全文获取类型
收费全文 | 1647篇 |
免费 | 96篇 |
国内免费 | 1篇 |
出版年
2023年 | 12篇 |
2022年 | 8篇 |
2021年 | 32篇 |
2020年 | 29篇 |
2019年 | 26篇 |
2018年 | 48篇 |
2017年 | 43篇 |
2016年 | 65篇 |
2015年 | 100篇 |
2014年 | 81篇 |
2013年 | 119篇 |
2012年 | 155篇 |
2011年 | 123篇 |
2010年 | 70篇 |
2009年 | 60篇 |
2008年 | 109篇 |
2007年 | 67篇 |
2006年 | 78篇 |
2005年 | 71篇 |
2004年 | 48篇 |
2003年 | 62篇 |
2002年 | 44篇 |
2001年 | 18篇 |
2000年 | 15篇 |
1999年 | 14篇 |
1998年 | 9篇 |
1996年 | 8篇 |
1995年 | 6篇 |
1992年 | 8篇 |
1991年 | 7篇 |
1990年 | 10篇 |
1989年 | 14篇 |
1988年 | 8篇 |
1987年 | 8篇 |
1985年 | 12篇 |
1981年 | 5篇 |
1978年 | 5篇 |
1977年 | 6篇 |
1976年 | 5篇 |
1975年 | 6篇 |
1974年 | 12篇 |
1973年 | 20篇 |
1972年 | 9篇 |
1971年 | 10篇 |
1970年 | 11篇 |
1969年 | 6篇 |
1968年 | 9篇 |
1967年 | 6篇 |
1966年 | 9篇 |
1965年 | 9篇 |
排序方式: 共有1744条查询结果,搜索用时 19 毫秒
101.
Rocio Duque‐Jamaica Azucena Arévalo‐Galvis Raúl A. Poutou‐Piñales Alba A. Trespalacios 《Helicobacter》2010,15(4):303-312
Background: Colonization of the gastric mucosa by Helicobacter pylori is one of the most important causes of acute and chronic gastric pathologies in humans. Achieving the growth of H. pylori in liquid media is of great importance in the development of clinical studies. In this study, we developed a sequential optimization strategy based on statistical models to improve the conditions of liquid culture of H. pylori. Materials and Methods: Four statistical models were sequentially used. First, a Box‐Behnken design was used to select the best process conditions (shaking speed, inoculum concentration, and final volume of culture). Secondly, a general factorial design was used to evaluate the influence of adding gel blocks or gel beads (shape and composition). Then a D‐optimal reduce design was carried out to allow the selection of the most influential factors in increasing the cell concentration (culture media components). Finally, another Box‐Behnken design was used to optimize the concentration of the culture media components previously selected. Results: After 12 hours of liquid culture a concentration of 25 × 108 cells per mL (9.4 log10 cells per mL) of H. pylori was obtained, compared with a predicted 32 × 108 (9.5 log10 cells per mL), which means between 1 and 5 log10 units higher than some previous reports. Conclusions: The sequential statistical approach increased the planktonic H. pylori cell culture. The final culture media and conditions were: Brain Heart Infusion, blood agarose (1.5% w/v), lamb’s blood (3.18% v/v), DENT (0.11% v/v), and Vitox (0.52% v/v) at 60 rpm and 37 °C with filtered CO2 (5% v/v) bubbled directly into the culture media in a final volume of 76.22 mL. 相似文献
102.
103.
104.
105.
106.
Marek Rác Michal K?upka Svatopluk Binder Michaela Sedlá?ová Zuzana Matu?ková Milan Ra?ka Pavel Pospí?il 《PloS one》2015,10(3)
The exposure of human cells to oxidative stress leads to the oxidation of biomolecules such as lipids, proteins and nuclei acids. In this study, the oxidation of lipids, proteins and DNA was studied after the addition of hydrogen peroxide and Fenton reagent to cell suspension containing human leukemic monocyte lymphoma cell line U937. EPR spin-trapping data showed that the addition of hydrogen peroxide to the cell suspension formed hydroxyl radical via Fenton reaction mediated by endogenous metals. The malondialdehyde HPLC analysis showed no lipid peroxidation after the addition of hydrogen peroxide, whereas the Fenton reagent caused significant lipid peroxidation. The formation of protein carbonyls monitored by dot blot immunoassay and the DNA fragmentation measured by comet assay occurred after the addition of both hydrogen peroxide and Fenton reagent. Oxidative damage of biomolecules leads to the formation of singlet oxygen as conformed by EPR spin-trapping spectroscopy and the green fluorescence of singlet oxygen sensor green detected by confocal laser scanning microscopy. It is proposed here that singlet oxygen is formed by the decomposition of high-energy intermediates such as dioxetane or tetroxide formed by oxidative damage of biomolecules. 相似文献
107.
