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881.
Marco Moracci Maria Ciaramella Roberto Nucci Laurence H. Pearl Ian Sanderson Antonio Trincone Mos Rossi 《Biocatalysis and Biotransformation》1994,11(2):89-103
The Sulfolobus solfataricus β-glycosidase (Sβgly) is a thermostable and thermophilic glycosyl-hydrolase with broad substrate specificity. The enzyme hydrolizes β-D-gluco-, fuco-, and galactosides, and a large number of /Winked glycoside dimers and oligomers, linked β1-3, β1-4, and β1-6, It is able to hydrolize oligosaccharides with up to 5 glucose residues. Furthermore, it is also able to promote transglycosylation reactions. The corresponding gene has been cloned and overexpressed both in yeast and Escherichia coli. Based on sequence and functional data, the Sβgly has been assigned to the so-called BGA family of glycosyl-hydrolases, including β-glycosidases, β-galactosidases and phosho-β-galactosidases from mesophilic and thermophilic organisms of the three domains. The Sβgly has been crystallized and the resolution of its structure is in progress. Because of its special properties, the enzymes has considerable biotechnological potential. 相似文献
882.
Giovanna Berruti 《Molecular reproduction and development》1994,38(4):386-392
The majority of cellular responses to changing environmental conditions is regulated by protein kinases. Spermatozoa have many special properties, including motility with demonstrated chemotaxis, the ability to undergo capacitation, and the acrosome reaction, which are in part controlled by extracellular signals and in which sperm kinases are considered to be involved. We have previously reported that there is a protein kinase activity, which phosphorylates the synthetic substrate poly-(Glu, Tyr) with a Km value of 2.3 μM, and is inhibited by the tyrosine kinase inhibitor tyrphostin, in the protein extract from boar spermatozoa (Berruti and Porzio, 1992: Biochim Biophys Acta 1118:149–154). Now we have demonstrated that the enzyme is cytosolic, is active as a monomer of Mr 42,000, is stimulated by Mg2+ > Mn2+ but not by Ca2+, is renaturable, and can phosphorylate native protein substrates such as microtubule-associated protein 2 (MAP2) and histone H2B both on the tyrosine and serine residues. N-terminal sequence analysis suggests that it is a novel protein. These new findings imply that the boar sperm 42 kD kinase may be a novel member of the emerging class of dual-specificity protein kinases, and they raise the intriguing question of its function in the protein kinase network mediating signal transduction in mammalian spermatozoa. © 1994 Wiley-Liss, Inc. 相似文献
883.
Methenamine-Silver Staining: a Simple and Sensitive Staining Method for Senile Plaques and Neurofibrillary Tangles 总被引:1,自引:0,他引:1
Chie Haga Kenji Ikeda Kiyoshi Iwabuchi Haruhiko Akiyama Hiromi Kondoh Kenji Kosaka 《Biotechnic & histochemistry》1994,69(5):295-300
An improved methenamine-silver impregnation method is presented which exhibits sensitivity for amyloid substances comparable to that of anti-β protein immunostaining. In optimally treated sections, this technique stained both β-amyloid deposits and neurofibrillary tangles, which are known to have a β-pleated structure. This simple procedure allows a large number of sections to be stained for routine examination. 相似文献
884.
Einar Lystad Arne T. Høstmark Cecilie Kiserud Aage Haugen 《In vitro cellular & developmental biology. Animal》1994,30(9):568-573
Summary The protective influence of bovine serum albumin against growth inhibition caused by fatty acids was studied in human hepatoma
(HepG2) and immortalized human kidney epithelial (IHKE) cells. In general, growth inhibition by unsaturated fatty acids (0.15
mmol/liter) increased with increasing number of double bonds. For HepG2 cells crude albumin (1g/100 ml) did not greatly modify growth inhibition by arachidonic, eicosapentaenoic, and docosahexaenoic acid. With oleic,
linoleic, and linolenic acids, crude and defatted albumin stimulated cell growth. In contrast, for IHKE cells both albumins
counteracted growth inhibition by unsaturated fatty acids to approximately the same extent. When HepG2 cells were cultured
in the presence of saturated fatty acids (0.3 mmol/liter), C2, C6, and C8 had no or little inhibitory effect. C10 and C12
inhibited cell growth appreciably, whereas C14, and especially C16, had poor inhibitory effects. Crude albumin counteracted
growth inhibition by all these fatty acids. In contrast, defatted albumin had little or no effect (except against C10 and
C12), and even increased the growth inhibition by C14 and C16. With unsaturated fatty acids there seemed to be an inverse
relationship between cell growth and the concentration of thiobarbituric acid reactive substances (TBARS) in media. Vitamin
E abolished growth inhibition (and the increase in TBARS concentration) by unsaturated fatty acids. The complex interaction
between fatty acids and albumins calls for great caution when interpreting data on growth effects. 相似文献
885.
