全文获取类型
收费全文 | 3274篇 |
免费 | 123篇 |
国内免费 | 39篇 |
专业分类
3436篇 |
出版年
2023年 | 53篇 |
2022年 | 66篇 |
2021年 | 84篇 |
2020年 | 74篇 |
2019年 | 162篇 |
2018年 | 155篇 |
2017年 | 106篇 |
2016年 | 74篇 |
2015年 | 54篇 |
2014年 | 140篇 |
2013年 | 138篇 |
2012年 | 78篇 |
2011年 | 105篇 |
2010年 | 60篇 |
2009年 | 65篇 |
2008年 | 79篇 |
2007年 | 91篇 |
2006年 | 85篇 |
2005年 | 96篇 |
2004年 | 110篇 |
2003年 | 112篇 |
2002年 | 88篇 |
2001年 | 47篇 |
2000年 | 40篇 |
1999年 | 50篇 |
1998年 | 43篇 |
1997年 | 49篇 |
1996年 | 48篇 |
1995年 | 41篇 |
1994年 | 58篇 |
1993年 | 49篇 |
1992年 | 49篇 |
1991年 | 44篇 |
1990年 | 28篇 |
1989年 | 42篇 |
1988年 | 41篇 |
1987年 | 34篇 |
1986年 | 28篇 |
1985年 | 73篇 |
1984年 | 102篇 |
1983年 | 52篇 |
1982年 | 101篇 |
1981年 | 61篇 |
1980年 | 70篇 |
1979年 | 28篇 |
1978年 | 35篇 |
1977年 | 30篇 |
1976年 | 37篇 |
1974年 | 31篇 |
1973年 | 21篇 |
排序方式: 共有3436条查询结果,搜索用时 15 毫秒
71.
Evaluation of monitoring approaches and effects of culture conditions on recombinant protein production in baculovirus-infected insect cells 总被引:2,自引:0,他引:2
The baculovirus infection process ofSpodoptera frugiperda (Sf9) insect cells in oxygen-controlled bioreactors in serum-free medium was investigated using a recombinantAutographa californica (AcNPV) virus expressing -galactosidase enzyme as a model system. A variety of monitoring techniques including trypan blue exclusion, fluorescent dye staining, oxygen uptake rate (OUR) measurements, and glucose consumption were applied to infected cells to determine the best way of evaluating cell integrity and assessing the course of baculovirus infection. The metabolism of newly-infected cells increased 90% during the first 24 hours, but as infection proceeded, and cells gradually succumbed to the baculovirus infection, the cytopathic effect of the baculovirus on the cells became evident. Oxygen and glucose uptake rate measurements appeared to more accurately assess the condition of infected cells than conventional trypan blue staining, which tended to overestimate cell viability in the mid stages of infection. The optimal harvest time varied, depending on which technique — SDS-PAGE, chromogenic (ONPG) or fluorometric (C12FDG) — was used to monitor -galactosidase production. Specific -galactosidase production was found to be insensitive to a wide range of culture dissolved oxygen tensions, whereas resuspending cells in fresh medium prior to infection increased volumetric productivity approximately two-fold (800,000 units -galactosidase/ml) compared to cultures infected in batch mode and allowed successful infections to occur at higher cell densities.Abbreviations ONPG
ortho-phenyl 2--D-galactopyranoside
- OUR
oxygen uptake rate (-mol O2/liter/hour)
- qglucose
specific glucose uptake rate (mg glucose/106cell/hour)
- qglutamine
specific glutamine uptake rate (mg glutamine/106cell/hour)
- qO2
specific oxygen uptake rate (-mol O2/106cell/hour)
- MOI
virus multiplicity of infection (viral plaque forming units/cell) 相似文献
72.
73.
The expression of an Arabidopsis acyl carrier protein (ACP) gene promoter has been examined in transgenic tobacco plants by linking it to the reporter gene -glucuronidase (GUS). Fluorometric analysis showed that the ACP gene promoter was most active in developing seeds. Expression was also high in roots, but significantly lower in young leaves and downregulated upon their maturation. Etiolated and light-grown seedlings showed the same level of GUS activity, indicating that this promoter is not tightly regulated by light. Histochemical studies revealed that expression was usually highest in apical/ meristematic zones of vegetative tissues. Young flowers (ca. 1 cm in length) showed GUS staining in nearly all cell types, however, cell-specific patterns emerged in more mature flowers. The ACP gene promoter was active in the stigma and transmitting tissue of the style, as well as in the tapetum of the anther, developing pollen, and ovules. The results provide evidence that this ACP gene is regulated in a complex manner and is responsive to the array of signals which accompany cell differentiation, and a demand for fatty acids and lipids, during organogenesis. 相似文献
74.
