全文获取类型
收费全文 | 1585篇 |
免费 | 146篇 |
专业分类
1731篇 |
出版年
2023年 | 5篇 |
2022年 | 17篇 |
2021年 | 27篇 |
2020年 | 16篇 |
2019年 | 19篇 |
2018年 | 31篇 |
2017年 | 22篇 |
2016年 | 53篇 |
2015年 | 72篇 |
2014年 | 95篇 |
2013年 | 98篇 |
2012年 | 145篇 |
2011年 | 123篇 |
2010年 | 88篇 |
2009年 | 78篇 |
2008年 | 120篇 |
2007年 | 123篇 |
2006年 | 101篇 |
2005年 | 104篇 |
2004年 | 75篇 |
2003年 | 72篇 |
2002年 | 89篇 |
2001年 | 15篇 |
2000年 | 14篇 |
1999年 | 19篇 |
1998年 | 31篇 |
1997年 | 11篇 |
1996年 | 12篇 |
1995年 | 8篇 |
1994年 | 6篇 |
1993年 | 14篇 |
1992年 | 6篇 |
1991年 | 5篇 |
1990年 | 3篇 |
1989年 | 1篇 |
1988年 | 2篇 |
1987年 | 2篇 |
1986年 | 3篇 |
1982年 | 2篇 |
1976年 | 2篇 |
1975年 | 1篇 |
1971年 | 1篇 |
排序方式: 共有1731条查询结果,搜索用时 15 毫秒
31.
Both the concept and the application of the impact factor (IF) have been subject to widespread critique, including concerns over its potential manipulation. This study provides a systematic analysis of significant journal Impact Factor changes, based on the relative contribution of either one or both variables of the IF equation (i.e. citations / articles as the numerator / denominator of the quotient). A cohort of JCR-listed journals which faced the most dramatic absolute IF changes between 2013 and 2014 (ΔIF ≥ 3.0, n = 49) was analyzed for the causes resulting in IF changes that theses journals have experienced in the last five years. Along with the variation by number of articles and citations, this analysis includes the relative change of both variables compared to each other and offers a classification of `valid`and `invalid`scenarios of IF variation in terms of the intended goal of the IF to measure journal quality. The sample cohort features a considerable incidence of IF increases (18%) which are qualified as `invalid`according to this classification because the IF increase is merely based on a favorably changing number of articles (denominator). The results of this analysis point out the potentially delusive effect of IF increases gained through effective shrinkage of publication output. Therefore, careful consideration of the details of the IF equation and possible implementation of control mechanisms versus the volatile factor of number of articles may help to improve the expressiveness of this metric. 相似文献
32.
Achim von Bomhard Alexander Els?sser Lucas Maximilian Ritschl Silke Schwarz Nicole Rotter 《PloS one》2016,11(2)
Vitrification of endothelial cells (MHECT-5) has not previously been compared with controlled slow freezing methods under standardized conditions. To identify the best cryopreservation technique, we evaluated vitrification and standardized controlled-rate -1°C/minute cell freezing in a -80°C freezer and tested four cryoprotective agents (CPA), namely dimethyl sulfoxide (DMSO), ethylene glycol (EG), propylene glycol (PG), and glycerol (GLY), and two media, namely Dulbecco''s modified Eagle medium Ham’s F-12 (DMEM)and K+-modified TiProtec (K+TiP), which is a high-potassium-containing medium. Numbers of viable cells in proliferation were evaluated by the CellTiter 96® AQueous One Solution Cell Proliferation Assay (Promega Corporation, Mannheim, Germany). To detect the exact frozen cell number per cryo vial, DNA content was measured by using Hoechst 33258 dye prior to analysis. Thus, results could be evaluated unconstrained by absolute cell number. Thawed cells were cultured in 25 cm2 cell culture flasks to confluence and examined daily by phase contrast imaging. With regard to cell recovery immediately after thawing, DMSO was the most suitable CPA combined with K+TiP in vitrification (99 ±0.5%) and with DMEM in slow freezing (92 ±1.6%). The most viable cells in proliferation after three days of culture were obtained in cells vitrificated by using GLY with K+TiP (308 ±34%) and PG with DMEM in slow freezing (280 ±27%). 相似文献
33.
