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961.
Hacker UT Schildhauer I Barroso MC Kofler DM Gerner FM Mysliwietz J Buening H Hallek M King SB 《Cancer immunology, immunotherapy : CII》2006,55(5):547-557
The modulated expression of MHC class I on tumour tissue is well documented. Although the effect of MHC class I expression on the tumorigenicity and immunogenicity of MHC class I negative tumour cell lines has been rigorously studied, less is known about the validity of gene transfer and selection in cell lines with a mixed MHC class I phenotype. To address this issue we identified a C26 cell subline that consists of distinct populations of MHC class I (H-2D/K) positive and negative cells. Transient transfection experiments using liposome-based transfer showed a lower transgene expression in MHC class I negative cells. In addition, MHC class I negative cells were more sensitive to antibiotic selection. This led to the generation of fully MHC class I positive cell lines. In contrast to C26 cells, all transfectants were rejected in vivo and induced protection against the parental tumour cells in rechallenge experiments. Tumour cell specificity of the immune response was demonstrated in in vitro cytokine secretion and cytotoxicity assays. Transfectants expressing CD40 ligand and hygromycin phosphotransferase were not more immunogenic than cells expressing hygromycin resistance alone. We suggest that the MHC class I positive phenotype of the C26 transfectants had a bearing on their immunogenicity, because selected MHC class I positive cells were more immunogenic than parental C26 cells and could induce specific anti-tumour immune responses. These data demonstrate that the generation of tumour cell transfectants can lead to the selection of subpopulations that show an altered phenotype compared to the parental cell line and display altered immunogenicity independent of selection marker genes or other immune modulatory genes. Our results show the importance of monitoring gene transfer in the whole tumour cell population, especially for the evaluation of in vivo therapies targeted to heterogeneous tumour cell populations. 相似文献
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963.
A lost less is known about the morbidity and mortality consequences of male infertility. It was the aim of our study to analyse the association between sperm concentration and individual lifetime mortality in men. The data sources included medical records of 601 men who attended the andrological service at the Marburg University Hospital between 1949 and 1985, and vital data gathered from public registration offices and a statutory health insurance. A Cox regression model estimated a two-fold higher mortality risk for oligozoospermic men as compared to the normozoospermic group for cohorts born between 1892 and 1931. Since a selection bias could not be found, we assume there to be a connection between poor fertility status and a shorter lifespan in men. Possible explanations for the variation in mortality risk are: (i) Lifestyle and health behaviour in adulthood, (ii) conditions in utero, and (iii) genetic dispositions. 相似文献
964.
Light intensity is crucial for plant growth. In this study, the hypothesis was tested whether a sudden increase in light intensity leads to an immediate increase of root growth. Seedlings of Nicotiana tabacum grown in agar-filled Petri dishes were subjected to light intensities of 60 and 300 micromol m(-2) s(-1), respectively. Seedling biomass, sucrose, glucose and fructose concentration as well as primary root growth increased significantly with light intensity. The dynamics of the increase in root growth were analysed here in more detail. In transition experiments from low to high light intensities, root growth increased by a factor of four within 4 d, reaching the steady-state level measured in plants that were cultivated in high-light conditions. The distribution of relative elemental growth rates along the root growth zone retained a constant shape throughout this transition. During the first three hours after light increase, strong growth fluctuations were repeatedly observed with the velocity of the root tip cycling in a sinusoidal pattern between 120 and 180 microm h(-1). These dynamic patterns are discussed in the context of hydraulic and photosynthetic acclimation to the altered conditions. Experiments with externally applied sucrose and with transgenic plants having reduced capacities for sucrose synthesis indicated clearly that increasing light intensity rapidly enhanced root growth by elevating sucrose export from shoot to root. 相似文献
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966.
967.
