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101.
102.
The use of phospholipid vesicles as reaction containers, as vehicles for pharmaceutical drug delivery, and as model systems for cells has prompted the development of new methods for analyzing the structure of vesicles and their contents. The pH of the interior of vesicles is of particular interest when analytes are encapsulated and concentrated with the use of a pH gradient. While the interior pH is generally measured for large populations of vesicles, we report the measurement of the interior pH of individual vesicles as their buffer contents are titrated by transfer of N-methylbutylamine (NMBA) into the vesicle by a pH gradient. The initially acidic buffer within the vesicles is titrated along with a small concentration of an encapsulated pH sensitive dye, 5,6-carboxy SNARF-1-dextran. Images of the indicator fluorescence from each vesicle and its dispersed fluorescence spectrum are recorded using epi-illumination spectral fluorescence microscopy. From a fit of the spectra to the respective acid and base forms of the fluorescent indicator, the interior pH of individual vesicles as a function of the concentration of the NMBA titrant in the external solution could be determined. 相似文献
103.
When proteins are conjugated to fluorescent organic dyes, fluorescence emission of the dye molecules is usually decreased, sometimes up to 50-70%. This quenching phenomenon has been acknowledged for decades, but as yet, there are no simple, practical methods to control the fluorescence of dyes conjugated to proteins, especially for dyes conjugated to immunoglobulins. Here, we report that the addition of (2-hydroxypropyl)-β-cyclodextrin (HPβCD) to dye-antibody conjugates can increase fluorescence up to 2.5-fold in cell imaging and flow analysis. This method may be an effective way to increase the sensitivity of detection of fluorescent organic labels used in immunology, histochemistry, and cell biology. 相似文献
104.
While the water permeability of the plasma membranes of mammalian cells has been studied extensively, water transport across membranes of subcellular compartments (e.g., lysosomes, macropinosomes) has been difficult to study. Here we demonstrate a new method for measuring water flux in late endosomes and lysosomes of intact living cells using time-lapse fluorescence microscopy. Cells were loaded by fluid-phase uptake with a mixture of the Lucifer Yellow dextran (LY-dex), a D(2)O sensitive dye, and a D(2)O insensitive control dye, Alexa fluor 546 dextran (AF546-dex). LY-dex responded linearly to changes in D(2)O concentration and the LY-dex D(2)O sensitivity was not affected by changes in pH, physiological salt, and protein concentrations. The co-loaded control dye, AF546-dex, showed no signal changes as a function of D(2)O concentration. To measure membrane water flux, the LY-dex fluorescence in labeled organelles was recorded during rapid superfusion of cells with isotonic buffers prepared in D(2)O. The time constant of water exchange across the lysosomal membrane of intact cells was determined by fitting the data to a single exponential function. From these data, together with the measured area of the organelles, observed water permeability for intracellular CHO-K1 lysosomes was calculated to be 5.3 × 10(-3) ± 0.3 × 10(-3) cm/s. This work demonstrates the feasibility of measuring water flux into subcellular organelles in live cells using LY-dex. 相似文献
105.
Brian D. Badgley John FergusonAmy Vanden Heuvel Gregory T. KleinheinzColleen M. McDermott Todd R. SandrinJulie Kinzelman Emily A. JunionMuruleedhara N. Byappanahalli Richard L. WhitmanMichael J. Sadowsky 《Water research》2011,45(2):721-731
High concentrations of Escherichia coli in mats of Cladophora in the Great Lakes have raised concern over the continued use of this bacterium as an indicator of microbial water quality. Determining the impacts of these environmentally abundant E. coli, however, necessitates a better understanding of their ecology. In this study, the population structure of 4285 Cladophora-borne E. coli isolates, obtained over multiple three day periods from Lake Michigan Cladophora mats in 2007-2009, was examined by using DNA fingerprint analyses. In contrast to previous studies that have been done using isolates from attached Cladophora obtained over large time scales and distances, the extensive sampling done here on free-floating mats over successive days at multiple sites provided a large dataset that allowed for a detailed examination of changes in population structure over a wide range of spatial and temporal scales. While Cladophora-borne E. coli populations were highly diverse and consisted of many unique isolates, multiple clonal groups were also present and accounted for approximately 33% of all isolates examined. Patterns in population structure were also evident. At the broadest scales, E. coli populations showed some temporal clustering when examined by year, but did not show good spatial distinction among sites. E. coli population structure also showed significant patterns at much finer temporal scales. Populations were distinct on an individual mat basis at a given site, and on individual days within a single mat. Results of these studies indicate that Cladophora-borne E. coli populations consist of a mixture of stable, and possibly naturalized, strains that persist during the life of the mat, and more unique, transient strains that can change over rapid time scales. It is clear that further study of microbial processes at fine spatial and temporal scales is needed, and that caution must be taken when interpolating short term microbial dynamics from results obtained from weekly or monthly samples. 相似文献
106.