Catherine Jumarie Christiane Malo 《In vitro cellular & developmental biology. Animal》1994,30(11):753-760
Summary Caco-2 cell human colon adenocarcinoma cell line was used to study the hormonal regulation of small intestinal epithelial
cell differentiation. We had previously shown that insulin-transferrin-selenium and triiodothyronine (5 × 10−8
M)-supplemented medium can best replace serum after 2 days of culture for both the maintenance and differentiation of Caco-2
cells. The present study demonstrates that precoating petri dishes with complete serum allows the growth and differentiation
of Caco-2 cells seeded directly in serum-free medium. On the other hand, precoating with dialyzed serum inhibits alkaline
phosphatase and dipeptidyl-dipeptidase IV activities by more than 50%. The results obtained with complete serum-precoated
culture plates indicate that there is no synergy between insulin and triiodothyronine because cells maintained in transferrin-selenium
and triiodothyronine-supplemented medium, with or without insulin, express comparable enzyme activities. Moreover, large increases
in alkaline phosphatase and dipeptidyl-dipeptidase IV activities were observed when triiodothyronine was added to the culture
medium by the time confluency was reached. In contrast, γ-glutamyltransferase was lowered to a greater extent when triiodothyronine
was present from the beginning of culture. These findings show that triiodothyronine preferentially stimulates alkaline phosphatase
and dipeptidyl-dipeptidase IV activities during the differentiation period whereas it selectively inhibits γ-glutamyltransferase
during the proliferation phase. Triiodothyronine acts in a dose-dependent manner. 相似文献
886.
Wilfried Bautsch 《Molecular biotechnology》1994,2(1):29-44
Macrorestriction mapping is often the first step toward a thorough physical and genetic characterization of a bacterial genome.
The problem of deducing the order of partially or completely digested macrorestriction fragments to yield a physical genome
map may readily be solved by applying twodimensional pulsed-field gel electrophoresis (2D-PFGE) techniques. These powerful
methods are quick and technically easy to perform; specifically, they are independent of DNA probes and should therefore be
applicable to any bacterial species irrespective of its prior genetic characterization. In this article, detailed step-by-step
protocols are given to set up, run, and evaluate 2D pulsed-field gels. Two basic methods are described: partial/complete 2D
gels of one restriction enzyme and complete/complete 2D gels of two different restriction enzymes. Other topics include preparation
of bacterial genomic DNA, screening for suitable rare-cutting restriction enzymes and determination of optimal running conditions.
Accompanied by many notes, these protocols are meant to offer the novice a sound and rapid access to these important methods. 相似文献
887.
Gas chromatography-mass spectrometry has been applied to the analysis of plasma linoleic acid and one of its oxidation products, 4-hydroxy-2-nonenal (HNE), in adult patients with the acute respiratory distress syndrome (ARDS). Peak areas of total ion chromatograms showed there to be negative correlations between loss of linoleic acid and formation of HNE (measured by selective ion monitoring) in 7 out 10 patients studied. When HNE was quantitated by selective ion monitoring, with reference to a pure standard of HNE and an internal standard of nonanoic acid, ARDS patients showed significantly increased levels of HNE (0.412 ± 0.023 nmol/ml) compared with normal healthy controls (0.205 ± 0.018 nmol/ml). 相似文献
888.
