DNA content in fresh versus paraffin-embedded tissue. Flow cytometric analysis of 100 tumors.

DNA ploidy analysis was determined on 100 consecutive tumors from a wide variety of sites using both fresh and paraffin-embedded tissue on the same specimen. The correlation coefficient (r) value between the methods was 0.85. Aneuploidy was detected by both methods in 51/100 (51%) of the cases. Fresh tissue analysis yielded 10 additional cases (overall 61% aneuploidy) not detected on corresponding paraffin-embedded sections, whereas paraffin-embedded analysis detected 4 additional cases (overall 55% aneuploidy) not revealed by fresh tissue analysis. Fresh tissue analysis produced lower coefficients of variation and resulted in a cleaner preparation with less cellular debris. Fresh tissue analysis was also superior to paraffin for the detection of hypodiploid, near-diploid and multiple peaks. Analysis of paraffin-embedded material allows examination of archival tissue and provides a more rapid means of long-term follow-up and statistical correlations for prognostic studies. Although the overall correlation of both methodologies for DNA analysis showed a minimal variation in results, in our experience fresh tissue analysis has an advantage and is preferable, when available, for ploidy analysis.     

Effect of Zeitgeber Intensity Reduction on a Simulated Dual-Oscillator Human Circadian System: Classical and Dynamic Analysis

The two-oscillator model of human circadian rhythmicity was analyzed when a zeitgeber relative intensity of 1, 0.5, or 0.1 was introduced into the equations. Fourier analysis was compared with dynamic analysis such as attractor reconstruction or Liapunov exponent calculation. After a 50 or 90% reduction in zeitgeber intensity, the dynamics of the system became equivalent and differed significantly from those of a system with maximal zeitgeber intensity. When 10% aleatory noise was added to the data, the analysis was still applicable, and the results obtained were essentially the same as in the absence of noise. Dynamic analysis could thus provide a distinct classification for periodic data, based on the type of analysis.     

一种快速有效分析烟草花冠中花青素苷的方法

花青素苷（anthocyanins）成分和含量的分析是花色研究的重要内容之一。该文利用高效液相色谱-电喷雾离子化-质谱技术（HPLC-ESI-MS）, 建立了一种快速有效地分析烟草（Nicotiana tabacum）花冠中花青素苷成分及含量的方法。在保证良好分离效果的基础上, 将分析时间缩短至15分钟, 大大提高了分析速度, 降低了成本, 并成功应用于转基因烟草花冠中花青素苷成分的分析。为不具备高效分析仪器（如超高效液相色谱-串联质谱（UPLC-MS/MS））的一般实验室研究花青素苷合成相关基因在烟草中的异源表达提供了一种有效且实用的分析方法。     

Application of the exploratory data analysis for evaluating the toxicity of chlorinated phenol derivatives by various cell models

Exploratory data analysis based on multivariate statistical analysis techniques was introduced as a new approach to expressing the toxicity of chemical substances at the simultaneous acceptance of various cell models. Using principal component analysis and cluster analysis methods the toxicity of chlorinated phenol derivatives on employing some of the cell models (chlorococcal algae, cyanobacteria, bacteria, micromycetes, plant and animal cells) was characterized. The previous empirical experience that the toxicity of chlorinated phenol derivatives will increase with a growing degree of chlorination and that the presence of the methoxy group will cause a lowering of the toxic effect was demonstrated. The relationship between groups of tests used was presented.     

A weighted least squares method for inverse dynamic analysis

Internal forces in the human body can be estimated from measured movements and external forces using inverse dynamic analysis. Here we present a general method of analysis which makes optimal use of all available data, and allows the use of inverse dynamic analysis in cases where external force data is incomplete. The method was evaluated for the analysis of running on a partially instrumented treadmill. It was found that results correlate well with those of a conventional analysis where all external forces are known.     