Polyclonal antiserum raised against the native PG1 isoform of tomato fruit (Lycopersicon esculentum Mill.) polygalacturonase [poly(1,4--d-galacturonide) glycanohydrolase, EC 3.2.1.15] bound to each of the subunits of the protein and also to a range of other fruit proteins. Affinity purification was used to remove antibody molecules that bound to the native form of the PG2 isoform. The resulting serum bound to native PG1, denatured PG2 and -subunits of PG1 but not to native PG2 or other fruit proteins. This anti-PG1 serum was used to monitor the occurrence of the PG1 -subunit and PG2 in detergent extracts of tomato tissues. The -subunit polypeptide was detected in pericarp but not locule tissue of fruit, including fruit of the rin and nor mutants. It increased in amount in the pericarp tissues from an early stage to the mature green stage, clearly prior to any appreciable accumulation of the PG2 subunit. The -subunit polypeptide was not detected in stem or leaf tissues. A PG2-specific antiserum was used to study the interaction of PG2 with the isolated -subunit. The PG2 isoform was bound to the -subunit over a wide range of salt concentrations and pH; the interaction was independent of the presence of reducing agents. It is concluded that strong non-covalent forces are involved in the interaction. The results are consistent with a model in which the -subunit is positioned in the cell wall structure and provides a specific binding site for the active PG2 subunit when this is synthesised during ripening.Abbreviations B
breaker
- MG
mature green
- Mr
relative molecular mass
-
nor
non-ripening mutant
- PAGE
polyacrylamide gel electrophoresis
- PG
polygalacturonase
-
rin
ripening inhibitor mutant
- SDS
sodium dodecyl sulphate 相似文献
75.
Energy-using non-mitochondrial ATPases were assayed in rat cerebral cortex synaptosomes and synaptosomal subfractions, namely synaptosomal plasma membranes and synaptic vesicles. The following enzyme activities were evaluated: Na+, K+-ATPase; high- and low-affinity Ca2+-ATPase; basal Mg2+-ATPase; Ca2+, Mg2+-ATPase. The evaluations were performed after four week-treatment with saline [controls] or -adrenergic agents (-yohimbine, clonidine), energymetabolism interfering compound (theniloxazine), and oxygen-partial pressure increasing agent (almitrine), in order to define the plasticity and the selective changes in individual ATPases. In rat cerebral cortex, the enzyme adaptation to four-week-treatment with -yohimbine or clonidine was characterized by increase in both high- and low-affinity Ca2+-ATPase activities. The action involves the enzyme form located in the synaptic plasma membranes. The enzyme adaptation to the subchronic treatments with theniloxazine or almitrine was characterized by increase in Na+, K+-ATPase or Mg2+-ATPase activities, respectively. The action involves the enzymatic forms located in the synaptic plasma membranes. Thus, the pharmacodynamic effects of the agents tested should also be related to the changes induced in the activity of some specific synaptosomal nonmitochondrial ATPases. 相似文献
76.
Richard Darley Alan Morris James Passas Wendy Bateman 《Cancer immunology, immunotherapy : CII》1993,37(2):112-118
The cell-surface expression of major histocompatibility (MHC) antigens and the adhesion molecule intercellular adhesion molecule 1 (ICAM-1) is essential for target cell recognition by T lymphocytes. The expression of both classes of molecule is induced by various cytokines, notably interferon (IFN). Since transforming growth factor (TGF) has been recently reported to antagonise HLA-DR induction by IFN we have examined, using a number of murine and human cell lines, the effect of TGF on IFN-induced MHC class I and class II and ICAM-1 expression. All of the cell lines tested expressed elevated class I MHC following IFN treatment. Class II MHC induction was seen on most but not all of the cells, the exceptions being among a panel of human colorectal carcinoma cell lines. A striking difference between cells of different origin was noted in the response to TGF. TGF was found to antagonise IFN-induced class I and class II MHC expression on C3H 10T1/2 murine fibroblasts, early-passage BALB/c mouse embryo fibroblasts, a murine oligodendroglioma cell line, and on MRC5 human fibroblasts and two human glioblastoma cell lines. Class II MHC was much more strongly inhibited (sometimes completely) than class I MHC. TGF also inhibited induction of class I MHC expression by IFN. However, TGF did not inhibit class I or class II MHC induction by IFN in any of the nine colorectal carcinoma cell lines, although two of five of the lines tested were growth-inhibited by TGF. On the other hand, human ICAM-1 induction by IFN was not affected by simultaneous treatment with TGF in any of the cell lines. The down-regulation of IFN-induced MHC antigens by TGF is not, therefore, the result of a general antagonism of IFN. Retinoic acid has recently been reported to induce ICAM-1 expression on human tumour cells. We have confirmed this observation on MRC5, and the two human glioblastoma cell lines, however six colorectal carcinoma cell lines tested did not respond. In contrast to IFN-induced ICAM-1 expression, retinoic-acid-induced ICAM-1 expression was inhibited by TGF on two of the three responsive lines. 相似文献
77.
78.