William R. Mundy Prasada Rao S. Kodavanti Victoria F. Dulchinos Hugh A. Tilson 《Journal of biochemical and molecular toxicology》1994,9(1):17-23
Calcium is actively transported into intracellular organelles and out of the cytoplasm by Ca2+/Mg2+-ATPases located in the endoplasmic reticulum and plasma membranes. We studied the effects of aluminum on calcium transport in the adult rat brain. We examined 45Ca-uptake in microsomes and Ca2+-ATPase activity in microsomes and synaptosomes isolated from the frontal cortex and cerebellum of adult male Long-Evans rats. ATP-dependent45Ca-uptake was similar in microsomes from both brain regions. The addition of 50-800 μM AICI3 resulted in a concentration-dependent inhibition of 45Ca-uptake. Mg2+-dependent Ca2+-ATPase activity was significantly lower in synaptosomes compared to microsomes in both frontal cortex and cerebellum. In contrast to the uptake studies, AICI3 stimulated Mg2+-dependent Ca2+-ATPase activity in both microsomes and synaptosomes from both brain regions. To determine the relationship between aluminum and Mg2+, we measured ATPase activity in the presence of increasing concentrations of Mg2+ or AICI3. Maximal ATPase activity was obtained between 3 and 6 mM Mg2+. When we substituted AICI3 for Mg2+, ATPase activity was also stimulated in a concentration-dependent manner, but to a greater extent than with Mg2+. One interpretation of these data is that aluminum acts at multiple sites to displace both Mg2+ and Ca2+, increasing the activity of the Ca2+-ATPase, but disrupting transport of calcium. 相似文献
889.
Xiao-Chen Huang 《International journal of biometeorology》1993,37(4):222-228
The febrile response and sympathetic nervous response to hypothalamic microinjections of prostaglandin E2 (PGE2) were investigated in anesthetized rabbits. Microninjection of PGE2 (500–1000 ng) caused an increase in rectal temperature of more than 0.3°C in 13 of 50 loci in the preoptic and anterior hypothalamic area (PO/AH). At 8 of these 13 loci, PGE2 elicited response patterns in the sympathetic nervous system, such as an increase in cutaneous sympathetic nervous activity and decrease in renal sympathetic nervous activity. This pattern of sympathetic nervous responses was induced with a simultaneous increase in rectal temperature of more than 0.5°C. The 8 loci were distributed in the preoptic area, especially in the vicinity of the supraoptic nucleus. Electrolytic lesions of this region were made bilaterally, and intracerebroventricular injection of PGE2 (8 µg/kg) was found to inhibit fever and sympathetic activity. The results demonstrate that the action of PGE2 is responsible for the response patterns of sympathetic twigs during fever. The preoptic area, especially in the vicinity of the supraoptic nucleus, is most sensitive to PGE2 for the patternized response of sympathetic neurons and fever. 相似文献
890.
I. I. Pottosin P. R. Andjus D. Vučelić G. N. Berestovsky 《The Journal of membrane biology》1993,136(2):113-124
We studied the effects of H2O/D2O substitution on the permeation and gating of the large conductance Ca2+-activated K+ channels inChara gymnophylla droplet membrane using the patchclamp technique. The selectivity sequence of the channel was: K+>Rb+≫Li+, Na+, Cs+ and Cl−. The conductance of this channel in symmetric 100mm KCl was found to be 130 pS. The single channel conductance was decreased by 15% in D2O as compared to H2O. The blockade of channel conductance by cytosolic Ca2+ weakened in D2O as a result of a decrease in zero voltage Ca2+ binding affinity by a factor of 1.4. Voltage-dependent channel gating was affected by D2O primarily due to the change in Ca2+ binding to the channel during the activation step. The Hill coefficient for Ca2+ binding was 3 in D2O and around 1 in H2O. The values of the Ca2+ binding constant in the open channel conformation were 0.6 and 6 μm in H2O and D2O, respectively, while the binding in the closed conformation was much less affected by D2O. The H2O/D2O substitution did not produce a significant change in the slope of channel voltage dependence but caused a shift as large
as 60 mV with 1mm internal Ca2+. 相似文献