Funhoff EG Bauer U García-Rubio I Witholt B van Beilen JB 《Journal of bacteriology》2006,188(14):5220-5227
The first and key step in alkane metabolism is the terminal hydroxylation of alkanes to 1-alkanols, a reaction catalyzed by a family of integral-membrane diiron enzymes related to Pseudomonas putida GPo1 AlkB, by a diverse group of methane, propane, and butane monooxygenases and by some membrane-bound cytochrome P450s. Recently, a family of cytoplasmic P450 enzymes was identified in prokaryotes that allow their host to grow on aliphatic alkanes. One member of this family, CYP153A6 from Mycobacterium sp. HXN-1500, hydroxylates medium-chain-length alkanes (C6 to C11) to 1-alkanols with a maximal turnover number of 70 min(-1) and has a regiospecificity of > or =95% for the terminal carbon atom position. Spectroscopic binding studies showed that C6-to-C11 aliphatic alkanes bind in the active site with Kd values varying from approximately 20 nM to 3.7 microM. Longer alkanes bind more strongly than shorter alkanes, while the introduction of sterically hindering groups reduces the affinity. This suggests that the substrate-binding pocket is shaped such that linear alkanes are preferred. Electron paramagnetic resonance spectroscopy in the presence of the substrate showed the formation of an enzyme-substrate complex, which confirmed the binding of substrates observed in optical titrations. To rationalize the experimental observations on a molecular scale, homology modeling of CYP153A6 and docking of substrates were used to provide the first insight into structural features required for terminal alkane hydroxylation. 相似文献
968.
Lipopolysaccharides (LPS; endotoxin) activate immunocompetent cells of the host via a transmembrane signaling process. In this study, we investigated the function of the LPS-binding protein (LBP) in this process. The cytoplasmic membrane of the cells was mimicked by lipid liposomes adsorbed on mica, and the lateral organization of LBP in these membranes and its interaction with LPS aggregates were characterized by atomic force microscopy. Using cantilever tips functionalized with anti-LBP antibodies, single LBP molecules were localized in the membrane at low concentrations. At higher concentrations, LBP formed clusters of several molecules and caused cross-linking of lipid bilayers. The addition of LPS to LBP-containing liposomes led to the formation of LPS domains in the membranes, which could be inhibited by anti-LBP antibodies. Thus, LBP mediates the fusion of lipid membranes and LPS aggregates. 相似文献
969.
970.
Activation by diazoxide and inhibition by 5-hydroxydecanoate are the hallmarks of mitochondrial ATP-sensitive K+ (K(ATP)) channels. Opening of these channels is thought to trigger cytoprotection (preconditioning) through the generation of reactive oxygen species. However, we found that diazoxide-induced oxidation of the widely used reactive oxygen species indicator 2',7'-dichlorodihydrofluorescein in isolated liver and heart mitochondria was observed in the absence of ATP or K+ and therefore independent of K(ATP) channels. The response was blocked by stigmatellin, implying a role for the cytochrome bc1 complex (complex III). Diazoxide, though, did not increase hydrogen peroxide (H2O2) production (quantitatively measured with Amplex Red) in intact mitochondria, submitochondrial particles, or purified cytochrome bc1 complex. We confirmed that diazoxide inhibited succinate oxidation, but it also weakly stimulated state 4 respiration even in K+-free buffer, excluding a role for K(ATP) channels. Furthermore, we have shown previously that 5-hydroxydecanoate is partially metabolized, and we hypothesized that fatty acid metabolism may explain the ability of this putative mitochondrial K(ATP) channel blocker to inhibit diazoxide-induced flavoprotein fluorescence, commonly used as an assay of K(ATP) channel activity. Indeed, consistent with our hypothesis, we found that decanoate inhibited diazoxide-induced flavoprotein oxidation. Taken together, our data question the "mitochondrial K(ATP) channel" hypothesis of preconditioning. Diazoxide did not evoke superoxide (which dismutates to H2O2) from the respiratory chain by a direct mechanism, and the stimulatory effects of this compound on mitochondrial respiration and 2',7'-dichlorodihydrofluorescein oxidation were not due to the opening of K(ATP) channels. 相似文献