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51.
Shuang Liu Hongyan Zhang Jun Dai Shaohu Hu Ignacio Pino Daniel J Eichinger Huibin Lyu Heng Zhu 《MABS-AUSTIN》2015,7(1):110-119
Monoclonal antibodies (mAbs) against human proteins are the primary protein capture reagents for basic research, diagnosis, and molecular therapeutics. The 2 most important attributes of mAbs used in all of these applications are their specificity and avidity. While specificity of a mAb raised against a human protein can be readily defined based on its binding profile on a human proteome microarray, it has been a challenge to determine avidity values for mAbs in a high-throughput and cost-effective fashion. To undertake this challenge, we employed the oblique-incidence reflectivity difference (OIRD) platform to characterize mAbs in a protein microarray format. We first systematically determined the Kon and Koff values of 50 mAbs measured with the OIRD method and deduced the avidity values. Second, we established a multiplexed approach that simultaneously measured avidity values of a mixture of 9 mono-specific mAbs that do not cross-react to the antigens. Third, we demonstrated that avidity values of a group of mAbs could be sequentially determined using a flow-cell device. Finally, we implemented a sequential competition assay that allowed us to bin multiple mAbs that recognize the same antigens. Our study demonstrated that OIRD offers a high-throughput and cost-effective platform for characterization of the binding kinetics of mAbs. 相似文献
52.
Won Il Choi Kwang-Sik Choi Dong-Pyeo Lyu Jung-Su Lee Jongok Lim Seunghwan Lee Sang-Chul Shin Yeong-Jin Chung Young-Seuk Park 《Biodiversity and Conservation》2010,19(8):2291-2305
Fauna assemblages reflect their habitat relating to ecological function in an ecosystem. The functional groups are concerned
with how a resource is processed by different species to provide a specific ecosystem service or function. We elucidated seasonal
changes of coleopteran functional groups in forests, and evaluated their ecological roles related to available food resources.
Coleopteran communities were collected weekly or biweekly using Malaise traps at nine study sites in Japanese red pine forests
in Korea from late June to September 2005. Compositions of the functional groups were compared at the different sites and
at sampling times with respect to taxa richness and abundance. Cluster analysis and non-metric multidimensional scaling were
used to characterize spatial and temporal changes of functional groups. Herbivores and dead/live wood feeders regulating primary
production in the pine forests were the dominant coleopteran groups in July, followed by detritivores and predators that dominated
from July to August, resulting from the accumulation of detritus. Then, fungivores became dominant due to increased fungal
biomass in the forest. Seasonal changes of coleopteran functional groups shifted from regulators of primary production to
regulators of decomposition, reflecting their available food resources. In addition, abundance of detritivores and predators
were dependent on total abundance of coleopterans, suggesting that these two groups reflect their habitat condition. 相似文献
53.
Evolutionarily conserved SR proteins (serine/arginine-rich proteins) are important factors for alternative splicing and their activity is modulated by SRPKs (SR protein-specific kinases). We previously identified Dsk1p (dis1-suppressing protein kinase) as the orthologue of human SRPK1 in fission yeast. In addition to its similarity of gene structure to higher eukaryotes, fission yeast Schizosaccharomyces pombe is a unicellular eukaryotic organism in which alternative splicing takes place. In the present study, we have revealed for the first time that SR proteins, Srp1p and Srp2p, are the in vivo substrates of Dsk1p in S. pombe. Moreover, the cellular localization of the SR proteins and Prp2p splicing factor is dependent on dsk1(+): Dsk1p is required for the efficient nuclear localization of Srp2p and Prp2p, while it promotes the cytoplasmic distribution of Srp1p, thereby differentially influencing the destinations of these proteins in the cell. The present study offers the first biochemical and genetic evidence for the in vivo targets of the SRPK1 orthologue, Dsk1p, in S. pombe and the significant correlation between Dsk1p-mediated phosphorylation and the cellular localization of the SR proteins, providing information about the physiological functions of Dsk1p. Furthermore, the results demonstrate that the regulatory function of SRPKs in the nuclear targeting of SR proteins is conserved from fission yeast to human, indicating a general mechanism of reversible phosphorylation to control the activities of SR proteins in RNA metabolism through cellular partitioning. 相似文献
54.
