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11.
The cellular trafficking and zinc dependence of secretory and lysosomal sphingomyelinase, two products of the acid sphingomyelinase gene 总被引:2,自引:0,他引:2
SL Schissel GA Keesler EH Schuchman KJ Williams I Tabas 《Canadian Metallurgical Quarterly》1998,273(29):18250-18259
The acid sphingomyelinase (ASM) gene, which has been implicated in ceramide-mediated cell signaling and atherogenesis, gives rise to both lysosomal SMase (L-SMase), which is reportedly cation-independent, and secretory SMase (S-SMase), which is fully or partially dependent on Zn2+ for enzymatic activity. Herein we present evidence for a model to explain how a single mRNA gives rise to two forms of SMase with different cellular trafficking and apparent differences in Zn2+ dependence. First, we show that both S-SMase and L-SMase, which contain several highly conserved zinc-binding motifs, are directly activated by zinc. In addition, SMase assayed from a lysosome-rich fraction of Chinese hamster ovary cells was found to be partially zinc-dependent, suggesting that intact lysosomes from these cells contain subsaturating levels of Zn2+. Analysis of Asn-linked oligosaccharides and of N-terminal amino acid sequence indicated that S-SMase arises by trafficking through the Golgi secretory pathway, not by cellular release of L-SMase during trafficking to lysosomes or after delivery to lysosomes. Most importantly, when Zn2+-dependent S-SMase was incubated with SMase-negative cells, the enzyme was internalized, trafficked to lysosomes, and became zinc-independent. We conclude that L-SMase is exposed to cellular Zn2+ during trafficking to lysosomes, in lysosomes, and/or during cell homogenization. In contrast, the pathway targeting S-SMase to secretion appears to be relatively sequestered from cellular pools of Zn2+; thus S-SMase requires exogeneous Zn2+ for full activity. This model provides important information for understanding the enzymology and regulation of L- and S-SMase and for exploring possible roles of ASM gene products in cell signaling and atherogenesis. 相似文献
12.
AH Lauber TJ Barrett M Subramaniam M Schuchard TC Spelsberg 《Canadian Metallurgical Quarterly》1997,272(39):24657-24665
The receptor-binding factor (RBF) for the avian oviduct progesterone (Pg) receptor (PR) has previously been shown to be a unique 10-kDa nuclear matrix protein that generates high affinity PR-binding sites on avian DNA. This paper describes the use of Southwestern blot and DNA gel shift analyses with RBF protein to identify a minimal 54-base pair RBF-binding element in the matrix-associated region (MAR) of the Pg-regulated c-myc gene promoter. This element contains a 5'-GC-rich domain and a 3'-AT-rich domain, the latter of which has a homopurine/homopyrimidine structure. The gel shift assays required the generation of an RBF-maltose fusion protein (RBF-MBP), which specifically binds this element and is supershifted when the anti-RBF polyclonal antibody is added. Computer analysis of the full-length amino acid sequence for RBF predicts a DNA-binding motif involving a beta-sheet structure at the N-terminal domain. Southern blot analyses using nuclear matrix DNA suggests that there are dual MAR sites in the c-myc promoter, which flank an intervening domain containing the RBF element. The co-transfection of this MAR sequence, containing the RBF element and cloned into a luciferase reporter vector, together with an RBF expression vector construct, into steroid treated human MCF-7 cells, results in a decrease of the c-myc promoter activity relative to control transfections containing only the parent vector of the RBF expression construct. These data suggest that a unique chromatin/nuclear matrix structure, composed of the RBF-DNA element complex which is flanked by nuclear matrix attachment sites, serves to bind the PR and repress the c-myc promoter. 相似文献
13.
