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251.
Eva Adamová Marcela Lišková Eva Matalová Karel Klepárník 《Analytical and bioanalytical chemistry》2014,406(22):5389-5394
Caspases are key enzymes activated during the apoptotic machinery. Apoptosis as a way of programmed cell death becomes deregulated in some pathologies including cancer transformations, neurodegenerative, or autoimmune diseases. Most of the methods available for the detection of apoptosis and caspases provide qualitative information only or quantification data as an average from cell populations or cell lysates. Several reports point to the importance of more accurate single-cell analyses in biomedical studies due to heterogeneity at tissue as well as cell level. To meet these requirements, we developed a miniaturized device enabling detection and quantification of active caspase-3/7 in individual cells at a femtogram level (10?15 g). The active caspase-3/7 detection protocol is based on the bioluminescence chemistry commercially available as a Caspase-Glo? 3/7 reagent developed by Promega. As a model, we used human stem cells treated by camptothecin to induce apoptosis. Individual apoptotic cells were captured from a culture medium under a microscope and transferred by a micromanipulation system into a detection capillary containing 2 μl of the reagent. Cells without activation by camptothecin served as negative controls. The detection limit of active caspase-3/7 achieved in the miniaturized system was determined as 0.20 and limit of quantification as 0.65 of the amount found in a single apoptotic human stem cell. Such a sensitive method could have a wide application potential in laboratory medicine and related clinically oriented research. Figure
Bioluminescence detection assembly 相似文献
252.
Yamile Jalit Mónica Moreno Fabiana A. Gutierrez Alberto Sanchez Arribas Manuel Chicharro Esperanza Bermejo Antonio Zapardiel Concepción Parrado Gustavo A. Rivas Marcela C. Rodríguez 《Electroanalysis》2013,25(5):1116-1121
We report the advantages of the adsorption and electrooxidation of oligonucleotides and calf‐thymus double stranded DNA (dsDNA) at glassy carbon electrodes (GCE) modified with a dispersion of multiwalled carbon nanotubes (MWCNT) in poly‐L ‐lysine (Plys) (GCE/MWCNT‐Plys). Important enhancement in the guanine oxidation signal was obtained by adsorptive stripping voltammetry (AdSV) due to a most favorable interaction between the negatively charged DNAs and the positively charged Plys that support the MWCNT. The layer of oligo(dG)11 immobilized at GCE/MWCNT‐Plys was successfully used for the selective detection of the hybridization event using oligo(dG)11/oligo(dC)11 as model. 相似文献
253.
Marcela P. Garcia Ammar Shahid Jennifer Y. Chen Jun Xi 《Analytical and bioanalytical chemistry》2013,405(4):1153-1158
Epidermal growth factor receptor (EGFR) plays a major role in cell migration and invasion and is considered to be the primary source of activation of various malignant tumors. To gain insight into how elevated levels of EGFR influence cellular function, particularly cell motility, we used a quartz crystal microbalance with dissipation monitoring (QCM-D) to examine restructuring of focal adhesions in MCF-10A cells induced by epidermal growth factor. Engineered cells that overexpress epidermal growth factor receptor (EGFR) exhibited a very different kinetic profile from wildtype MCF-10A cells that have a lower level of EGFR with a higher rate for the initial disassembly of focal adhesion and a much lower rate for the later reassembly of focal adhesions. It is conceivable that these effects exhibited by EGFR-overexpressing cells may promote the initiation and maintenance of a more favorable adhesion state for cell migration. This study has demonstrated the capability of the dissipation monitoring function of the QCM-D to quantitatively assess kinetic aspects of cellular processes with a high temporal resolution and sensitivity. Figure
Characterization of the effects of the expression level of epidermal growth factor receptor on the kinetics of the epidermal growth factor-induced restructuring of focal adhesions with the quartz crystal microbalance with dissipation monitoring. 相似文献
254.
Marcela Pavan Bagagli Hélia Harumi Sato 《Applied biochemistry and biotechnology》2013,170(5):1057-1065
Transglutaminase catalyzes the cross-linking reaction between a glutamine residue and a free amine residue of proteins leading to the formation of protein aggregates. In this research, the effects of temperature, agitation, and aeration on the production of transglutaminase in a bench reactor by a newly isolated Streptomyces sp. from Brazilian soils were investigated using a factorial experimental design. The parameters evaluated influenced the enzyme production, and the data showed that the best conditions to enhance cell growth were different from those leading to enhanced transglutaminase production. Thus, a temperature and agitation shift strategy was adopted to increase transglutaminase productivity. The temperature and agitation were first set at 34 °C and 350 rpm, respectively, and after 24 h decreasing to 26 °C and 150 rpm until the end of fermentation. The transglutaminase activity obtained was 2.18 U/mL after 42 h of fermentation, which was twice than that obtained using a constant temperature and agitation fermentation strategy. 相似文献
255.