In vivo gait analysis in a mouse femur fracture model

Although the mouse has become a preferred species for molecular studies on fracture healing, gait analysis after fracture fixation and during bone healing has not yet been performed in mice. Herein, we introduce a novel technique for gait analysis in mice and report the change of motion pattern after fracture and fixation. A standardized femur fracture was stabilized by a common pin. The non-fractured tibia was additionally marked with a pin, allowing continuous analysis of the tibio-femoral angle by digital video-radiography. Dynamic gait analysis was performed at day fourteen after surgery in a radio-opaque running wheel. Fracture fixation resulted in a significantly reduced range and maximum of the tibio-femoral angle compared to non-fractured controls. This was associated with a significantly reduced stride length. Because stride frequency was slightly increased and, thus, stride time diminished, stride velocity was not significantly reduced compared to controls. Thus, our study demonstrates distinct alterations of the gait of mice at 2 weeks after femur fracture and stabilization. Our results support the need of gait analysis in fracture healing studies to assess the animals’ well-being.     

Contextual analysis and intermediate cell markers enhance high-resolution cell image analysis for automated cervical smear diagnosis.

Until now, efforts to automate cervical smear diagnosis have focused on analyzing features of individual cells. In a complex specimen such as that obtained from a cervical scrape, diagnostically significant cells may not be adequately represented or may elude detection by the automated technology. An approach is needed that extracts additional quantitative information from cervical smears beyond what the cell-by-cell approach can provide. A new methodology, contextual analysis, was developed to extract global quantitative information about cells, cell clusters, and background debris. This pilot study was designed to compare the efficacy of contextual analysis with high-resolution, single cell analysis and the analysis of intermediate cell markers. Thirty-four samples prepared as monolayers and stained with the Feulgen-Thionin/Congo Red stain were measured. Contextual analysis alone was able to classify 91% of the smears correctly; single cell analysis classified 94% of the cells correctly; and the intermediate cell analysis correctly identified the smear diagnosis for 84% of the cells. When all three analysis methods were combined into a simple smear level classifier, the overall smear classification accuracy was improved over those obtained using the three methodologies alone.     

Chromosomal abnormalities and DNA image cytometry of haematological neoplasms in fine needle aspirates of lymph nodes

The current diagnostics of haematological neoplasms along with morphological analysis, immunophenotyping and molecular analysis inevitably includes cytogenetic analysis. In this work the possibility of cytomorphological subclassification of haematological neoplasms from lymph node fine needle aspirates was examined without depending upon the referential histological diagnosis and cytogenetic analysis. In addition, the feasibility of cytogenetic analysis of the material obtained by lymph node fine needle aspiration (FNA) was examined. By analysing the findings of cytogenetic analysis and DNA image cytometry, it was decided to examine the possibility of comparing the findings and supplementing diagnostic possibilities of these methods. In 15 cases cytological diagnoses and cytogenetic analysis of haematological neoplasms were performed on the material obtained by lymph node FNA. In 12 of 15 cases histological diagnosis was made separately. A good cytohistological correlation was available in 9 of 12 cases (75%). Cytomorphological diagnoses in 10 of 15 cases (76%) were confirmed by the finding of a specific chromosomal translocation. In two cases cytological diagnosis did not correlate with the histological diagnosis and was confirmed only with specific chromosomal translocations. The lymphocytes obtained by lymph node FNA were adequate material for cytogenetic analysis - in 15 of 18 (83%) cases mitoses in cell cultures were obtained. In 13 of 15 (87%) cases clonal chromosomal abnormalities were detected, whereas in 2 of 15 (13%) cases a normal karyotype was found. DNA image cytometry was performed on nine samples, whereas in six samples the material was not sufficient. Although a small number of samples was analysed in the cases with identical cytomorphological diagnoses, the analysed histograms regarding the DNA index values showed heterogeneity. In conclusion, a cell culture sampled by FNA of lymph nodes is an adequate method for the chromosomal analysis. The specific cytogenetic abnormality associated with cytological diagnosis provides an opportunity to make a definitive diagnosis and provides a powerful approach when reference diagnosis on biopsy material cannot be obtained.     

Genome analysis for nucleotide interactions in fully sequenced genomes of selective prokaryotes

Fully sequenced prokaryotic genomes ofEscherichia coli, Haemophilus influenzae andMethanococcus janaschii were subjected .to genome analysis for nucleotide interactions. The analysis was restricted to inter-nucleotide relations like two nucleotides in a dinucleotide, three nucleotides in a codon and two codons in a dicon. This relational analysis was carried out in C language and was compiled on a C++ compiler. The relational analysis showed a preferential dinucleotide frequency (the observed frequencies of AA and TT were higher than the expected frequency and the observed frequencies of CC and GG). From codon frequency distribution analysis, sub-codonic elements have been noticed, exerting that the first one or first two nucleotide may reasonably determine the next nucleotide(s) in a codon. The analysis further reveals the existence of short-range randomness or chaotic behaviour in prokaryotic genomes, which might be a forerunner for the origin of introns in eukaryotes, besides being involved in a regulatory role.     

