Comparison of Doxycycline Delivery Methods for Tet-Inducible Gene Expression in a Subcutaneous Xenograft Model

Doxycycline (Dox) controlled Tet systems provide a powerful and commonly used method for functional studies on the consequences of gene overexpression/downregulation. However, whereas Dox delivery in tissue culture in vitro is relatively simple, the situation in vivo is more complex. Several methods of Dox delivery in vivo have been described-e.g., in drinking water containing alcohol, in drinking water containing various concentrations of sucrose, and in feed. Unfortunately there are no reports directly comparing the advantages and disadvantages of these diverse methods, and there is no generally accepted standard. We therefore compared four non-invasive methods of Dox delivery in vivo-in drinking water, by gavage, as a jelly, and in standard feed. To assess the delivery of Dox by these methods, we used a subcutaneous xenograft model based on colorectal carcinoma cells engineered for Dox-inducible expression of an activated mutant of c-Src and the luciferase reporter gene. Our results indicate that feed represents the most favorable method of Dox administration.     

水产病原菌抗菌药物敏感试验标准化探讨

病原菌药物敏感试验是临床微生物学研究的重要手段,相比人类和兽医临床,国内外水产养殖中细菌性病原的药敏试验还未有公认的标准化参考方法。概括近几年国外对水产病原菌药敏试验标准化方法建立的研究进展,探讨了试验中关键因子的影响,指出我国水产药敏试验中存在的不足及对未来发展提出展望。     

Choice of method of place cell classification determines the population of cells identified

Place cells, spatially responsive hippocampal cells, provide the neural substrate supporting navigation and spatial memory. Historically most studies of these neurons have used electrophysiological recordings from implanted electrodes but optical methods, measuring intracellular calcium, are becoming increasingly common. Several methods have been proposed as a means to identify place cells based on their calcium activity but there is no common standard and it is unclear how reliable different approaches are. Here we tested four methods that have previously been applied to two-photon hippocampal imaging or electrophysiological data, using both model datasets and real imaging data. These methods use different parameters to identify place cells, including the peak activity in the place field, compared to other locations (the Peak method); the stability of cells’ activity over repeated traversals of an environment (Stability method); a combination of these parameters with the size of the place field (Combination method); and the spatial information held by the cells (Information method). The methods performed differently from each other on both model and real data. In real datasets, vastly different numbers of place cells were identified using the four methods, with little overlap between the populations identified as place cells. Therefore, choice of place cell detection method dramatically affects the number and properties of identified cells. Ultimately, we recommend the Peak method be used in future studies to identify place cell populations, as this method is robust to moderate variations in place field within a session, and makes no inherent assumptions about the spatial information in place fields, unless there is an explicit theoretical reason for detecting cells with more narrowly defined properties.     

检测纳米材料毒性的若干实验方法

纳米材料进入生命体和环境以后可能带来的生物安全问题需要定量的测定。现有的检测纳米材料生物安全性的方法大致可分为体内和体外实验两种,但是还没有证据表明既有的某个方法单独能作为一种检测所有纳米材料毒性的通用方法,也没有证据表明这些方法合在一起就能全面评估纳米材料的生物安全性。本文在总结既有的若干方法的同时,报道了一种基于rpsL基因的复制保真性来定量检测纳米材料毒性的方法。纳米材料对rpsL基因的体内体外复制过程保真性的影响均可方便地定量测定。     

Resampling methods for variance estimation of singular value decomposition analyses from microarray experiments

Microarray experiments offer the ability to generate gene expression measurements for thousands of genes simultaneously. Work has begun recently on attempting to reconstruct genetic networks based on analyses of microarray experiments in time-course studies. An important tool in these analyses has been the singular value decomposition method. However, little work has been done on assessing the variability associated with singular value decomposition analyses. In this report, we discuss use of the bootstrap as a method of obtaining standard errors for singular value decomposition analyses. We consider use of this method both when there are replicates and when no replicates exist. The proposed methods are illustrated with an application to two datasets: one involving a human foreskin study, the other involving yeast. Electronic Publication     

A matter of bacterial life and death

Over 50 years ago, standard microbiological methods were established for determining whether bacterial cells were dead or alive. Recently there has been a flurry of reports suggesting that bacteria may exist in an eclipsed state, escaping detection by standard methods. Whether there really is such a state is of more than academic interest, considering the implications for public health. The ensuing debate has been unusually energetic for the normally cultured community of microbiologists.     