Strunecký Otakar Kopejtka Karel Goecke Franz Tomasch Jürgen Lukavský Jaromír Neori Amir Kahl Silke Pieper Dietmar H. Pilarski Plamen Kaftan David Koblížek Michal 《Extremophiles : life under extreme conditions》2019,23(1):35-48
Extremophiles - Genotypic and morphological diversity of cyanobacteria in the Rupite hot spring (Bulgaria) was investigated by means of optical microscopy, cultivation, single-cell PCR, and 16S... 相似文献
34.
Saranna Fanning Aftabul Haque Thibaut Imberdis Valeriya Baru M. Inmaculada Barrasa Silke Nuber Daniel Termine Nagendran Ramalingam Gary P.H. Ho Tallie Noble Jackson Sandoe Yali Lou Dirk Landgraf Yelena Freyzon Gregory Newby Frank Soldner Elizabeth Terry-Kantor Tae-Eun Kim Dennis Selkoe 《Molecular cell》2019,73(5):1001-1014.e8
35.
Jezierska Sylwia Claus Silke Ledesma-Amaro Rodrigo Van Bogaert Inge 《Journal of industrial microbiology & biotechnology》2019,46(12):1697-1706
Journal of Industrial Microbiology & Biotechnology - Free fatty acids are basic oleochemicals implemented in a range of applications including surfactants, lubricants, paints, plastics, and... 相似文献
36.
Evaluation of a DC pulsed magnetic tracking system in neurosurgical navigation: technique, accuracies, and influencing factors] 总被引:1,自引:0,他引:1
Olaf Suess Silke Suess Sven Mularski Bj?rn Kühn Thomas Picht Sven Sch?nherr Theodoros Kombos 《Biomedizinische Technik》2007,52(3):223-233
Navigation systems are useful instruments in cranial neurosurgery. For specification of position, so-called sensor-based navigation techniques use: (a) a signal emitter that generates a defined electromagnetic field in the area of the operation site; and (b) small sensors that detect the position of various operating instruments in the electromagnetic field. For a long time, owing to a lack of clinical data and long-term studies, electromagnetic systems have been regarded as error-prone and imprecise. With the development of a pulsed direct current (DC) technique, precision levels can now be reached that are comparable with those of established optical and mechanical measuring procedures. However, it must be noted that the influence on the measuring accuracy within the operating field increases with increasing susceptibility of the various metals used in the operating theatre (titanium相似文献
37.
Pauff JM Hemann CF Jünemann N Leimkühler S Hille R 《The Journal of biological chemistry》2007,282(17):12785-12790
The rapid reaction kinetics of wild-type xanthine dehydrogenase from Rhodobacter capsulatus and variants at Arg-310 in the active site have been characterized for a variety of purine substrates. With xanthine as substrate, k(red) (the limiting rate of enzyme reduction by substrate at high [S]) decreased approximately 20-fold in an R310K variant and 2 x 10(4)-fold in an R310M variant. Although Arg-310 lies on the opposite end of the substrate from the C-8 position that becomes hydroxylated, its interaction with substrate still contributed approximately 4.5 kcal/mol toward transition state stabilization. The other purines examined fell into two distinct groups: members of the first were effectively hydroxylated by the wild-type enzyme but were strongly affected by the exchange of Arg-310 to methionine (with a reduction in k(red) greater than 10(3)), whereas members of the second were much less effectively hydroxylated by wild-type enzyme but also much less significantly affected by the amino acid exchanges (with a reduction in k(red) less than 50-fold). The effect was such that the 4000-fold range in k(red) seen with wild-type enzyme was reduced to a mere 4-fold in the R310M variant. The data are consistent with a model in which "good" substrates are bound "correctly" in the active site in an orientation that allows Arg-310 to stabilize the transition state for the first step of the overall reaction via an electrostatic interaction at the C-6 position, thereby accelerating the reaction rate. On the other hand, "poor" substrates bound upside down relative to this "correct" orientation. In so doing, they are unable to avail themselves of the additional catalytic power provided by Arg-310 in wild-type enzyme but, for this reason, are significantly less affected by mutations at this position. The kinetic data thus provide a picture of the specific manner in which the physiological substrate xanthine is oriented in the active site relative to Arg-310 and how this residue is used catalytically to accelerate the reaction rate (rather than simply bind substrate) despite being remote from the position that is hydroxylated. 相似文献
38.
