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The results presented here provide the first single-cell genetic assay for Tay-Sachs disease based on real-time PCR. Individual lymphoblasts were lysed with an optimized lysis buffer and assayed using one pair of primers that amplifies both the wild type and 1278 + TATC Tay-Sachs alleles. The resulting amplicons were detected in real time with two molecular beacons each with a different colored fluorochrome. The kinetics of amplicon accumulation generate objective criteria by which to evaluate the validity of each reaction. The assay had an overall utility of 95%, based on the detection of at least one signal in 235 of the 248 attempted tests and an efficiency of 97%, as 7 of the 235 samples were excluded from further analysis for objective quantitative reasons. The accuracy of the assay was 99.1%, because 228 of 230 samples gave signals consistent with the genotype of the cells. Only two of the 135 heterozygous samples were allele drop-outs, a rate far lower than previously reported for single-cell Tay-Sachs assays using conventional methods of PCR. Copyright © 2002 John Wiley & Sons, Ltd. 相似文献
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Prior to a prospective application of amniotic fluid (AF) cell filtration to early amniocentesis, we tested the technique on a surplus from mid-trimester samples. By using the same sample size of 5 ml in experiments with a filter and in routine diagnostic procedures (control), we evaluated an optimal filter system. The prolonged culture time of filtered cells and the reduced number of clones are most probably due to mechanical stress (filtration pressure), whereas loss of the cells by adhesion to the filter system, and an AF-free culture medium (growth factors) are suggested to be less important. The AF cells are very sensitive to mechanical stress. Slow filtration (⩽3 ml AF/min) through filters with a high porosity and the largest possible pore size should be preferred. A mixed cellulose ester filter membrane with a pore size of 5·0 μm proved to be the most efficient, allowing harvest of the filtered cells after only a slight prolongation of the culture time (+2·4 days) compared with unfiltered aliquots. A filter set with a bypass connected by three-way taps allows cell filtration during either aspiration or reinjection of the AF. Cell filtration after amniocentesis and consecutive reverse flushing of the membrane with the appropriate amount of culture medium proved to be the best with regard to easy handling and reducing the risk of bacterial contamination. 相似文献
937.
构建了双室型微生物燃料电池(MFC),探讨了硫酸盐对MFC阴极脱氮效果和产电性能的影响。实验结果表明:缺氧阴极室中,硝酸盐氮、硫酸盐的还原存在一定的竞争关系,且硫酸盐对电子的竞争能力比硝酸盐氮强;当S与N的质量比(记为S/N)为2∶1时,72 h后的硫酸根去除率达96.41%,而硝酸盐氮去除率仅为36.86%;在S/N为2∶1的条件下,当电压达307.6 m V时,功率密度达最大值15.77 W/m2,电池内阻为100Ω,与未加入硫酸盐时相比产电性能明显提高。 相似文献
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二氧化硫(SO_2)是一种常见的大气污染物,目前关于SO_2对木本植物的毒害作用及相关机制并不清楚.本文以木本植物胡杨的愈伤细胞为材料,研究SO_2衍生物对胡杨细胞的致死效应,以及过氧化氢(H_2O_2)与一氧化氮(NO)在SO_2诱导胡杨细胞死亡中的信号调节作用.研究发现:SO_2衍生物处理(1~5 mmol·L-1)可诱发胡杨细胞死亡,且SO_2衍生物浓度越大、处理时间越长,细胞死亡率越高.2 mmol·L-1SO_2衍生物处理胡杨细胞后,胞内H_2O_2和NO水平显著升高,且H_2O_2水平的升高先于NO.一定浓度的外源H_2O_2或NO供体SNP能够提高SO_2胁迫下胡杨细胞的死亡率;而使用H_2O_2清除剂CAT和ASA、NO清除剂c PTIO、NO合成抑制剂钨酸钠后,SO_2诱导的细胞死亡率明显降低.进一步实验发现,外源H_2O_2可以提高SO_2胁迫下胡杨细胞的硝酸还原酶(NR)活性,促进胞内NO产生;而利用CAT和ASA清除H_2O_2后,细胞NR活性和NO产生均受到明显抑制.此外,SO_2胁迫下,外源SNP能够抑制抗氧化酶(CAT和APX)活性,增加胡杨细胞内的H_2O_2水平,而一定浓度的c PTIO和钨酸钠均可提高CAT和APX活性,降低胞内H_2O_2水平.结果表明:SO_2胁迫下,胡杨细胞快速产生的H_2O_2能够激活硝酸还原酶活性,促进NO生成,同时NO能够通过抑制抗氧化酶活性而提高H_2O_2水平.H_2O_2与NO互作调控SO_2诱导的胡杨细胞死亡. 相似文献
940.
Qian Xu Weiqi Zhang Jian Zhao Lei Xing Qiang Ma Li Xu 《International Journal of Green Energy》2018,15(3):181-188
Unlike the situation in the direct methanol fuel cell (DMFC) fed with dilute liquid methanol solution, the required water in anode for a DMFC fed with neat methanol is entirely transported from cathode. In this study, the water concentration in anode catalyst layer of such a DMFC operating with fully active mode is theoretically analyzed, followed by the experimental investigations on the effects of air flow rate and operating temperature on cell performance. The results revealed that the air flow rate has a strong impact on cell performance, especially at larger current density. Overmuch air causes rapid decline of cell performance, which results from the dehydration of membrane and lack of water in the anode reaction sites. Raising temperature induces faster reaction kinetics, while undesired stronger water dissipation from the DMFC. In practice, the stable cell resistance can be used as a criterion to help the DMFC to achieve a high and sustainable performance by finely combining the air flow rate and operating temperature. 相似文献