Stain efficiency and MALDI-TOF MS compatibility of seven visible staining procedures

Visible stain is still the most popular protein staining method used in proteomic approaches. However, most published data have been derived from comparisons between visible dyes and fluorescent dyes. In this work, we have focused on seven widely used visible staining procedures—Neuhoff CCB, blue silver, and five silver stains (LKB SN, He SN, Yan SN, Vorum SN, and Blum SN)—and studied their stain efficiencies and MALDI-TOF MS compatibilities on 1-D and 2-D PAGE. It was concluded that blue silver is slightly better in terms of stain efficiency than Neuhoff CCB, but it presented worse MS compatibility. Neuhoff CCB presented better MS compatibility and superior linearity but worse sensitivity than silver stains. Among the five silvering procedures, He SN showed the best MS compatibility and a reasonable staining efficiency; Yan SN lowered the chances of obtaining the protein identity by PMF but gave the best stain efficiency; Vorum SN gave a very clear background and a great contrast, while Blum SN was the worst in this respect. The implications of these results for the selection of a convenient stain are discussed according to specific objectives as well as practical aspects.     

High‐throughput negative detection of SDS‐PAGE separated proteins and its application for proteomics

A negative detection method for proteins on SDS‐PAGE is described. In this method, Eosin Y (EY) was selectively precipitated in the gel background, which is absent from those zones where proteins are located through the formation of a stable water‐soluble protein–dye complex. Negative staining of proteins using EY, allows high‐sensitivity, low‐cost, and simple protocol. The new described method takes less than an hour to complete all the protocol, with a detection limit of 0.5 ng of single protein band. Comparing with imidazole‐zinc negative stain, EY dye provides broader linear dynamic range, higher sensitivity and reproducibility, and better obvious contrast between the protein bands or spots and background. Furthermore, the novel technique developed here presented a real practical method for simultaneous processing of multiple gels, which makes it possible to perform high‐throughput staining for proteome research. Additionally, we have also compared the influence of staining method on the quality of mass spectra by PMF.     

Enhanced visualization of polysaccharides from aqueous suspensions

Aqueous suspensions of polysaccharides such as those prepared for domestic and industrial applications or present in natural waters, although difficult to visualize by conventional transmission electron microscopy (TEM) because of their poor electron density, can be characterized at the ultrastructural level by using milden bloc staining and contrast enhancement by energy-filtered TEM (EF-TEM). The advantages and drawbacks of the proposed method are discussed in relation to the different parameters controlling the quality of final images. It is shown, with synthetic polysaccharides, purified algal fibrils and lacustrine exocellular polymers as key examples, that optimizing specimen preparation and visualization parameters allows unbiased identification of organic substructures never revealed or strongly degraded by classical microscopic procedures.     

光谱法用于血氧饱和度无损测量

综述了血氧饱和度无损测量的方法,泰勒级数法、近红外光谱法、氧指数(IOK)法以及基于蒙特卡罗的多元线性回归法等,分析了这些方法的优缺点.基于光动力疗法治疗机制(PDT),提出了一种修正的氧指数法,初步分析表明该法能够监测光动力疗法治疗鲜红斑痣(PWS)过程中血氧饱和度变化情况.     

Design and synthesis of a novel fluorescent protein probe for easy and rapid electrophoretic gel staining by using a commonly available UV‐based fluorescent imaging system

A new fluorescent molecular probe, methyl 3‐(3,5‐bis((bis(pyridin‐2‐ylmethyl)amino)‐methyl)‐4‐hydroxyphenyl)‐2‐(5‐(dimethylamino)naphthalene‐1‐sulfonamido) propanoate, dizinc(II) chloride salt (Dansyl‐ 1 ‐Zn(II)), which possesses Zn(II) complexes and a dansyl group, was designed and synthesized to enable the detection of proteins in solution and in high‐throughput electrophoresis by using a UV‐based detection system. Dansyl‐ 1 ‐Zn(II) exhibited weak fluorescence in the absence of proteins and strong green fluorescence at approximately 510 nm in the presence of BSA upon irradiation with light at a wavelength of 345 nm. Compared with conventional protocols for in‐gel SDS‐PAGE protein staining (e.g. silver staining, SYPRO Ruby, and Oriole), the operating times of which range from 90 min to overnight, Dansyl‐ 1 ‐Zn(II) allowed 1‐step protein staining (SDS‐PAGE →Staining →Detection) and shortened the operating time (35 min) with high sensitivity (LOD: 1 ng or less) under 312‐nm or 365‐nm light excitation with orange or red emission filters, respectively. Moreover, Dansyl‐ 1 ‐Zn(II) was successfully applied to protein identification by MS via in‐gel tryptic digestion, Western blotting, and Native‐PAGE. Accordingly, Dansyl‐ 1 ‐Zn(II) may facilitate highly sensitive and high‐throughput protein detection, and it may be widely applicable as a convenient tool in various scientific and medical fields.     

鲜红斑痣光谱测定系统与临床实验

对进行激光治疗的鲜红斑痣患者的皮肤进行光谱定量分析,有利于正确分析患者病变皮肤的光学特性、调整手术设备的工作参数和提高激光治疗的效果。基于对鲜红斑痣形成机理分析及激光临床治疗过程观察,文章建立了一套自动数据采集和分析的光谱系统,可以用于测定鲜红斑痣皮肤微区在380～780 nm范围内的光谱,光谱分辨率为1 nm。实验可测定不同年龄、不同色素痣的光谱曲线,分析并讨论了影响系统工作的原因和解决方法。     

一种新的混合斑精子分离方法

制备了精子表面膜抗原受精素β蛋白(Fertilinβ)的特异性抗体,通过抗原-抗体反应和抗体固相偶联技术,将抗体连接到琼脂糖球珠上,建立了混合斑精子分离纯化的新方法.首先,通过聚合酶链式反应(PCR)扩增编码人受精素β蛋白第341~373位氨基酸的基因序列,构建PGEX-4T-1/Fertilinβ原核表达质粒;其次,诱导表达GST-Fertilinβ融合蛋白,制备受精素β蛋白多克隆抗体,免疫荧光检测结果表明,受精素β蛋白定位于精子的头后部,并且阴道上皮细胞没有受精素β蛋白的表达;最后,将受精素β抗体与ProteinA琼脂糖球珠连接构建固相抗体系统,将精子吸附于表面,可从混合斑(精子与阴道上皮细胞混合液)中分离纯化精子.本文为性犯罪案件侦破提供了新的方法.     

化学染色法在pn结分析中的应用研究

化学染色法是pn结分析中较普遍的方法,不同的化学染色法所显现出pn结的剖面形貌会受到化学试剂、pn结固有性质、染色工艺参数的影响。研究了常用的铬酸溶液染色法、醋酸溶液染色法、BOE溶液染色法和硫酸铜溶液染色法之间的差异。在一定范围内,铬酸溶液染色法、醋酸溶液染色法、BOE溶液染色法适合染出p区,硫酸铜溶液染色法适合染出n区,铬酸溶液染色法可以染出较小尺寸的pn结。染色时间的增加将会使所染pn结形貌变大,而染色溶液温度的升高对所染pn结形貌影响不大,但会使部分在常温下不能染出的pn结染出,BOE溶液染色法所得出的pn结较接近真实形貌。