Synthesis of a low-molecular-weight form of exopolysaccharide by Bradyrhizobium japonicum USDA 110

A novel extracellular low-molecular-weight polysaccharide was detected as a contaminant within extracellular cyclic beta-1,6-beta-1,3-glucan preparations from Bradyrhizobium japonicum USDA 110 cultures. Compositional analysis, methylation analysis, and nuclear magnetic resonance analysis revealed that this low-molecular-weight polysaccharide was composed of the same pentasaccharide repeating unit previously described for the high-molecular-weight form of the exopolysaccharide (EPS) synthesized by B. japonicum strains. Mass spectrometry analysis indicated that the size of this low-molecular-weight form of EPS was consistent with a dimeric form of the pentasaccharide repeating unit.     

Evaluation of capillary supercritical fluid chromatography with mass spectrometric detection for the analysis of a drug (mebeverine) in a dog plasma matrix

Supercritical fluid chromatography with mass spectrometric detection was evaluated as a technique for the analysis of drugs in biological fluids. Dog plasma was spiked with a model drug, mebeverine, and with a deuterium-labeled analog of mebeverine. The spiked plasma was prepared for analysis by solid-phase extraction on octadecylsilane cartridges. Mebeverine levels in the spiked dog plasma samples were determined by interpolation from a standard curve. Accuracy and precision of the analysis were determined within and between days. In general, accuracy was found to be 100 ± 15% for plasma samples spiked with 6 to 60 ng mebeverine/ml. The relative standard deviation for replicate sample analysis over this concentration range was between 5 and 12.5%.     

Terminal Restriction Fragment Length Polymorphism Data Analysis for Quantitative Comparison of Microbial Communities

Terminal restriction fragment length polymorphism (T-RFLP) is a culture-independent method of obtaining a genetic fingerprint of the composition of a microbial community. Comparisons of the utility of different methods of (i) including peaks, (ii) computing the difference (or distance) between profiles, and (iii) performing statistical analysis were made by using replicated profiles of eubacterial communities. These samples included soil collected from three regions of the United States, soil fractions derived from three agronomic field treatments, soil samples taken from within one meter of each other in an alfalfa field, and replicate laboratory bioreactors. Cluster analysis by Ward's method and by the unweighted-pair group method using arithmetic averages (UPGMA) were compared. Ward's method was more effective at differentiating major groups within sets of profiles; UPGMA had a slightly reduced error rate in clustering of replicate profiles and was more sensitive to outliers. Most replicate profiles were clustered together when relative peak height or Hellinger-transformed peak height was used, in contrast to raw peak height. Redundancy analysis was more effective than cluster analysis at detecting differences between similar samples. Redundancy analysis using Hellinger distance was more sensitive than that using Euclidean distance between relative peak height profiles. Analysis of Jaccard distance between profiles, which considers only the presence or absence of a terminal restriction fragment, was the most sensitive in redundancy analysis, and was equally sensitive in cluster analysis, if all profiles had cumulative peak heights greater than 10,000 fluorescence units. It is concluded that T-RFLP is a sensitive method of differentiating between microbial communities when the optimal statistical method is used for the situation at hand. It is recommended that hypothesis testing be performed by redundancy analysis of Hellinger-transformed data and that exploratory data analysis be performed by cluster analysis using Ward's method to find natural groups or by UPGMA to identify potential outliers. Analyses can also be based on Jaccard distance if all profiles have cumulative peak heights greater than 10,000 fluorescence units.     