The large archives of formalin-fixed paraffin-embedded (FFPE) tissue specimens that exist are a highly valuable source of sample material for molecular biological analysis, including gene expression profiling. However, current data on adverse effects of standard pathological practice on the usefulness of biomolecular analytes obtained from such archived specimens is largely anecdotal. Here, we present a systematic examination of the most relevant parameters for integrity and useability of RNA obtained from FFPE samples, including storage time and conditions, fixation time, and specimen size. The results are particularly relevant for any application relying on cDNA synthesis as an initial step of the procedure, such as RT-PCR, and microarray analysis. 相似文献
39.
Objective: The study of human preadipocytes is hampered by the limited availability of adipose tissue and low yield of cell preparation. Proliferation of preadipocytes using common protocols, including fetal bovine serum (FBS), results in a markedly reduced differentiation capacity. Therefore, we were interested in developing an improved culture system that allows the proliferation of human preadipocytes without loss of differentiation capacity. Research Methods and Procedures: Adipose tissue samples were taken from subjects undergoing elective abdominal surgery. Cells were seeded at various densities and cultured using different formulations of proliferation media including factors such as fibroblast growth factor‐2 (basic fibroblast growth factor), epidermal growth factor, insulin, and FBS either alone or in combination. Cells were counted and induced to differentiate after confluence. After complete differentiation, cells were harvested, and glycerol‐3‐phosphate dehydrogenase activity was measured. Cells were subcultured for up to five passages. Results: The proliferation medium with 4 growth factors (PM4), consisting of 2.5% FBS, 10 ng/mL epidermal growth factor, 1 ng/mL basic fibroblast growth factor, and 8.7 µM insulin, resulted in lower doubling times at all seeding densities tested (0.05 × 104 to 1.5 × 104) compared with medium supplemented with 10% FBS. In contrast to cells in FBS medium, cells grown with PM4 medium retained full differentiation rate (glycerol‐3‐phosphate dehydrogenase activity, 493 ± 215 vs. 41 ± 17 mU/mg, p < 0.01). Differentiation capacity was fully retained at least for up to three passages in PM4 medium. Discussion: The use of PM4 medium results in substantial proliferation of human preadipocytes with preserved differentiation capacity. This novel technique represents a valuable tool for the study of human adipose tissue development and function starting from small samples. 相似文献
40.
Sequence alignments of human molybdopterin synthase sulfurase, MOCS3, showed that the N-terminal domain is homologous to Escherichia coli MoeB, whereas the C-terminal domain is homologous to rhodanese-like proteins. Previous studies showed that the activity of the separately purified rhodanese-like domain of MOCS3 displayed 1000-fold lower activity in comparison to bovine rhodanese with thiosulfate as sulfur source. When the six amino acid active site loop of MOCS3 rhodanese-like domain was exchanged with the loop found in bovine rhodanese, thiosulfate:cyanide sulfurtransferase activity was increased 165-fold. Site-directed mutagenesis of each individual residue of the active site loop of the MOCS3 rhodanese-like domain showed that the charge of the last amino acid determines thiosulfate sulfurtransferase activity. Replacing Asp417 by threonine resulted in 90-fold increased activity, whereas replacing it by arginine increased the activity 470-fold. Using a fully defined in vitro system containing precursor Z, MOCS2A, E. coli MoaE, E. coli MoeB, Mg-ATP, MOCS3 rhodanese-like domain, and thiosulfate, it was shown that sulfur transfer to MOCS2A was also affected by the alterations, but not as drastically. Our studies revealed that in humans and most eukaryotes thiosulfate is not the physiologic sulfur donor for MOCS3, whereas in bacterial homologs, which have an arginine at the last position of the active site loop, thiosulfate can be used as a sulfur source for molybdenum cofactor biosynthesis. The phylogenetic analysis of MoeB homologs showed that eukaryotic homologs are of bacterial origin. Furthermore, it could be shown that an MoeB homolog named MoeZ, where the dual CXXC zinc-binding motif of the MoeB domain is not present, arose independently several times during evolution. 相似文献