Bladder cancer-associated protein gene (BLCAP) is a novel candidate tumor suppressor gene identified from the human bladder
carcinoma. Our previous studies have shown that BLCAP overexpression could inhibit cell growth by inducing apoptosis in HeLa
cells [Zuo Z, Zhao M, Liu J, Gao G, Wu X: Tumor Biol 27: 221–226, 2006]. Such evidence suggests the alterations in BLCAP may
play an important role in tumorigenesis. To further study the biological function of the BLCAP gene, we constructed a recombinant
retroviral vector encoding BLCAP cDNA. Overexpressed BLCAP, via stable infection of exogenous BLCAP, resulted in growth inhibition
of the human tongue cancer cell line Tca8113 in vitro, accompanied by S phase cell cycle arrest and apoptosis. The growth inhibition was correlated with up-regulation of p21WAF1/CIP1 expression and down-regulation of Bcl-XL and Bcl-2 expressions. However, p53 expression and NF-κB activity remained unchanged
post infection. Furthermore, no changes in p53 phosphorylation at Ser46 and nuclear localization, which are critical to p53
function, were observed in BLCAP-overexpressed cells. Taken together, BLCAP may play a role not only in regulating cell proliferation
but also in coordinating apoptosis and cell cycle via a novel way independent of p53 and NF-κB.
Jun Yao and Li Duan contributed equally to this work. 相似文献
55.
56.
57.
Background
Protein transduction is safer than viral vector-mediated transduction for the delivery of a therapeutic protein into a cell. Fusion proteins with an arginine-rich cell-penetrating peptide have been produced in E. coli, but the low solubility of the fusion protein expressed in E. coli impedes the large-scale production of fusion proteins from E. coli.Results
Expressed protein ligation is a semisynthetic method to ligate a bacterially expressed protein with a chemically synthesized peptide. In this study, we developed expressed protein ligation-based techniques to conjugate synthetic polyarginine peptides to Cre recombinase. The conjugation efficiency of this technique was higher than 80%. Using this method, we prepared semisynthetic Cre with poly-L-arginine (ssCre-R9), poly-D-arginine (ssCre-dR9) and biotin (ssCre-dR9-biotin). We found that ssCre-R9 was delivered to the cell to a comparable level or more efficiently compared with Cre-R11 and TAT-Cre expressed as recombinant fusion proteins in E. coli. We also found that the poly-D-arginine cell-penetrating peptide was more effective than the poly-L-arginine cell-penetrating peptide for the delivery of Cre into cell. We visualized the cell transduced with ssCre-dR9-biotin using avidin-FITC.Conclusions
Collectively, the results demonstrate that expressed protein ligation is an excellent technique for the production of cell-permeable Cre recombinase with polyarginine cell-penetrating peptides. In addition, this approach will extend the use of cell-permeable proteins to more sophisticated applications, such as cell imaging.Electronic supplementary material
The online version of this article (doi:10.1186/s12896-015-0126-z) contains supplementary material, which is available to authorized users. 相似文献58.