Manual and semi-automated morphological analysis of Penicillium chrysogenum chemostat cultures

Measurement of key morphological indices of chemostat cultures of Penicillium chrysogenum, by image analysis, was carried out manually at Strathclyde University and semi-automatically at Birmingham University using identical preserved samples. Using both methods, the value of the mean hyphal growth unit was found to decrease with decreasing dilution rate. Although similar trends were observed for data obtained at Strathclyde and Birmingham, the values of key morphological indices measured by semi-automated analysis were consistently higher than the values obtained by manual analysis. This discrepancy was as a result of the different analysis methods, particularly with respect to clump analysis. An electronic image transfer method is discussed which would allow analysis of a given image set at either site by either method. © Rapid Science Ltd. 1998     

Fe3+ immobilized metal affinity chromatography with silica monolithic capillary column for phosphoproteome analysis

Immobilized metal affinity chromatography (IMAC) is a commonly used technique for phosphoproteome analysis due to its high affinity for adsorption of phosphopeptides. Miniaturization of IMAC column is essential for the analysis of a small amount of sample. Nanoscale IMAC column was prepared by chemical modification of silica monolith with iminodiacetic acid (IDA) followed by the immobilization of Fe3+ ion inside the capillary. It was demonstrated that Fe3+-IDA silica monolithic IMAC capillary column could specifically capture the phosphopeptides from tryptic digest of alpha-casein with analysis by MALDI-TOF MS. The silica monolithic IMAC capillary column was manually coupled with nanoflow RPLC/nanospray ESI mass spectrometer (muRPLC-nanoESI MS) for phosphoproteome analysis. The system was validated by analysis of standard phosphoproteins and then it was applied to the analysis of protein phosphorylation in mouse liver lysate. Besides MS/MS spectra, MS/MS/MS spectra were also collected for neutral loss peak. After database search and manual validation with conservative criteria, 29 singly phosphorylated peptides were identified by analyzing a tryptic digest of only 12 mug mouse liver lysate. The results demonstrated that the silica monolithic IMAC capillary column coupled with muRPLC-nanoESI MS was very suitable for the phosphoproteome analysis of minute sample.     

Terminal restriction fragment length polymorphism data analysis for quantitative comparison of microbial communities

Terminal restriction fragment length polymorphism (T-RFLP) is a culture-independent method of obtaining a genetic fingerprint of the composition of a microbial community. Comparisons of the utility of different methods of (i) including peaks, (ii) computing the difference (or distance) between profiles, and (iii) performing statistical analysis were made by using replicated profiles of eubacterial communities. These samples included soil collected from three regions of the United States, soil fractions derived from three agronomic field treatments, soil samples taken from within one meter of each other in an alfalfa field, and replicate laboratory bioreactors. Cluster analysis by Ward's method and by the unweighted-pair group method using arithmetic averages (UPGMA) were compared. Ward's method was more effective at differentiating major groups within sets of profiles; UPGMA had a slightly reduced error rate in clustering of replicate profiles and was more sensitive to outliers. Most replicate profiles were clustered together when relative peak height or Hellinger-transformed peak height was used, in contrast to raw peak height. Redundancy analysis was more effective than cluster analysis at detecting differences between similar samples. Redundancy analysis using Hellinger distance was more sensitive than that using Euclidean distance between relative peak height profiles. Analysis of Jaccard distance between profiles, which considers only the presence or absence of a terminal restriction fragment, was the most sensitive in redundancy analysis, and was equally sensitive in cluster analysis, if all profiles had cumulative peak heights greater than 10,000 fluorescence units. It is concluded that T-RFLP is a sensitive method of differentiating between microbial communities when the optimal statistical method is used for the situation at hand. It is recommended that hypothesis testing be performed by redundancy analysis of Hellinger-transformed data and that exploratory data analysis be performed by cluster analysis using Ward's method to find natural groups or by UPGMA to identify potential outliers. Analyses can also be based on Jaccard distance if all profiles have cumulative peak heights greater than 10,000 fluorescence units.     

Prenatal diagnosis of Duchenne muscular dystrophy: prospective linkage analysis and retrospective dystrophin cDNA analysis.

The accuracy of DNA-based prenatal diagnosis of Duchenne muscular dystrophy (DMD) was determined by study of 174 families. Only 60% of families had a living affected male, and 63% had history of a single affected male. Prenatal diagnosis was declined by 47% of mothers whose DNA studies predicted a carrier risk below 2%, and none have had affected sons. Fetal risk was estimated prospectively by linkage analysis using intragenic and flanking RFLPs and retrospectively using dystrophin cDNA analysis for families whose linkage estimates lacked precision. Diagnostic accuracy was determined by comparing predictions with 40 male pregnancy outcomes. On the basis of linkage analysis, we anticipated 3.2 DMD males and observed 3.0. Retrospective cDNA analysis identified deletions in 2 of these 3 males. The combined use of linkage and cDNA deletion analysis provided a highly accurate method for prenatal diagnosis of DMD.     

