has been detected with minimum flux when the less active G5 V star was in front. A possible periodicity in the X-ray flare-like events was also found.

Nonclassical light revealed by the joint statistics of simultaneous measurements

Nonclassicality cannot be a single-observable property, since the statistics of any quantum observable is compatible with classical physics. We develop a general procedure to reveal nonclassical behavior of light states from the joint statistics arising in the practical measurement of multiple observables. Beside embracing previous approaches, this protocol can disclose nonclassical features for standard examples of classical-like behavior, such as SU(2) and Glauber coherent states. When combined with other criteria, this would imply that every light state is nonclassical.

What We Don't Know Can Hurt Us: Clinical Inference Bias and Constructed Memories of Sexual Abuse.

The false memory/recovered memory debate, research regarding the malleability of memory, and the current lack of methods for validating recovered memories all support the view that heightened care is required of therapists dealing with clients whom they suspect have been sexually abused. The judgmental heuristics that underlie the major clinical inference biases of confirmatory bias, biased covariation, base rate fallacies, and schematic processing errors are all relevant to the processes leading to therapist-client constructions of memories of sexual abuse. Suggestions for minimizing each of these biases are offered. Personal motivations of the client and client suggestibility are factors that may contribute to the construction of memories of sexual abuse, and suggestions for minimizing the impact of these motivations are offered. In conclusion, general suggestions for minimizing the impact of clinical inference biases within the sexual abuse treatment context are summarized.

Simultaneous implementation of the Bachelor, Masters and PhD degrees in nursing in the Universidad Jaume I. Castellón de la Plana, Spain

We present an experience in Nursing Education, accredited and implemented under the Spanish University System Reform in a Public University (Jaume I, Castellón) which had no previous nursing studies. The academics offered included all three educational levels (Bachelor, Master's and Doctorate), with an integrated theoretical-practical-clinical teaching methodology for the Bachelor Degree, competence acquisition in research in the Master's degree, and a doctorate formed by lines of research in the field of Nursing. Studies are accredited by the National Agency for Quality Assessment, which were authorized by the Spanish Ministry of Education and implanted between 2009 and 2011.

Simultaneous synthesis of heat exchanger networks with pressure recovery: optimal integration between heat and work

The optimal integration between heat and work may significantly reduce the energy demand and consequently the process cost. This paper introduces a new mathematical model for the simultaneous synthesis of heat exchanger networks (HENs) in which the pressure levels of the process streams can be adjusted to enhance the heat integration. A superstructure is proposed for the HEN design with pressure recovery, developed via generalized disjunctive programming (GDP) and mixed-integer nonlinear programming (MINLP) formulation. The process conditions (stream temperature and pressure) must be optimized. Furthermore, the approach allows for coupling of the turbines and compressors and selection of the turbines and valves to minimize the total annualized cost, which consists of the operational and capital expenses. The model is tested for its applicability in three case studies, including a cryogenic application. The results indicate that the energy integration reduces the quantity of utilities required, thus decreasing the overall cost.

Impact of simultaneous pancreas-kidney transplantation: patients' perspectives

Background: Few qualitative studies of simultaneous pancreas-kidney transplantation (SPK Tx) have been published. The aims of this study were to explore from the perspective of patients, the experience of living with diabetes mellitus type 1 (T1DM), suffering from complications, and undergoing SPK Tx with good outcome; and to determine the impact of SPK Tx on patients and their social and cultural environment. Methods: We performed a focused ethnographic study. Twenty patients were interviewed. Data were analyzed using content analysis and constant comparison following the method proposed by Miles and Huberman. Results: A functioning SPK Tx allowed renal replacement therapy and insulin to be discontinued. To describe their new situation, patients used words and phrases such as "miracle", "being reborn" or "coming back to life". Although the complications of T1DM, its surgery and treatment, and associated psychological problems did not disappear after SPK Tx, these were minimized when compared with the pretransplantation situation. Conclusion: For patients, SPK Tx represents a recovery of their health and autonomy despite remaining problems associated with the complications of T1DM and SPK Tx. The understanding of patients' existential framework and their experience of disease are key factors for planning new intervention and improvement strategies.

Simultaneous synthesis of work exchange networks with heat integration

The optimal integration of work and its interaction with heat can represent large energy savings in industrial plants. This paper introduces a new optimization model for the simultaneous synthesis of work exchange networks (WENs), with heat integration for the optimal pressure recovery of process gaseous streams. The proposed approach for the WEN synthesis is analogous to the well-known problem of synthesis of heat exchanger networks (HENs). Thus, there is work exchange between high-pressure (HP) and low-pressure (LP) streams, achieved by pressure manipulation equipment running on common axes. The model allows the use of several units of single-shaft-turbine-compressor (SSTC), as well as stand-alone compressors, turbines and valves. Helper motors and generators are used to respond to any demand and excess of energy. Moreover, between the WEN stages the streams are sent to the HEN to promote thermal recovery, aiming to enhance the work integration. A multi-stage superstructure is proposed to represent the process. The WEN superstructure is optimized in a mixed-integer nonlinear programming (MINLP) formulation and solved with the GAMS software, with the goal of minimizing the total annualized cost. Three examples are conducted to verify the accuracy of the proposed method. In all case studies, the heat integration between WEN stages is essential to improve the pressure recovery, and to reduce the total costs involved in the process.

Simultaneous environmental and economic process synthesis of Isobutane Alkylation

This multidisciplinary study concerns the optimal design of processes with a view to both maximizing profit and minimizing environmental impacts. This can be achieved by a combination of traditional chemical process design methods, measurements of environmental impacts and advanced mathematical optimization techniques. More to the point, this paper presents a hybrid simulation-multiobjective optimization approach that at once optimizes the production cost and minimizes the associated environmental impacts of isobutane alkylation. This approach has also made it possible to obtain the flowsheet configurations and process variables that are needed to manufacture isooctane in a way that satisfies the above-stated double aim. The problem is formulated as a Generalized Disjunctive Programming problem and solved using state-of-the-art logic-based algorithms. It is shown, starting from existing alternatives for the process, that it is possible to systematically generate a superstructure that includes alternatives not previously considered. The optimal solution, in the form a Pareto curve, includes different structural alternatives from which the most suitable design can be selected. To evaluate the environmental impact, Life Cycle Assessment based on two different indicators is employed: Ecoindicator 99 and Global Warming Potential.

The Goodness of Simultaneous Fits in ISIS

In a previous work, we introduced a tool for analyzing multiple datasets simultaneously, which has been implemented into ISIS. This tool was used to fit many spectra of X-ray binaries. However, the large number of degrees of freedom and individual datasets raise an issue about a good measure for a simultaneous fit quality. We present three ways to check the goodness of these fits: we investigate the goodness of each fit in all datasets, we define a combined goodness exploiting the logical structure of a simultaneous fit, and we stack the fit residuals of all datasets to detect weak features. These tools are applied to all RXTE-spectra from GRO 1008−57, revealing calibration features that are not detected significantly in any single spectrum. Stacking the residuals from the best-fit model for the Vela X-1 and XTE J1859+083 data evidences fluorescent emission lines that would have gone undetected otherwise.

Inference of binding affinity from neuronal receptors in humans

Tese de mestrado, Bioinformática e Biologia Computacional (Bioinformática), Universidade de Lisboa, Faculdade de Ciências, 2016