Carolina Serena Enrique Calvo Mari Paz Clares María Luisa Diaz Javier U. Chicote Raúl Beltrán-Debon Ramón Fontova Alejandro Rodriguez Enrique García-Espa?a Antonio García-Espa?a 《PloS one》2015,10(3)
Background
The clinical use of purified SOD enzymes has strong limitations due to their large molecular size, high production cost and immunogenicity. These limitations could be compensated by using instead synthetic SOD mimetic compounds of low molecular weight.Background/Methodology
We have recently reported that two SOD mimetic compounds, the MnII complexes of the polyamines Pytren2Q and Pytren4Q, displayed high antioxidant activity in bacteria and yeast. Since frequently molecules with antioxidant properties or free-radical scavengers also have anti-inflammatory properties we have assessed the anti-inflammatory potential of Pytren2Q and Pytren4Q MnII complexes, in cultured macrophages and in a murine model of inflammation, by measuring the degree of protection they could provide against the cellular injury produced by lipopolisacharide, a bacterial endotoxin.Principal Findings
In this report we show that the MnII complex of Pytren4Q but not that of Pytren2Q effectively protected human cultured THP-1 macrophages and whole mice from the inflammatory effects produced by LPS. These results obtained with two molecules that are isomers highlight the importance of gathering experimental data from animal models of disease in assessing the potential of candidate molecules.Conclusion/Significance
The effective anti-inflammatory activity of the MnII complex of Pytren4Q in addition to its low toxicity, water solubility and ease of production would suggest it is worth taking into consideration for future pharmacological studies. 相似文献108.
Purification of anti‐colorectal cancer monoclonal antibody CO17‐1A from insect cell culture using a French press and sonication 下载免费PDF全文
Monoclonal antibody (mAb) CO17‐1A binds to GA733, which is a tumor‐associated glycoprotein antigen highly expressed on the colorectal cancer cell surface. Thus, mAb CO17‐1A is considered a useful biomolecule for diagnosis and treatment against colorectal cancer. Previously, we established a baculovirus–insect cell expression system for the production of mAb CO17‐1A. In order to use mAb CO17‐1A as a diagnostic and therapeutic tool, however, the antibody must be properly purified from the insect cells. In this study, our aim was to investigate effective purification processes of mAb CO17‐1A expressed in Spodoptera frugiperda (Sf9) insect cells, using a French press and sonication for cell disruption. SDS‐PAGE confirmed that both mAb CO17‐1A and mAb CO17‐1A fused to the KDEL endoplasmic reticulum (ER) retention signal (mAb CO17‐1AK) were expressed clearly in Sf9 insect cells. Western blot analysis showed that detection levels of mAb CO17‐1A and CO17‐1AK were higher when the insect cells were disrupted two times by the French press and then sonicated, compared to only one French press disruption plus sonication. Optical microscopy confirmed that insect cells treated with both the French press and sonication were properly disrupted. Analysis of gene sequence information on mAb CO17‐1A verified that a signal peptide is present but a transmembrane protein does not exist. These results suggest that cell disruption by the French press twice and sonication once is an effective method for improving purification efficiency. 相似文献
109.
Koldo Garcia-Etxebarria María Alma Bracho Juan Carlos Galán Tomàs Pumarola Jesús Castilla Raúl Ortiz de Lejarazu Mario Rodríguez-Dominguez Inés Quintela Núria Bonet Marc Garcia-Garcerà Angela Domínguez Fernando González-Candelas Francesc Calafell CIBERESP Cases Controls in Pandemic Influenza Working Group 《PloS one》2015,10(10)
110.
Eva Prats Elena Tejero Paloma Pardo Adelaida Gavilán Raúl Galera José Ramón Donado Miguel ángel Racionero Raquel Casitas Antonio Zapatero Francisco García-Río 《PloS one》2015,10(10)
BackgroundThe six-second spirometry has been proposed as an alternative to diagnose airflow limitation, although its prognostic value in patients with chronic obstructive pulmonary disease (COPD) remains unknown. The purpose of this study was to determine the prognostic value of the postbronchodilator forced expiratory volume in 1 second (FEV1)/forced expiratory volume in 6 seconds (FEV6) ratio and FEV6 in COPD patients.ConclusionsIn a general COPD outpatient population, airflow obstruction assessed by the FEV1/FEV6 is an independent risk factor for both death and hospitalization. 相似文献