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A new four-color flow cytometric assay to detect apoptosis in lymphocyte subsets of cultured peripheral blood cells 总被引:1,自引:0,他引:1
Hasper HJ Weghorst RM Richel DJ Meerwaldt JH Olthuis FM Schenkeveld CE 《Cytometry》2000,40(2):167-171
BACKGROUND: Human peripheral blood lymphocytes kept in culture after isolation die by an apoptotic process. Detection of apoptosis with labeled Annexin V to demonstrate loss of plasma membrane asymmetry is sensitive, specific, and easy using flow cytometry. This is true in lymphoblastic cell lines when combining Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide (PI). However, measurement of apoptosis by flow cytometry in isolated human lymphocytes using Annexin V-FITC/PI is disturbed by the presence of a variable percentage of erythrocytes in the isolated lymphocyte population. To overcome this problem, we have developed and tested a new four-color flow cytometric assay to detect apoptosis in lymphocyte subsets of cultured peripheral blood cells. METHODS: Peripheral blood lymphocytes are isolated by density gradient centrifugation. Nucleus-containing cells are selected using CD45-phycoerythrin (PE). The lymphocyte subset of interest is selected using CD4, CD8, or CD19 energy-coupled dye (ECD) labeling. Apoptosis is detected using Annexin V-FITC with 7-amino-Actinomycin-D (7-AAD) to distinguish early apoptotic from late apoptotic lymphocytes. RESULTS: We have developed a new technique to detect apoptosis in isolated human peripheral blood lymphocyte subsets with good reproducibility, coefficient of variation < 17%. CONCLUSIONS: We now have a validated tool to study apoptosis in subsets of isolated human lymphocytes to increase our knowledge of pathogenesis and therapies in lymphoreticular malignancies. 相似文献
23.
R Podstawski DJ Choszcz S Konopka J Klimczak M Starczewski 《Biology of sport / Institute of Sport》2014,31(4):315-321
The aim of the study was to evaluate anthropometric characteristics as determinants of 500 m rowing ergometer performance in physically inactive collegiate females. In this cross-sectional study, which included 196 collegiate females aged 19-23 years not participating in regular physical activities, body mass (BM), body height (BH), length of upper limbs (LA), length of lower limbs (LL), body mass index (BMI), slenderness index (SI), and the Choszcz-Podstawski index (CPI) were measured and a stepwise multiple regression analysis was performed. Participants performed 500 m maximal effort on a Concept II rowing ergometer. BM, BH, LA, LL, and the BMI, SI and CPI indices were found to be statistically significant determinants of 500 m performance. The best results (T) were achieved by females whose BH ranged from 170 to 180 cm, with LA and LL ranging from 75 to 80 cm and 85 to 90 cm, respectively. The best fitting statistical model was identified as: T = 11.6793 LR – 0.1130 LR2 – 0.0589 LN2 + 29.2157 CPI2 + 0.1370 LR·LN - 2.6926 LR·CPI – 211.7796. This study supports a need for additional studies focusing on understanding the importance of anthropometric differences in rowing ergometer performance, which could lead to establishing a better quality reference for evaluation of cardiorespiratory fitness tested using a rowing ergometer in collegiate females. 相似文献
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Rosan Luijcks Hermie J. Hermens Lonneke Bodar Catherine J. Vossen Jim van. Os Richel Lousberg 《PloS one》2014,9(4)
Experience of stress may lead to increased electromyography (EMG) activity in specific muscles compared to a non-stressful situation. The main aim of this study was to develop and validate a stress-EMG paradigm in which a single uncontrollable and unpredictable nociceptive stimulus was presented. EMG activity of the trapezius muscles was the response of interest. In addition to linear time effects, non-linear EMG time courses were also examined. Taking into account the hierarchical structure of the dataset, a multilevel random regression model was applied. The stress paradigm, executed in N = 70 subjects, consisted of a 3-minute baseline measurement, a 3-minute pre-stimulus stress period and a 2-minute post-stimulus phase. Subjects were unaware of the precise moment of stimulus delivery and its intensity level. EMG activity during the entire experiment was conform a priori expectations: the pre-stimulus phase showed a significantly higher mean EMG activity level compared to the other two phases, and an immediate EMG response to the stimulus was demonstrated. In addition, the analyses revealed significant non-linear EMG time courses in all three phases. Linear and quadratic EMG time courses were significantly modified by subjective anticipatory stress level, measured just before the start of the stress task. Linking subjective anticipatory stress to EMG stress reactivity revealed that subjects with a high anticipatory stress level responded with more EMG activity during the pre-stimulus stress phase, whereas subjects with a low stress level showed an inverse effect. Results suggest that the stress paradigm presented here is a valid test to quantify individual differences in stress susceptibility. Further studies with this paradigm are required to demonstrate its potential use in mechanistic clinical studies. 相似文献
26.