Emily Y.X. Soh Belinda Yuen 《Cities》2011,28(1):3-10
This paper interrogates the socio-spatial environment that is emerging in Singapore with its development of a globalised economy. Using the case studies of two of the city’s dichotomised spaces - the spectacular downtown and gentrified heartland - the paper seeks to examine the resulting spatial reconfigurations, challenges and unintended consequences. While the common assertion is that state-led planning is effective at the macro level, at the micro level against the accelerated change and emergence of the multi-layered package of spaces of globalisation, the planning of the city is no longer limited to physical planning. Like many emergent economies of the global south, the challenge of how to expand emerging sectors, support Singapore’s economic aspirations and meet social objectives remains. 相似文献
107.
Vitalii Kim Emily Gudvangen Oleg Kondratiev Luis Redondo Shu Xiao Andrei G. Pakhomov 《International journal of molecular sciences》2021,22(13)
Intense pulsed electric fields (PEF) are a novel modality for the efficient and targeted ablation of tumors by electroporation. The major adverse side effects of PEF therapies are strong involuntary muscle contractions and pain. Nanosecond-range PEF (nsPEF) are less efficient at neurostimulation and can be employed to minimize such side effects. We quantified the impact of the electrode configuration, PEF strength (up to 20 kV/cm), repetition rate (up to 3 MHz), bi- and triphasic pulse shapes, and pulse duration (down to 10 ns) on eliciting compound action potentials (CAPs) in nerve fibers. The excitation thresholds for single unipolar but not bipolar stimuli followed the classic strength–duration dependence. The addition of the opposite polarity phase for nsPEF increased the excitation threshold, with symmetrical bipolar nsPEF being the least efficient. Stimulation by nsPEF bursts decreased the excitation threshold as a power function above a critical duty cycle of 0.1%. The threshold reduction was much weaker for symmetrical bipolar nsPEF. Supramaximal stimulation by high-rate nsPEF bursts elicited only a single CAP as long as the burst duration did not exceed the nerve refractory period. Such brief bursts of bipolar nsPEF could be the best choice to minimize neuromuscular stimulation in ablation therapies. 相似文献
108.
109.
Woods Steven Paul; Rippeth Julie D.; Conover Emily; Gongvatana Assawin; Gonzalez Raul; Carey Catherine L.; Cherner Mariana; Heaton Robert K.; Grant Igor 《Canadian Metallurgical Quarterly》2005,19(1):35
Methamphetamine (MA) dependence is associated with deficits in episodic verbal memory, but the cognitive mechanisms underlying such impairments are not known. The authors evaluated a component process model of episodic verbal memory in 87 persons with MA dependence (MA+) and 71 demographically similar non-MA-using controls (MA-). Compared with MA- controls, MA+ participants demonstrated deficient overall learning, free recall, and utilization of semantic clustering, as well as higher rates of repetitions and intrusions. No between-groups differences were evident on measures of serial clustering, retention, or recognition discrimination. Taken together, these findings indicate that MA dependence is associated with deficient strategic (i.e., executive) control of verbal encoding and retrieval, which is consistent with the sequelae of MA-related prefronto-striatal circuit neurotoxicity. (PsycINFO Database Record (c) 2010 APA, all rights reserved) 相似文献
110.