256.
Pablo C. Schulz Paula Messina Marcela A. Morini Bruno Vuano 《Colloid and polymer science》2002,280(12):1104-1109
The scarcely studied sodium dehydrocholate-water system was investigated at low concentration by using H+-, Na+- and dehydrocholate-ion-selective electrodes. The findings at concentrations below the critical micelle concentration are consistent with those of previous work. A dimeric chelate structure entrapping Na+ ions at very low concentration is proposed. The composition of the micelles and the degree of ionisation were studied and explained as the result of the possibility of a back-to-face mechanism of aggregation of this steroid salt. This mechanism also explains the possibility of formation of acid soaplike aggregates at intermediate concentrations. 相似文献
257.
Carolina V. Barra Fillipe V. Rocha Adelino V. G. Netto B. Shimura Regina C. G. Frem Antonio E. Mauro Iracilda Z. Carlos Sandra R. Ananias Marcela B. Quilles 《Journal of Thermal Analysis and Calorimetry》2011,106(2):483-488
The pyrazole ligand 3,5-dimethyl-4-iodopyrazole (HdmIPz) has been used to obtain a series of palladium(II) complexes (1–4) of the type [PdX2(HdmIPz)2] {X = Cl− (1); Br− (2); I− (3); SCN− (4)}. All compounds have been isolated, purified, and characterized by means of elemental analysis, IR spectroscopy, 1H and 13C{1H}-NMR experiments, differential thermal analysis (DTA), and thermogravimetry (TG). The TG/DTA curves showed that the compounds
released ligands in the temperature range 137–605 °C, yielding metallic palladium as final residue. The complexes and the
ligand together with cisplatin have been tested in vitro by MTT assay for their cytotoxicity against two murine cancer cell
lines: mammary adenocarcinoma (LM3) and lung adenocarcinoma (LP07). 相似文献
258.
Wang H Ko YJ García LG Sen P Beltrán MR Bowen KH 《Physical chemistry chemical physics : PCCP》2011,13(17):7685-7691
Anion photoelectron spectroscopic experiments and calculations based on density functional theory have been used to investigate and uniquely identify the structural, electronic, and magnetic properties of both neutral and anionic (Rh(m)Co(n)) and (Rh(m)Co(n))(-) (m=1-5, n=1-2) clusters, respectively. Negative ion photoelectron spectra are presented for electron binding energies up to 3.493 eV. The calculated electron affinities and vertical detachment energies are in good agreement with the measured values. Computational results for geometric structures and magnetic moments of both cluster anions and their neutrals are presented. 相似文献
259.
260.
Rosende M Miró M Segundo MA Lima JL Cerdà V 《Analytical and bioanalytical chemistry》2011,400(7):2217-2227
An automated dynamic leaching test integrated in a portable flow-based setup is herein proposed for reliable determination
of readily bioaccessible Cr(VI) under worst-case scenarios in soils containing varying levels of contamination. The manifold
is devised to accommodate bi-directional flow extraction followed by processing of extracts via either in-line clean-up/preconcentration
using multi-walled carbon nanotubes or automatic dilution at will, along with Cr(VI) derivatization and flow-through spectrophotometric
detection. The magnitude of readily mobilizable Cr(VI) pools was ascertained by resorting to water extraction as promulgated
by current standard leaching tests. The role of carbon nanomaterials for the uptake of Cr(VI) in soil leachates and the configuration
of the packed column integrated in the flow manifold were investigated in detail. The analytical performance of the proposed
system for in vitro bioaccessibility tests was evaluated in chromium-enriched soils at environmentally relevant levels and
in a standard reference soil material (SRM 2701) with a certified value of total hexavalent chromium. The automated method
was proven to afford unbiased assessment of water-soluble Cr(VI) in soils as a result of the minimization of the chromium
species transformation. By combination of the kinetic leaching profile and a first-order leaching model, the water-soluble
Cr(VI) fraction in soils was determined in merely 6 h against >24 h taken in batchwise steady-state standard methods. 相似文献