BACKGROUND: To determine the best cutoff values of aspartate aminotransferase (AST) and alanine amino-transferase (ALT) in detecting viral hepatitis C infection among patients of continuous ambulatory peritoneal dialysis (CAPD). METHODS: 90 (44 male and 46 female) CAPD patients and 526 adult controls (266 male, 260 female) were enrolled. Serum AST and ALT were measured by an auto-analyser monthly. Serum HBsAg was examined using a RIA method and anti-HCV by an second-generation EIA method. The best cutoff values of AST and ALT for detecting viral hepatitis were obtained from the ROC (receiver-operating characteristic) curve. RESULTS: The prevalence of anti-HCV(+) was significantly higher in CAPD patients (16.7%) than in normal controls (4.9%), while that of HBsAg(+) was similar in both groups. CAPD patients had significantly lower levels of serum aminotransferases compared to normal controls. Mean AST were 23.8 IU/l in normal control and 18.8 IU/l in the CAPD patients (P < 0.001). Mean ALT were 21.9 IU/l in normal controls and 15.3 IU/l in the CAPD patients (P < 0.001). CAPD patients with HCV infection had higher serum AST and ALT levels than those without. However, HBV infection did not cause significant serum aminotransferase elevation in patients. The conventional cutoff values of AST (40 IU/l) and ALT (40 IU/l) for detecting viral hepatitis yielded only a sensitivity of 27.3 and 18.2% respectively; on the contrary, our revised cutoff values of AST (24 IU/l) and ALT (17 IU/l) had better sensitivities (AST, 72.7%; ALT, 63.6%). For serial aminotransferase values, the sensitivity of AST and ALT for detecting HCV were 36.4 and 27.3% by conventional criteria, and were both 81.8%, by our newly revised criteria. CONCLUSIONS: Serum aminotransferase cutoff values should be modified for screening viral hepatitis in a CAPD population. Our new cutoff criteria had important clinical implications in providing benefits of earlier detection and possible prevention from chronic hepatic deteriorations. 相似文献
14.
RM Campbell EP Heimer M Ahmad HG Eisenbeis TJ Lambros Y Lee RW Miller PR Stricker AM Felix 《Canadian Metallurgical Quarterly》1997,49(6):527-537
In the present study, human growth hormone-releasing factor (hGRF) and analogs were successfully pegylated at the carboxy-terminus using a novel solid- and solution-phase strategy. Following synthesis, these pegylated hGRF analogs were evaluated for in vitro and in vivo biological activity. Specifically, hGRF (1-29)-NH2, [Ala15]-hGRF (1-29)-NH2, [desNH2Tyr1, D-Ala2, Ala15]-hGRF(1-29)-NH2 and [His1, Val2, Gln8, Ala15, Leu27]-hGRF(1-32)-OH were each C-terminally extended using a Gly-Gly-Cys-NH2 spacer (previously demonstrated not to alter intrinsic biological activity), and then monopegylated via coupling to an activated dithiopyridyl-PEG reagent. PEG moieties of 750, 2000, 5000 or 10,000 molecular weight (MW) were examined to determine the effect of polymer weight on activity. Initial biological evaluations in vitro revealed that all C-terminally pegylated hGRF analogs retained high growth hormone (GH)-releasing potencies, regardless of the MW of PEG polymer employed. Two of these pegylated hGRF analogs, [desNH2Tyr1, D-Ala2, Ala15]-hGRF (1-29)-Gly-Gly-Cys(NH2)-S-Nle-PEG5000 and [His1, Val2, Gln8, Ala15, Leu27]-hGRF(1-32)-Gly-Cys(NH2)-S-Nle-PEG5000, were subsequently evaluated in both pig and mouse models and found to be highly potent (in vivo potency range = 12-55-fold that of native hGRF). Relative to their non-pegylated counterparts, these two pegylated hGRF analogs exhibited enhanced duration of activity. 相似文献
15.
J Brake TJ Walsh CE Benton JN Petitte R Meijerhof G Pe?alva 《Canadian Metallurgical Quarterly》1997,76(1):144-151
The relative importance of fat and lean tissue mass in determining bone mineral mass among postmenopausal women was examined in this 1-year longitudinal study. Fifty postmenopausal Caucasian women entered the study; 45 of them completed a 1-year follow-up. Dual-energy X-ray absorptiometry was employed for measuring total and regional bone mineral density (BMD) and bone mineral content (BMC), fat tissue mass (FTM), lean tissue mass (LTM), and body weight. Results from linear regression analysis using the cross-sectional data (n = 50) of the study indicated that LTM explained a larger percentage of variation in bone mineral mass than did FTM. FTM and LTM were found to be moderately correlated (r = 0.55); when FTM was entered in the same predicting regression models, LTM was a significant predictor (p < 0.05) of the total and regional BMC, but not BMD. The percent FTM (and inversely %LTM) was correlated with BMD and BMC, but significant correlation was primarily found only for total body BMD (or BMC). Weight was the best predictor of total body BMD and BMC. Longitudinally (n = 45), annual changes in both FTM and weight were significantly associated with annual changes in regional BMD after adjustment for initial bone mineral values (p < 0.05). We conclude that bone mineral mass is more closely related to LTM than to FTM, while annual changes in regional BMD are more closely correlated with changes in FTM in healthy postmenopausal women. Meanwhile, increased body weight is significantly associated with increased bone mineral mass. 相似文献
16.
17.
Although not well known, the University of Toronto had a very early computer-development program and in 1952 was one of the first few institutions with an operable computer in North America. This article describes the university's initial attempt to build the UTEC computer and how it acquired the pioneering Ferut machine 相似文献
18.