Evolution at the tip and base of the X chromosome in an African population of Drosophila melanogaster 总被引:1,自引:0,他引:1
Hitchhiking effects of advantageous mutations have been invoked to explain
reduced polymorphism in regions of low crossing-over in Drosophila. Besides
reducing DNA heterozygosity, hitchhiking effects should produce strong
linkage disequilibrium and a frequency spectrum skewed toward an excess of
rare polymorphisms (compared to the neutral expectation). We measured DNA
polymorphism in a Zimbabwe population of D. melanogaster at three loci,
yellow, achaete, and suppressor of forked, located in regions of reduced
crossing-over. Similar to previously published surveys of these genomic
regions in other populations, we observed low levels of nucleotide
variability. However, the frequency spectrum was compatible with a neutral
model, and there was abundant evidence for recombination in the history of
the yellow and ac genes. Thus, some aspects of the data cannot be accounted
for by a simple hitchhiking model. An alternative hypothesis, background
selection, might be compatible with the observed patterns of linkage
disequilibrium and the frequency spectrum. However, this model cannot
account for the observed reduction in nucleotide heterozygosity. Thus,
there is currently no satisfactory theoretical model for the data from the
tip and base of the X chromosome in D. melanogaster.
相似文献
27.
Oligogalacturonic acids (OGAs), derived from plant cell wall pectin, have
been implicated in a number of signal transduction pathways involved in
growth, development and defense responses of higher plants. This study
investigates the size range of OGAs capable of inducing ethylene synthesis
in tomato plants, and demonstrates that in contrast with many other
effects, only short chain OGAs are active. Oligomers across a range of DP
from 2-15 were separated and purified to homogeneity by QAE-Sephadex anion
exchange chromatography using a novel elution system. The OGAs were applied
to tomato plants and assayed for their ability to induce ethylene gas
release and changes in steady state levels of mRNA encoding the ethylene
forming enzyme aminocyclopropane-1-carboxylic acid oxidase (ACO). The study
demonstrated that only OGAs in the size range of DP4-6 were active both in
eliciting ACO expression and in the production of ethylene.
相似文献
28.
A change in twist of actin provides the force for the extension of the acrosomal process in limulus sperm: the false-discharge reaction 总被引:6,自引:5,他引:1 下载免费PDF全文
One of the most spectacular motions is the generation of the acrosomal process in the limulus sperm. On contact with the egg, the sperm generates a 60-mum-long process that literally drills its way through the jelly surrounding the egg. This irresversible reaction takes only a few seconds. We suggested earlier that this motion is driven by a change in twist of the actin filaments comprising the acrosomal process. In this paper we analyze the so-called false discharge, a reversible reaction, in which the acrosomal filament bundle extends laterally from the base of the sperm and not anteriorly from the apex. Unlike the true discharge, which is straight, the false discharge is helical. Before extension, the filament bundle is coiled about the base of the sperm. In the coil, the bundle is not smoothly bent but consists of arms (straight segments) and elbows (corners) so that the coil looks like a 14-sided polygon. The extension of the false discharge works as follows: starting at the base of the bundle, the filaments change their twist which concomitantly changes the orientations of the elbows relative to each other; that is, in the coil, the elbows all like in a common plane, but after the change in twist, the plane of each elbow is rotated to be perpendicular to that of its neighbors. This change transforms the bundle from a compact coil into an extended left- handed helix. Because the basal end of the bundle is unconstrained, the extension is lateral. The true discharge works the same way but starts at the apical end of the bundle. The apical end, however, is constrained by its passage through the nuclear canal, which directs the extention anteriorly. Unlike the false discharge, during the true discharge the elbows are melted out, making the reaction irreversible. This study shows that rapid movement can be regenerated by actin without myosin and gives us insight into the molecular mechanism. 相似文献
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