利用烟草高效转化功能基因体系的建立

对农杆菌介导的烟草遗传转化条件进行探索,以20 mg/L的潮霉素作为转化苗的筛选压,稀释至OD600为O.2～O.3的农杆菌对外植体浸染15 min,于添加300 mg/L头孢霉素的生根培养基中诱导生根,可稳定获得较高的阳性转化率.经PCR和PCR-Souttlern检测,阳性转基因植株达84.8%以上.通过本研究建立了一种稳定高效的功能基因烟草遗传转化体系.     

Requirements and tests for HPLC apparatus and methods in pharmaceutical quality control

Summary One possible method has been demonstrated for pharmaceutical quality control which is not restricted to this field and fulfils and documents the prerequisites for reliable, accurate and precise HPLC analysis. This includes validation which shows that the method is able in principle to fulfil the requirements, the apparatus test which shows that the apparatus generally works correctly and precisely and the system suitability test which shows that the method provides accurate and precise results on this apparatus and with this column for the analysis in question when other non-equipment and non-method-induced errors are excluded.     

The practical advantages of hospital quality systems such as NIAZ/PACE

 The Netherlands Institute for Accreditation of Hospitals (NIAZ) was established at the end of 1998. It was founded by the PACE foundation, the Society of University Hospitals, the Netherlands Association of General Hospitals and the Society of Medical Specialists in the Netherlands. Since then in the Netherlands 19 pilot accreditations have been performed based upon 35 NIAZ-PACE standards for hospital departments and functions, and the overall standard 'quality system'. The aims and methods of the accreditation system were inspired by examples from the other side of the Atlantic, especially from Canada. The characteristics are: voluntary-based, self-evaluation, peer-review and aiming at continuous improvement of quality of care. Received: 15 April 2000 · Accepted: 15 April 2000     

Chemical Profile of Danning Tablets by LC-DAD Coupled with ESI-MS

HPLC-DAD coupled to an electrospray tandem mass spectrometer (ESI-MS) has been developed to analysis the chemical profile of Danning tablets. Separation was performed on a ZORBAX Extend C₁₈ analytical column by gradient elution with acetonitrile and formate buffer (containing 12.6 mmol L⁻¹ formic acid, adjusted to pH 5.0 with ammonia) at a flow rate of 0.8 mL⁻¹ min. Twenty-three peaks were identifies as the major compounds in this preparation. Similarity measurements of the HPLC-UV chromatograms were investigated by a Similarity Evaluation System for Chromatographic Fingerprinting of Traditional Chinese Medicine (TCM) in the range of 0.84–0.99, and principal components analysis (PCA) on the quantitative analysis data was further performed to evaluate the variation among batches. The proposed method gives chemical profiles of multi-components in one run, and therefore can be readily utilized as a comprehensive quality control approach for TCM.     

Ultrasound freeze-thawing style pretreatment to improve the efficiency of the vacuum freeze-drying of okra (Abelmoschus esculentus (L.) Moench) and the quality characteristics of the dried product

Vacuum freeze-drying is a new and high technology on agricultural product dehydrating dry, but it faces the high cost problem caused by high energy consumption. This study investigated the effect of ultrasound (US), freeze-thawing (including the freeze-air thawing (AT), freeze-water thawing (WT), freeze-ultrasound thawing (UST), and freeze-air ultrasound thawing (AT + US)) pretreatments on the vacuum freeze-drying efficiency and the quality of dried okra. The results indicated that the application of ultrasound and different freeze-thawing pretreatments reduced the drying time by 25.0%–62.50% and the total energy consumption was 24.28%–62.35% less. The AT pretreatment reduced the time by of okra slices by 62.50% and the total energy consumption was 62.35% less. The significant decrease in drying time was due to a change in the microstructure caused by pretreatment. Besides, the okra pretreated with the US retained most of the quality characteristics (flavor, color, hardness, and frangibility) among all methods, while, AT + US had the most changeable characteristics in quality, which is deprecated in our study. The okra pretreated with the US and AT, separately, had the best dry matter content loss (9.008%, 5.602%), lower chlorophyll degradation (5.05%, 5.44% less), and higher contents of total phenolics, total flavonoids, and pectin, with strong antioxidant capacity, compared to other methods. The pretreatments did not have a large effect on the functional groups and the structure of pectin, but slightly affected the viscosity. It can be concluded that AT and US pretreatment methods are better than others.     

Suspension culture in a T-flask with acoustic flow induced by ultrasonic irradiation

Suspension culture is an essential large-scale cell culture technique for biopharmaceutical development and regenerative medicine. To transition from monolayer culture on the culture surface of a flask to suspension culture in a bioreactor, a pre-specified cell number must first be reached. During this period of preparation for suspension culture, static suspension culture in a flask is generally performed because the medium volume is not large enough to use a paddle to circulate the medium. However, drawbacks to this static method include cell sedimentation, leading to high cell density near the bottom and resulting in oxygen and nutrient deficiencies. Here, we propose a suspension culture method with acoustic streaming induced by ultrasonic waves in a T-flask to create a more homogeneous distribution of oxygen, nutrients, and waste products during the preparation period preceding large-scale suspension culture in a bioreactor. To demonstrate the performance of the ultrasonic method, Chinese hamster ovary cells were cultured for 72 h. Results showed that, on average, the cell proliferation was improved by 40% compared with the static method. Thus, the culture time required to achieve a 1000-fold increase could be reduced by 32 h (a 14% reduction) compared with the static method. Furthermore, the ultrasonic irradiation did not compromise the metabolic activity of the cells cultured using the ultrasonic method. These results demonstrate the effectiveness of the ultrasonic method for accelerating the transition to large-scale suspension culture.     