A Meneghetti WV Cardoso JS Brody MC Williams 《Canadian Metallurgical Quarterly》1996,44(10):1173-1182
Explants of embryonic lung are often used to characterize lung growth, bronchial tree pattern, and cell differentiation. Most investigators culture lungs for 3-7 days in defined media lacking, e.g., added growth factors or hormones. If growth and differentiation are comparable to that in vivo, these cultures show considerable promise for identifying developmental regulatory molecules and target genes, and for elucidating molecular responses. We used in situ hybridization and RT-PCR to compare times and sites of expression of mRNAs of six epithelial genes in cultured and uncultured fetal rat lungs. These genes, expressed in distal lung of adult rats, are surfactant proteins (SP) A, B, and C; LAR, a receptor-type tyrosine phosphatase; Clara cell secretory protein (CC10, CCSP); and T1alpha. SP-A, SF-B, LAR, and CC10 are expressed by both Clara and Type II cells in adult animals. SP-C and T1alpha are unique markers for Type II and Type I cells, respectively. SP-C, LAR, and T1alpha are expressed before the lung is explanted (Day 13.5); SP-A, -B, and CC10 mRNAs are first detected later. The onset of expression is similar in vivo and in vitro. Although the patterns of expression differ for each mRNA, their sites of expression in culture match those in vivo relative to the bronchial tree. The explanted embryonic lung appears to be an excellent experimental model. 相似文献
19.
C Chryssanthopoulos C Williams W Wilson L Asher L Hearne 《Canadian Metallurgical Quarterly》1994,4(4):374-386
The purpose of this study was to compare the effects of a carbohydrate-electrolyte solution, ingested during exercise, with the effects of a preexercise carbohydrate meal on endurance running performance. Ten endurance-trained males completed two 30-km treadmill runs. In one trial subjects consumed a placebo solution 4 hr before exercise and a carbohydrate-electrolyte solution immediately before exercise and every 5 km (C). In the other trial, subjects consumed a 4-hr preexercise high-carbohydrate meal and water immediately before exercise and every 5 km (M). Performance times were identical for M and C, and there was no difference in the self-selected speeds. Oxygen uptake, heart rates, perceived rate of exertion, and respiratory exchange ratios were also similar. However, blood glucose concentration was higher in C during the first 20 km of the 30-km run. In M, blood glucose concentration was maintained above 4.5 mmol.L-1 throughout exercise. Thus, the two conditions produced the same 30-km treadmill running performance time. 相似文献
20.
J. C. Williams R. G. Baggerly N. E. Paton 《Metallurgical and Materials Transactions A》2002,33(3):837-850
Single crystals of Ti-Al alloys containing 1.4, 2.9, 5, and 6.6 pct Al (by weight) were oriented for 〈a〉 slip on either basal or prism planes or loaded parallel along the c-axis to enforce a nonbasal deformation mode. Most of the tests were conducted in compression and at temperatures between
77 and 1000 K. Trace analysis of prepolished surfaces enabled identification of the twin or slip systems primarily responsible
for deformation. Increasing the deformation temperature, Al content, or both, acted to inhibit secondary twin and slip systems,
thereby increasing the tendency toward strain accommodation by a single slip system having the highest resolved stress. In
the crystals oriented for basal slip, transitions from twinning to multiple slip and, finally, to basal slip occurred with
increasing temperature in the lower-Al-content alloys, whereas for Ti-6.6 pct Al, only basal slip was observed at all temperatures
tested. A comparison of the critically resolved shear stress (CRSS) values for basal and prism slip as a function of Al content
shows that prism slip is favored at room temperature in pure Ti, but the stress to activate these two systems becomes essentially
equal in the Ti-6.6 pct Al crystals over a wide range of temperatures.
Compression tests on crystals oriented so that the load was applied parallel to the c-axis showed extensive twinning in lower Al concentrations and 〈c+a〉 slip at higher Al concentrations, with a mixture of 〈c+a〉 slip and twinning at intermediate compositions. A few tests also were conducted in tension, with the load applied parallel
to the c-axis. In these cases, twinning was observed, and the resolved shear for plastic deformation by twinning was much lower that
that for 〈c+a〉 slip observed in compression loading.
This article is based on a presentation made in the symposium entitled “Defect Properties and Mechanical Behavior of HCP Metals
and Alloys” at the TMS Annual Meeting, February 11–15, 2001, in New Orleans, Louisiana, under the auspices of the following
ASM committees: Materials Science and Critical Technology Sector, Structural Materials Division, Electronic, Magnetic & Photonic
Materials Division, Chemistry & Physics of Materials Committee, Joint Nuclear Materials Committee, and Titanium Committee. 相似文献