《L'Anthropologie》2023,127(2):103134
The most abundant evidence of Pampean Pleistocene human presence are modified bones, as lithic procurement sites located farther than 300 km away. Therefore, we focused on the study of bone modifications, in particular cut and percussion marks. We studied Pampean paleontological collections of Argentine and European museums as an alternative resource of previously unnoticed human modification evidences. We compared marks characteristics with those of archaeological collections from diverse climatic and cultural adaptations, from middle Pleistocene sites (Vallonet, Atapuerca, Lazaret, Arago and Terra Amata) to terminal Pleistocene ones (Abri Pataud, Isturiz and La Vache). Marks typologies were defined, leading to the identification of a particular cutmark that we named double parallel considered as highly diagnostic of anthropic use of lithic artifacts. We also propose that hafted artifacts were used on carcass processing. Perimortem bone modifications are classified in relation with mark morphology (e.g., percussion striae), inferred gesture or action (e.g., breakage of diaphysis), and inferred objective of that action (e.g., marrow extraction). The megafauna specimens analyzed from the historical collections (d’Angelis-Vilardebó, 1847, Muñiz-Dupotet, 1842 and Breton-Bonnement, 1881) present modifications compatible with dismemberment, defleshing, tongue and masseter extraction, and utilization as anvils. The lack of contextual data (location, stratigraphy) and the loss due to museum selection/collecting of accompanying material prevent paleoenvironmental and paleoecological inferences. Regional geology indicates that most of the historical collections were exhumed in riverbank cuts with ages between 70 and 13 ky BP. Preliminary direct dating presents evidence of a Pampean human occupation, at least, since OIS 2. 相似文献
79.
《Revista espa?ola de geriatría y gerontología》2023,58(3):148-154
Introduction and objectivesThere is a considerable lack of structure in training health professionals in geriatrics. The narratives can promote a collaborative reflection on different topics and might be used as a pedagogic strategy for undergraduate health students. This study aimed to explore the adoption of new perspectives on ageing after the implementation of dynamic narratives in the first graduate year for physiotherapy students.Materials and methodsAn exploratory qualitative study was performed. Participants were included if they were: ≥18 years old, physiotherapy students and agreed to participate. Physiotherapy students (n = 44) were recruited from the School of Health Sciences, Polytechnic Institute of Leiria. Two gaming sessions were performed to help students, as narrators, express their visions and ways to deal with the geriatrics field. Students’ perspectives about ageing at baseline (T1) and after the narratives experience (T2) were collected by answering the following question: What is your perception regarding ageing? Two evaluators were involved in qualitative data analysis by performing: (i) an individual analysis of themes/subthemes and a discussion meeting to evaluate discrepancies and to reach a consensus.ResultsNegative perceptions about ageing were referred 39 times at T1 (most of them related to restriction; deterioration subthemes). There were no negative perceptions registered at T2. Positive perceptions increased at T2, from n = 39 to n = 52, and three new subthemes emerged (beginning of something, fighting ageism, challenge).ConclusionThis study demonstrated the potential of narrative-based experiences as a desirable pedagogic methodology (board games-oriented) for geriatric education in undergraduate health students. 相似文献
80.
《Revista espa?ola de geriatría y gerontología》2023,58(3):125-133
IntroductionBNT162b2 (BioNTech and Pfizer) is a nucleoside-modified mRNA vaccine that provides protection against SARS-CoV-2 infection and is generally well tolerated. However, data about its efficacy, immunogenicity and safety in people of old age or with underlying chronic conditions are scarce.PurposeTo describe BNT162b2 (BioNTech and Pfizer) COVID-19 vaccine immunogenicity, effectiveness and reactogenicity after complete vaccination (two doses), and immunogenicity and reactogenicity after one booster, in elders residing in nursing homes (NH) and healthy NH workers in real-life conditions.MethodsObservational, ambispective, multicenter study. Older adults and health workers were recruited from three nursing homes of a private hospital corporation located in three Spanish cities. The primary vaccination was carried out between January and March 2021. The follow-up was 13 months. Humoral immunity, adverse events, SARS-CoV-2 infections, hospitalizations and deaths were evaluated. Cellular immunity was assessed in a participant subset.ResultsA total of 181 residents (mean age 84.1 years; 89.9% females, Charlson index ≥2: 45%) and 148 members of staff (mean age 45.2 years; 70.2% females) were surveyed (n:329). After primary vaccination of 327 participants, vaccine response in both groups was similar; ≈70% of participants, regardless of the group, had an antibody titer above the cut-off considered currently protective (260 BAU/ml). This proportion increased significantly to ≈ 98% after the booster (p < 0.0001 in both groups). Immunogenicity was largely determined by a prior history of COVID-19 infection. Twenty residents and 3 workers were tested for cellular immunity. There was evidence of cellular immunity after primary vaccination and after booster. During the study, one resident was hospitalized for SARS-CoV-2. No SARS-CoV-2-related deaths were reported and most adverse events were mild.ConclusionsOur results suggest that the BNT162b2 mRNA COVID-19 vaccine is immunogenic, effective and safe in elderly NH residents with underlying chronic conditions. 相似文献