Quantitative detection of gold nanoparticles on individual, unstained cancer cells by scanning electron microscopy

Gold nanoparticles are rapidly emerging for use in biomedical applications. Characterization of the interaction and delivery of nanoparticles to cells through microscopy is important. Scanning electron microscopes have the intrinsic resolution to visualize gold nanoparticles on cells. A novel sample preparation protocol was developed to enable imaging of cells and gold nanoparticles with a conventional below lens scanning electron microscopes. The negative influence of 'charging' on the quality of scanning electron microscopes' images could be limited by deposition of biological cells on a conductive (gold) surface. The novel protocol enabled high-resolution scanning electron microscopes' imaging of small clusters and individual gold nanoparticles on uncoated cell surfaces. Gold nanoparticles could be counted on cancer cells with automated routines.     

Germline stem cell lineage tracing by free-floating immunofluorescent assay of mouse seminiferous tubule

Whole-mount immunohistochemistry (whole-mount IH) of the seminiferous tubule is widely used to investigate the self-renewal and differentiation of spermatogonial stem cells (SSCs). Examination of the length of spermatogonial cysts is critical for tracing SSCs lineage by using Whole-mount IH. However, it is difficult for antibody molecules to penetrate into the depth of seminiferous epithelium because its thickness and the tight peritubular myoid and basement membrane outside. Here, we developed a free-floating immunofluorescent procedure of mouse seminiferous tubules using regular incubation time and normal antibody concentration. Microscopic results showed that undifferentiated spermatogonia were positively labeled by promyelocytic leukemia zinc finger protein, E-cadherin, and glial cell line-derived neurotrophic factor family receptor alpha 1, respectively. Spermatogonial cysts in varied length were revealed clearly and spermatogonia subpopulations including A(single) (A(s)), A(paired) (A(pr)), and A(aligned) (A(al)) were distinguished in lower background images. This method provides us an alternate simple way to trace the lineage of individual SSCs and show their three-dimensional locations and distributions within their niches anatomically in next step.     

The intracellular distribution of vinculin and alpha2 integrin in epithelial cells and chondrocytes

The purpose of this study was to demonstrate the presence of vinculin and alpha₂ integrin in chondrocytes in situ and epithelial cells. We also determined that the appropriate fixation and extraction protocols for immunohistochemistry and laser scanning confocal microscopy for an integral membrane protein and an actin-associated protein in cultured cells and whole tissue was different. Cultured epithelial cells, whole mount human cornea and avian cartilage were fixed and prepared using a number of standard procedures used for indirect fluorescence immunohistochemistry. The distribution of vinculin was cell-type and fixation-specific. Chondrocytes and cultured epithelial cells demonstrated vinculin in areas that appear to be associated with filamentous actin. Vinculin was associated with cell membranes in human cornea. The expression of alpha₂ integrin observed in chondrocytes fixed with methanol, paraformaldehyde, or formaldehyde is consistent with its role in cell-substrate interaction, but may also suggest a role in dividing and differentiating cells. The localization of alpha₂ integrin in human corneal epithelia supports its role as a cell-cell adhesion molecule. The cytoplasmic distribution of vinculin and alpha₂ integrin in tissues fixed without detergent extraction suggests that the fixation step may be sufficient for antibody penetration and antigen extraction. These studies are the first report of vinculin and alpha₂ integrin in embryonic chondrocytes. In addition we have shown that confocal laser scanning microscopy combined with proper fixation and extraction protocols may optimize the localization of antigens in cultured and whole mount cells.     

High-resolution subunit detection of glutamate receptor by ultrasmall gold nanoparticles

In this study, we aimed to increase the sensitivity of protein labeling using 1.4 nm gold nanoparticles and glutamate δ2 receptor (GluD2) from the postsynaptic membrane of the Purkinje cells. The very small marker size of the particles reduces the steric hindrance between antibodies leading to a higher labeling efficiency of more than one subunit per single receptor molecule. The nanoparticles are visible in 200 kV dark‐field scanning transmission electron microscope on freeze‐fractured carbon replica of nervous tissue after plasma cleaning treatment. The different elemental composition of nanoparticles as Au nanogold or CdS quantum dot can be distinguished by energy dispersive X‐ray spectroscopy. This method ensures detection of an average of three subunits per GluD2 and often labels all four of them with 1.4 nm Au nanoparticles. It is concluded that this high‐resolution microscopic method is useful for exploring the quaternary structure of membrane proteins. Microsc. Res. Tech. 75:1159–1164, 2012. © 2012 Wiley Periodicals, Inc.     