Some protein interaction data do not exhibit power law statistics

It has been claimed that protein-protein interaction (PPI) networks are scale-free, and that identifying high-degree "hub" proteins reveals important features of PPI networks. In this paper, we evaluate the claims that PPI node degree sequences follow a power law, a necessary condition for networks to be scale-free. We provide two PPI network examples which clearly do not have power laws when analyzed correctly, and thus at least these PPI networks are not scale-free. We also show that these PPI networks do appear to have power laws according to methods that have become standard in the existing literature. We explain the source of this error using numerically generated data from analytic formulas, where there are no sampling or noise ambiguities.     

林木非同化器官CO2 通量的测定方法及对结果的影响

林木非同化器官的气体交换特性是研究森林 CO2 通量过程中的一个必须考虑的因子 ,但是目前对于如何测定并没有标准方法。综述研究根系呼吸的 6类 10种方法、研究树干和树枝呼吸的 2类方法 ,并对各种方法的优缺点和最新研究成果进行了讨论。在此基础之上 ,收集前人研究结果 (6 5个根系呼吸占土壤总呼吸比例的数据 ,5 9个美国黄松 (Pinus ponderosa)树干呼吸数据 )对不同研究方法对测定结果的可能影响进行了评述。根系呼吸对土壤呼吸的贡献率在 5 %～ 90 %之间 ,而近 6 0 %的研究结果显示土壤呼吸中根呼吸所占比例为 4 0 %～ 70 %。不同研究方法测定的根系呼吸结果不同 ,其中 ,使用同位素标记法测定的根系呼吸占土壤总呼吸的比例最低 (4 0 %) ,而根系排除法和树干环剥法测定结果最高 ,较同位素标记法 (人工同位素标记法和天然同位素丰度法 )的测定结果高 33%,较根系分离法的测定结果高 7%,表明根系排除法和树干环剥法对根际环境的扰动破坏可能导致估计偏高 ,而根系分离法中根系死亡导致呼吸速率降低和根系受伤导致呼吸增加之间的补偿作用 ,仅仅使测定结果稍微偏高。对树干呼吸的统计结果显示 ,当以树干表面积为基准时 ,离体测定结果较活体测定结果平均高 74 %(p=0 .0 13) ;而当用边材体积为基准时 ,离体测     

Meta-analysis of the difference of medians

We consider the problem of meta-analyzing two-group studies that report the median of the outcome. Often, these studies are excluded from meta-analysis because there are no well-established statistical methods to pool the difference of medians. To include these studies in meta-analysis, several authors have recently proposed methods to estimate the sample mean and standard deviation from the median, sample size, and several commonly reported measures of spread. Researchers frequently apply these methods to estimate the difference of means and its variance for each primary study and pool the difference of means using inverse variance weighting. In this work, we develop several methods to directly meta-analyze the difference of medians. We conduct a simulation study evaluating the performance of the proposed median-based methods and the competing transformation-based methods. The simulation results show that the median-based methods outperform the transformation-based methods when meta-analyzing studies that report the median of the outcome, especially when the outcome is skewed. Moreover, we illustrate the various methods on a real-life data set.     

Components of resistance of leaf rust (Melamspora laricii epitea Kleb./Melampsora ribesii-viminalis Kleb.) in Salix viminalis L.

Selection on partial resistance components, namely latent period, development rate of uredia and number and size of uredia, has been suggested as a means to achieve durable resistance. Three experiments were carried out in growth chambers to assess the impact of environmental and genetic factors on these components in the Salix-Melampsora host-pathogen system. They confirmed the environmental ability of the Melampsora-Salicaceae relationship and provided no definite answer on the possibility of attaining durable resistance through selection on partial resistance components. On the one hand, there is a large amount of heritable variation among clones for most components; on the other hand, all components were extremely sensitive to environmental conditions and isolates, making the development of standard selection methods difficult.     

Computational methods for prediction of T-cell epitopes--a framework for modelling, testing, and applications

Computational models complement laboratory experimentation for efficient identification of MHC-binding peptides and T-cell epitopes. Methods for prediction of MHC-binding peptides include binding motifs, quantitative matrices, artificial neural networks, hidden Markov models, and molecular modelling. Models derived by these methods have been successfully used for prediction of T-cell epitopes in cancer, autoimmunity, infectious disease, and allergy. For maximum benefit, the use of computer models must be treated as experiments analogous to standard laboratory procedures and performed according to strict standards. This requires careful selection of data for model building, and adequate testing and validation. A range of web-based databases and MHC-binding prediction programs are available. Although some available prediction programs for particular MHC alleles have reasonable accuracy, there is no guarantee that all models produce good quality predictions. In this article, we present and discuss a framework for modelling, testing, and applications of computational methods used in predictions of T-cell epitopes.     