Xiangyang Wu Yong Wu Ruijuan Zheng Fen Tang Lianhua Qin Detian Lai Lu Zhang Lingming Chen Bo Yan Hua Yang Yang Wang Feifei Li Jinyu Zhang Fei Wang Lin Wang Yajuan Cao Mingtong Ma Zhonghua Liu Jianxia Chen Xiaochen Huang Jie Wang Ruiliang Jin Peng Wang Qin Sun Wei Sha Liangdong Lyu Pedro MouraAlves Anca Dorhoi Gang Pei Peng Zhang Jiayu Chen Shaorong Gao Felix Randow Gucheng Zeng Chang Chen XinShan Ye Stefan H E Kaufmann Haipeng Liu Baoxue Ge 《EMBO reports》2021,22(7)
Mycobacterial arabinogalactan (AG) is an essential cell wall component of mycobacteria and a frequent structural and bio‐synthetical target for anti‐tuberculosis (TB) drug development. Here, we report that mycobacterial AG is recognized by galectin‐9 and exacerbates mycobacterial infection. Administration of AG‐specific aptamers inhibits cellular infiltration caused by Mycobacterium tuberculosis (Mtb) or Mycobacterium bovis BCG, and moderately increases survival of Mtb‐infected mice or Mycobacterium marinum‐infected zebrafish. AG interacts with carbohydrate recognition domain (CRD) 2 of galectin‐9 with high affinity, and galectin‐9 associates with transforming growth factor β‐activated kinase 1 (TAK1) via CRD2 to trigger subsequent activation of extracellular signal‐regulated kinase (ERK) as well as induction of the expression of matrix metalloproteinases (MMPs). Moreover, deletion of galectin‐9 or inhibition of MMPs blocks AG‐induced pathological impairments in the lung, and the AG‐galectin‐9 axis aggravates the process of Mtb infection in mice. These results demonstrate that AG is an important virulence factor of mycobacteria and galectin‐9 is a novel receptor for Mtb and other mycobacteria, paving the way for the development of novel effective TB immune modulators. 相似文献
59.
Zhong J Kim HT Lyu J Yoshikawa K Nakafuku M Lu W 《Development (Cambridge, England)》2011,138(3):409-419
GABAergic neurons and oligodendrocytes originate from progenitors within the ventral telencephalon. However, the molecular mechanisms that control neuron-glial cell-fate segregation, especially how extrinsic factors regulate cell-fate changes, are poorly understood. We have discovered that the Wnt receptor Ryk promotes GABAergic neuron production while repressing oligodendrocyte formation in the ventral telencephalon. We demonstrate that Ryk controls the cell-fate switch by negatively regulating expression of the intrinsic oligodendrogenic factor Olig2 while inducing expression of the interneuron fate determinant Dlx2. In addition, we demonstrate that Ryk is required for GABAergic neuron induction and oligodendrogenesis inhibition caused by Wnt3a stimulation. Furthermore, we showed that the cleaved intracellular domain of Ryk is sufficient to regulate the cell-fate switch by regulating the expression of intrinsic cell-fate determinants. These results identify Ryk as a multi-functional receptor that is able to transduce extrinsic cues into progenitor cells, promote GABAergic neuron formation, and inhibit oligodendrogenesis during ventral embryonic brain development. 相似文献
60.
A three-step electrodeposition method has been successfully adopted to fabricate morphology-controlled novel Au microspheres on self-doped polyaniline nanofibers (nanoSPAN) modified glassy carbon electrode. The deposition conditions, such as HAuCl(4) concentration and deposition step, have significant influences on the morphologies and electrochemical properties of the resulted Au microspheres. Well hierarchical and homogeneously dispersed flower-like Au microspheres (HHFAu) were obtained under optimal conditions by the three-step electrodeposition strategy in 5.0mM HAuCl(4) solution. HHFAu possess large surface area, excellent electron transfer ability and good biocompatibility. The DNA probe could be effectively attached to HHFAu and thus a high-performance DNA biosensor was constructed by using electrochemical impedance spectroscopy as detection method. A gene fragment of the cauliflower mosaic virus 35S gene, which is related to one of the screening genes for the transgenically modified plants, has been satisfactorily detected. The linear range was from 1.0 × 10(-13)M to 1.0 × 10(-6)M and the detection limit was 1.9 × 10(-14)M. This HHFAu/nanoSPAN-based impedance biosensing platform holds great promise for the detection of other biological and chemical molecules. 相似文献