Noninvasive 3D vital imaging and characterization of notochordal cells of the intervertebral disc by femtosecond near-infrared two-photon laser scanning microscopy and spatial-volume rendering

The central region of the intervertebral disc (IVD) in infant humans is made and maintained by notochordal cells (NCs). These cells disappear during maturation to be replaced by mature chondrocyte-like cells. NCs are completely different morphologically from the mature chondrocyte-like IVD cells and have complex and essential functions but little is known about them. Recently, two-photon laser scanning microscopy (TPLSM) using near-infrared (NIR) femtosecond pulsed lasers has emerged as a promising noninvasive optical technique for observing unfixed living 3D biological specimens in situ and in vitro. Several lines of evidence suggest that compared with conventional laser scanning confocal microscopy (LSCM), femtosecond NIR laser-based TPLSM has any number of advantages including 3D resolution without a spatial filter (confocal pinhole), minimal photobleaching, and photodamage above and below the focal plane, and importantly, greater depth penetration. We have thus taken advantage of these unique features of femtosecond laser-based TPLSM for vital 3D imaging in conjunction with advanced spatial-volume rendering modalities to compare morphologies of NCs/clusters from pig caudal discs with chondrocyte-like IVD cells from bovine caudal discs, both in ex vivo tissue and when isolated and grown in vitro within 3D alginate scaffolds. Our results provide evidence that (a) ex vivo notochordal tissue consists of areas with NC clusters, and those dominated by tubular structures of low cell density (b) within 3D in vitro scaffolds the morphology of NC is heterogeneous and the cells contain distinct cytoplasmic vacuole-like structures occasionally including acidic subinclusions (c) a quantitative determination based on 3D spatial and volumetric-rendering reveals an average NC diameter of 22.05 microm (range 11.96-46.63 microm) and NC volume of 9701 microm(3) (2041-36427 microm(3)) whereas chondrocyte-like cells have a mean volume of 3279 microm(3) and diameter of 12.20 microm. Taken together, this study demonstrates that femtosecond TPLSM has unique advantages over other conventional histological and in particular LSCM for high resolution noninvasive vital characterization of notochordal and chondrocyte-like cells of IVD over extended depths beyond 300-500 microm.     

高速锯床防护罩抗冲击能力的有限元分析

利用ANSYS/LS_DYNA的动力学分析功能对GS3050型超高速数控锯床安全防护罩的抗冲击能力进行了数值分析,对刀具碎片侵彻有机玻璃的过程进行了数值模拟.结果表明,采用厚度为15 mm的改性丙烯酸脂有机玻璃作为GS3050锯床的安全防护罩是可靠的.数值分析和试验的结果吻合.     

黄河三小间水情自动测报系统遥测站供电系统设计计算

供电系统是水情自动测报系统遥测站的心脏，可靠与否至关重要。黄河三小间水情自动测报系统过几年的使用，效果良好，证明其供电系统方案正确、设计合理。本文介绍了黄河三小间水情自动测报系统遥测站供电系统的方案选择和太阳能电池、蓄电池容量的计算。     

有机碳源对厌氧氨氧化污泥颗粒化的影响

为探究有机碳源对厌氧氨氧化(Anammox)污泥颗粒形成的影响,采用两组平行的SBR,通过改变进水中有机碳源的质量浓度进行研究.结果表明:适量的有机碳源(130 mg/L)可通过提高反应器中的胞外聚合物(EPS)含量从而加速颗粒污泥的形成,提高污泥的沉降性能.添加与不添加有机碳源的两组反应器分别于28和35 d颗粒化成功,平均颗粒粒径分别达450和409μm.过量有机碳源( 230 mg/L)会使污泥出现解体,粒径减小,污泥的沉降性能明显变差.当有机碳源小于110 mg/L时,可通过反硝化作用促进厌氧氨氧化反应从而提高脱氮效率;但是当有机碳源质量浓度大于110 mg/L时,会抑制厌氧氨氧化反应,并降低脱氮效率.在厌氧氨氧化工艺实际运行中,应避免有机物质量浓度超过110 mg/L.     

Highly reliable hybrid nano-stratified moisture barrier for encapsulating flexible OLEDs

Herein, a novel thin-film encapsulation for flexible organic light-emitting diodes (FOLEDs) is proposed, and its long-term reliability in tensile stress conditions was tested. The hybrid nano-stratified moisture barrier consists of 2.5 dyads of an Al₂O₃/ZnO nano-stratified structure and a S-H nanocomposite organic layer. The nano-stratified structure is prepared by low-temperature atomic layer deposition and the S-H nanocomposite by spin-coating at a thickness of 30 and 120 nm, respectively. An optical transmittance of 89.05% was measured with the 2.5-dyad hybrid nano-stratified moisture barrier with a total thickness of 330 nm. A low water vapor transmission rate (WVTR) of 1.91 × 10⁻⁵ g/m²day was recorded based on an electrical Ca test at 30 °C and 90% R.H. without losing its properties after a bending test. With this highly reliable hybrid nano-stratified moisture barrier, FOLEDs were successfully encapsulated. After 30 days under conditions of 30 °C and 90% R.H. with tensile stress, the J-V-L performances of the FOLEDs were comparable to those of the initial state without dark spots. These results suggest that this hybrid nano-stratified moisture barrier is an excellent method for encapsulating FOLEDs.     

Research on temperature field and temperature effect of steel silos under solar radiation

综合使用光线追踪法和点盒位置判断理论，提出了一种高效的钢板筒仓太阳辐射阴影区计算方法，并通过试验验证了该方法的计算精度。分析了钢板筒仓加盖和不加盖、满仓和空仓时太阳辐射非均匀温度场的时空分布规律，其中不加盖空仓时的太阳辐射非均匀温度场对结构最为不利。不加盖空仓时钢板筒仓结构正温差温度效应最大，与不考虑太阳辐射均匀温度作用工况相比，最大温度变形、最大温度应力、支座最大径向反力分别增加了135%、85%、85%，支座环向反力从0kN增加到132 kN，建议在工程设计中合理考虑太阳辐射非均匀温度效应。