Percutaneous ethanol injections in the treatment of nodular thyroid disease--fourteen years of experience

Surgery, radioactive iodine, and suppressive doses of Lthyroxine are commonly used in the treatment of benign nodular thyroid disease, with the best results achieved with surgery. However, recent advances in cytological diagnostic methods enable patients to choose alternative therapies if there are some contraindications or there is no agreement for surgery. Over past 14 years percutaneous ethanol injections (PEI), used in the past as a therapy for liver, kidney, parathyroid and adrenal cortical tumors, have been developed as an alternative method in the management of thyroid nodules. This paper reviews the investigations of a number aspects of PEI in the treatment of thyroid nodules reported in the literature. The evidence demonstrate that PEI is effective in the case of both solid non-toxic and autonomously functioning toxic nodules as well as cystic nodules. In majority of cases restore normal serum fT4, fT3 and TSH concentrations and shrink the tumor volume. The method seems to be safe and generally well tolerated by the patients. However, the current research base on the efficacy and PEI-associated risks is still inadequate to determine definitively its role as a standard treatment of benign nodular thyroid disease. For example, there is no detailed data comparing the results of treatment with PEI to standard treatment with radioactive iodine and L-thyroxine. Despite it, we conclude that PEI is a valuable method that earned recognition among other methods of the thyroid nodules treatment.     

水稻耐冷性鉴定评价方法

水稻耐冷性是我国东北和西南高海拔地区、日本、韩国和朝鲜等国水稻育种的重要研究目标,至今许多学者对水稻耐冷性进行了广泛的研究,并且目前所采用的耐冷性鉴定评价方法已基本成熟,但国内还没有对其进行技术规范化.本文汇总了国内外至今所采用的水稻耐冷性鉴定方法和耐冷指标及分级标准等,将为水稻耐冷性种质资源鉴定和育种以及我国水稻耐冷性鉴定评价技术规范的制定提供参考依据.     

Multiple imputation methods for treatment noncompliance and nonresponse in randomized clinical trials

Summary .  Randomized clinical trials are a powerful tool for investigating causal treatment effects, but in human trials there are oftentimes problems of noncompliance which standard analyses, such as the intention-to-treat or as-treated analysis, either ignore or incorporate in such a way that the resulting estimand is no longer a causal effect. One alternative to these analyses is the complier average causal effect (CACE) which estimates the average causal treatment effect among a subpopulation that would comply under any treatment assigned. We focus on the setting of a randomized clinical trial with crossover treatment noncompliance (e.g., control subjects could receive the intervention and intervention subjects could receive the control) and outcome nonresponse. In this article, we develop estimators for the CACE using multiple imputation methods, which have been successfully applied to a wide variety of missing data problems, but have not yet been applied to the potential outcomes setting of causal inference. Using simulated data we investigate the finite sample properties of these estimators as well as of competing procedures in a simple setting. Finally we illustrate our methods using a real randomized encouragement design study on the effectiveness of the influenza vaccine.     

A novel method for the extraction and culture of progenitor stem cells from human peripheral blood for use in regenerative medicine

Human peripheral blood (HPB) contains both circulating endothelial cells (CECs) and endothelial progenitor stem cells (EPCs), which may be suitable for use in regenerative medicine. There has been considerable interest in using these cells, but there is no "gold standard" technique for isolating these cells. The aim of this study was to characterize and compare a number of different extraction and culture techniques to develop a system to isolate and culture cells. EPC and CEC were isolated from HPB using either Histopaque-1077 or Lymphoprep. The two isolation methods were compared for the number of cells isolated, cell metabolism, and RNA expression. Both isolations produced viable cells and were comparable. The tissue culture method employed does have a significant effect on the cell population with regard to medium choice, fetal bovine serum concentration, and surface modification of the culture surface. In conclusion, it can be seen that although this study and previous work can suggest a basis for culture, further work to develop an optimized and agreed "gold standard" culture regime for EPC from HPB is required to maximize the potential of this source of cells for regenerative medicine and to translate its clinical use in the future.     