基于时间聚类分析和独立成分分析的癫痫fMRI盲分析方法

提出了一种基于时间聚类分析和独立成分分析的癫痫fMRI数据盲分析方法,并将两种方法有效联合,提取发作间期的癫痫fMRI激活时空信息.该方法首先由时间聚类分析得到与激活相关的时间峰度特征曲线,以此特征作为时间参考信息;再由空间独立成分分析分解fMRI信号得到空间独立成分;最后将每个独立成分所对应的时间曲线与参考曲线做相关分析提取相应脑激活图.提出的方法无需任何关于癫痫fMRI的先验假设信息,有效解决了独立成分的排序问题,实现了对数据的盲分析.仿真试验结果阐明了这一方法的有效性及可靠性,对癫痫数据的试验结果显示空间定位准确性优于统计参数图方法.     

A Bayesian hierarchical non-linear regression model in receiver operating characteristic analysis of clustered continuous diagnostic data

Receiver operating characteristic (ROC) analysis is a useful evaluative method of diagnostic accuracy. A Bayesian hierarchical nonlinear regression model for ROC analysis was developed. A validation analysis of diagnostic accuracy was conducted using prospective multi-center clinical trial prostate cancer biopsy data collected from three participating centers. The gold standard was based on radical prostatectomy to determine local and advanced disease. To evaluate the diagnostic performance of PSA level at fixed levels of Gleason score, a normality transformation was applied to the outcome data. A hierarchical regression analysis incorporating the effects of cluster (clinical center) and cancer risk (low, intermediate, and high) was performed, and the area under the ROC curve (AUC) was estimated.     

Evaluation of a protocol for metabolic profiling studies on human blood plasma by combined ultra-performance liquid chromatography/mass spectrometry: From extraction to data analysis

The investigation presented here describes a protocol designed to perform high-throughput metabolic profiling analysis on human blood plasma by ultra-performance liquid chromatography/mass spectrometry (UPLC/MS). To address whether a previous extraction protocol for gas chromatography (GC)/MS-based metabolic profiling of plasma could be used for UPLC/MS-based analysis, the original protocol was compared with similar methods for extraction of low-molecular-weight compounds from plasma via protein precipitation. Differences between extraction methods could be observed, but the previously published extraction method was considered the best. UPLC columns with three different stationary phases (C8, C18, and phenyl) were used in identical experimental runs consisting of a total of 60 injections of extracted male and female plasma samples. The C8 column was determined to be the best for metabolic profiling analysis on plasma. The acquired UPLC/MS data of extracted male and female plasma samples was subjected to principal component analysis (PCA) and orthogonal projections to latent structures discriminant analysis (OPLS-DA). Furthermore, a strategy for compound identification was applied here, demonstrating the strength of high-mass-accuracy time-of-flight (TOF)/MS analysis in metabolic profiling.     

紫球藻多糖化学结构解析

为明确紫球藻多糖的化学结构,本文采用化学分析和光谱分析方法对紫球藻多糖的一级糖链结构进行了分析。GC分析表明该多糖由木糖、葡萄糖和半乳糖组成,为一种杂多糖,其摩尔比为:2.96∶1.25∶3.06;红外光谱分析结果显示紫球藻多糖为硫酸化多糖,糖苷键类型为β构型;化学分析结果推断紫球藻多糖糖链连接方式以1→3为主,存在少量1→2,1→4,1→6键型,且半乳糖在支链或主链末端有较大量的存在,木糖和葡萄糖在主链或靠近主链区域有特定分布;NMR分析显示紫球藻多糖的硫酸酯基连在C-6上,且多糖的糖苷键为β型;GC-MS联机分析进一步确定紫球藻多糖为一种主要含有1→3糖苷键,并含有1→4,1→6糖苷键的杂多糖。综合上述分析,推断出紫球藻多糖的糖链主链的重复单元结构。