Evaluation of a Gelatin-Modified Poly(ε-Caprolactone) Film as a Scaffold for Lung Disease

Lung transplantation is a necessary step for the patients with the end stage of chronic obstructive pulmonary disease. The use of artificial lungs is a promising alternative to natural lung transplantation which is complicated and is restricted by low organ donations. For successful lung engineering, it is important to choose the correct combination of specific biological cells and a synthetic carrier polymer. The focus of this study was to investigate the interactions of human lung epithelial cell line NCl-H292 that is involved in lung tissue development with the biodegradable poly(ε-caprolactone) before and after its chemical modification to evaluate potential for use in artificial lung formation. Also, the effect of polymer chemical modification on its mechanical and surface properties has been investigated. The poly(ε-caprolactone) surface was modified using aminolysis followed by immobilization of gelatin. The unmodified and modified polymer surfaces were characterized for roughness, tensile strength, and NCl-H292 metabolic cell activity. The results showed for the first time the possibility for NCI-H292 cells to adhere on this polymeric material. The resazurin assay showed that the metabolic activity at 24?h postseeding of 80% in the presence of the unmodified and greater than 100% in the presence of the modified polymer was observed. The roughness of the poly(ε-caprolactone) increased from 4 to 26?nm and the film strength increased from 0.01 to 0.045 kN when the material was chemically modified. The results obtained to date show potential for using modified poly(ε-caprolactone) as a scaffold for lung tissue engineering.     

Hydrogel microfluidic co‐culture device for photothermal therapy and cancer migration

We developed the photo‐crosslinkable hydrogel microfluidic co‐culture device to study photothermal therapy and cancer cell migration. To culture MCF7 human breast carcinoma cells and metastatic U87MG human glioblastoma in the microfluidic device, we used 10 w/v% gelatin methacrylate (GelMA) hydrogels as a semi‐permeable physical barrier. We demonstrated the effect of gold nanorod on photothermal therapy of cancer cells in the microfluidic co‐culture device. Interestingly, we observed that metastatic U87MG human glioblastoma largely migrated toward vascular endothelial growth factor (VEGF)‐treated GelMA hydrogel‐embedding microchannels. The main advantage of this hydrogel microfluidic co‐culture device is to simultaneously analyze the physiological migration behaviors of two cancer cells with different physiochemical motilities and study gold nanorod‐mediated photothermal therapy effect. Therefore, this hydrogel microfluidic co‐culture device could be a potentially powerful tool for photothermal therapy and cancer cell migration applications.     

结冷胶与聚乙二醇丙烯酸酯双网络凝胶的制备及生物相容性评价

针对结冷胶脆性较大的问题,将聚乙二醇丙烯酸酯(PEGDA)引入结冷胶,通过紫外交联制备了结冷胶/PEGDA双网络凝胶,并对单组分凝胶和双网络凝胶的溶胀性能、微观形貌、拉伸力学性能、动态压缩性能和流变性能等进行比较.结果表明,双网络凝胶在类生理环境中具有较小的溶胀率和较好的尺寸稳定性,PEGDA的引入能够大幅度提高结冷胶的韧性,双网络凝胶的拉断伸长率可达340%,断裂能达1.01×103J/m2,与天然关节软骨相当.将成纤维细胞种植在凝胶内部进行体外三维立体培养,结果显示,细胞在凝胶内部生存状态良好,双网络凝胶的细胞负载率高于单网络结冷胶,说明该体系在生物医用领域具有良好的应用前景.     

Biodegradation-induced surface change of polymer microspheres and its influence on cell growth

Biodegradable microspheres were fabricated by poly(?-caprolactone) (PCL) homopolymer and poly(?-caprolactone-b-ethylene oxide) (PCL-b-PEO) amphiphilic block copolymer. The regulation of microsphere surface morphology was successfully achieved by controlled enzymatic degradation. The morphological changes induced by biodegradation and their influences on the growth of MG-63 human osteosarcoma cells were studied. Results based on the evaluation of cytotoxicity and the morphological observation of MG-63 cells cultivated on microspheres showed better growth of cells on the surface of degraded microspheres than on the surface of those undegraded microspheres no matter they were fabricated by homopolymers or copolymers. The influences of morphological changes of microsphere surface before and after biodegradation on MG-63 cell growth were discussed. The results of this work indicated that the biodegradation-induced morphological changes of microspheres could be well controlled and were favorable for MG-63 cell attachment and proliferation.