Concatenation and concordance in the reconstruction of mouse lemur phylogeny: an empirical demonstration of the effect of allele sampling in phylogenetics

The systematics and speciation literature is rich with discussion relating to the potential for gene tree/species tree discordance. Numerous mechanisms have been proposed to generate discordance, including differential selection, long-branch attraction, gene duplication, genetic introgression, and/or incomplete lineage sorting. For speciose clades in which divergence has occurred recently and rapidly, recovering the true species tree can be particularly problematic due to incomplete lineage sorting. Unfortunately, the availability of multilocus or "phylogenomic" data sets does not simply solve the problem, particularly when the data are analyzed with standard concatenation techniques. In our study, we conduct a phylogenetic study for a nearly complete species sample of the dwarf and mouse lemur clade, Cheirogaleidae. Mouse lemurs (genus, Microcebus) have been intensively studied over the past decade for reasons relating to their high level of cryptic species diversity, and although there has been emerging consensus regarding the evolutionary diversity contained within the genus, there is no agreement as to the inter-specific relationships within the group. We attempt to resolve cheirogaleid phylogeny, focusing especially on the mouse lemurs, by employing a large multilocus data set. We compare the results of Bayesian concordance methods with those of standard gene concatenation, finding that though concatenation yields the strongest results as measured by statistical support, these results are found to be highly misleading. By employing an approach where individual alleles are treated as operational taxonomic units, we show that phylogenetic results are substantially influenced by the selection of alleles in the concatenation process.     

Statistics of local multiple alignments

SUMMARY: BLAST statistics have been shown to be extremely useful for searching for significant similarity hits, for amino acid and nucleotide sequences. Although these statistics are well understood for pairwise comparisons, there has been little success developing statistical scores for multiple alignments. In particular, there is no score for multiple alignment that is well founded and treated as a standard. We extend the BLAST theory to multiple alignments. Following some simple assumptions, we present and justify a significance score for multiple segments of a local multiple alignment. We demonstrate its usefulness in distinguishing high and moderate quality multiple alignments from low quality ones, with supporting experiments on orthologous vertebrate promoter sequences.     

Multiplex bead array assays: performance evaluation and comparison of sensitivity to ELISA

The measurement of soluble cytokines and other analytes in serum and plasma is becoming increasingly important in the study and management of many diseases. As a result, there is a growing demand for rapid, precise, and cost-effective measurement of such analytes in both clinical and research laboratories. Multiplex bead array assays provide quantitative measurement of large numbers of analytes using an automated 96-well plate format. Enzyme-linked immunosorbent assay (ELISAs) have long been the standard for quantitative analysis of cytokines and other biomarkers, but are not well suited for high throughput multiplex analyses. However, prior to replacement of ELISA assays with multiplex bead array assays, there is a need to know how comparable these two methods are for quantitative analyses. A number of published studies have compared these two methods and it is apparent that certain elements of these assays, such as the clones of monoclonal antibodies used for detection and reporting, are pivotal in obtaining similar results from both assays. By careful consideration of these variables, it should be possible to utilize multiplex bead array assays in lieu of ELISAs for studies requiring high throughput analysis of numerous analytes.     

Comparison of methods of enumerating coliforms after UV disinfection.

In view of the differences that have been found between the most-probable-number and membrane filtration methods for the recovery of coliforms from chlorinated samples, the survival of total and fecal coliforms in UV-irradiated effluent samples, as tested by the most-probable-number and standard single-step membrane filtration methods, was compared. There were no significant differences in the survival of total and fecal coliforms, as tested by the two methods. In a separate set of experiments comparing total and fecal coliform survival, as tested by the most-probable-number method, only a very small but statistically significant difference of 0.1 log survival units was found. For UV-disinfected wastewater effluents, standard one-step membrane filtration procedures are comparable to standard most-probable-number procedures.     

Indices of oxidative stress. 2. Lipid peroxides

Two methods of the determination of lipid peroxidation products have been compared which are based on Fe(II) oxidation by them at acid pH values in the presence of xylenol orange which binds Fe(III) have been compared. The first method uses cumene hydropeoxide as an internal standard. In the second one, lipid peroxides are previously reduced by triphenylphosphine and these substances content is measured as a difference of the production of complexes with xylenol orange and iron ions in the control (with reduction) and experimental sample (without reduction). The optimization of measurement conditions is described. The levels of lipid peroxides in goldfish tissues assayed simultaneously by two methods were similar. The method with cumene hydroperoxide needs less amounts of biological material; moreover, there is no necessity in a calibration curve. Effects of hyperoxia on lipid peroxide levels in goldfish tissues were studied with the cumene method. Within the first hours of hyperoxia this index increased 13-times in the liver and 2-times in the brain and muscle. The further exposure rebounded this parameter to the initial level. Levels of lipid peroxides positively correlated with levels of end products of lipid peroxidation (thiobarbiturate acid reactive substances) in the goldfish tissues. The method of quantification of lipid peroxides with cumene is recommended for wide using in biological investigations.