Evolution of the silver and gold stains in neurohistology

Scientific investigations depend on the reliability of the observations that can be made. This reliability is determined in part by the understanding of the techniques and technology used to make the observations. The limitations and the strengths of the methodology and the equipment used must be evaluated thoroughly. The extent to which this is and has been the case for the use of the metal based stains in neuroscience is the subject of this paper. I evaluate the metallic stains used for neuroscience from several perspectives. I review briefly the state of neurohistology prior to its “golden years,” 1870-1910. Then I trace the development of the silver based stains used for neurohistology. I wanted to discuss the reasoning used by the originators of the silver based techniques in developing their specific procedures, but discovered that while procedures may be published, the methods and ideas used to arrive at the final procedures are not usually described in published work.     

Evolution of the silver and gold stains in neurohistology

Scientific investigations depend on the reliability of the observations that can be made. This reliability is determined in part by the understanding of the techniques and technology used to make the observations. The limitations and the strengths of the methodology and the equipment used must be evaluated thoroughly. The extent to which this is and has been the case for the use of the metal based stains in neuroscience is the subject of this paper. I evaluate the metallic stains used for neuroscience from several perspectives. I review briefly the state of neurohistology prior to its “golden years,” 1870–1910. Then I trace the development of the silver based stains used for neurohistology. I wanted to discuss the reasoning used by the originators of the silver based techniques in developing their specific procedures, but discovered that while procedures may be published, the methods and ideas used to arrive at the final procedures are not usually described in published work.     

长链非编码RNA的作用机制及其研究方法

长链非编码RNA(Long non-coding RNA, lncRNA)通过多种机制发挥其生物学功能, 这些机制包括基因印记、染色质重塑、细胞周期调控、剪接调控、mRNA降解和翻译调控等。lncRNA通过这些作用机制在不同水平进行基因表达调控。在研究lncRNA功能的过程中, 研究方法的建立和应用起着非常重要的作用。目前用于lncRNA研究的主要方法有：微阵列、转录组测序、Northern印迹、实时荧光定量逆转录-聚合酶链反应、荧光原位杂交、RNA干扰和RNA结合蛋白免疫沉淀等。文章着重介绍了3种前沿方法, 即：在线快速预测RNA与蛋白质相互作用的catRAPID、RNA纯化的染色质分离(Chromatin isolation by RNA purification, ChIRP)以及非编码RNA沉默与定位分析技术(Combined knockdown and localization analysis of non-coding RNAs, c-KLAN)。     

寄生性膜翅目昆虫毒素的生态机制及应用前景

膜翅目昆虫利用高效的毒素进行自身防卫、攻击猎物和调节寄主生长发育.从寄生性膜翅目昆虫毒素的产生、类别、组份、性质、毒素的生态功能以及毒素的作用机制等方面综述了寄生性膜翅目昆虫毒素的研究概况.膜翅目的泌毒器官起源于外胚层,由生殖系统的附腺演化而来.毒液由成熟雌蜂的毒腺或酸腺所分泌,并贮于毒囊中.昆虫毒素是成分复杂的混合物,已知膜翅目昆虫毒素中含有烃类、醇类、醛类、酮类、羧酸类、酯类、内酯类、酶类等多种化合物.寄生性膜翅目昆虫的毒素在提高自身适应能力方面的作用是巨大的,如通过麻痹寄主提高产卵成功的概率、通过抑制寄主的生长发育和免疫功能提高后代的存活率、通过干扰寄主的生理活动改善后代的营养需求等.体外寄生蜂毒素可造成寄主幼虫停止发育、永久性的麻痹甚至死亡,这类毒素常为抑性的、广谱的,一般作用于中枢神经系统或神经-肌肉连接点.而体内寄生蜂多为容性寄生,其毒液中含有多分DNA病毒（PDV）,PDV通过抑制寄主免疫系统而巧妙地调节寄主的生理活动和发育,影响寄主的正常变态,大多数种类直到寄主结茧或做好蛹室时才将其杀死在安全的场所,从而使寄生蜂后代能够顺利完成发育.容性寄生蜂毒素对PDV的功能具有显著的增效或协同作用,而不会使寄主产生永久性麻痹.PDV对寄生蜂本身是非致病性的,与寄生蜂是一种分子水平上的共生或依生关系.寄生性膜翅目昆虫毒素显示了良好的应用前景,特别是在开发人类医药和特异性生物杀虫剂方面.但分离和纯化毒液中各个活性成分是应用的前提,也是生化和毒理研究的需要.     

抗菌肽抗菌机制及其应用研究进展

抗菌肽是一类小分子肽,具有广谱的抗菌活性。以往对抗菌肽抗菌机制的研究主要集中在细菌细胞膜的作用上,包含桶板模型、毯式模型,环形孔模型和凝聚模型。近年来相继发现某些抗菌肽可以作用于细菌细胞内部,与核酸物质结合,阻断DNA复制、RNA合成;影响蛋白质合成;抑制隔膜、细胞壁合成,阻碍细胞分裂;抑制胞内酶的活性。本文从胞内机制和胞外机制两个角度对抗菌肽的抗菌机制进行综述,以期阐明各类抗菌肽的作用机制,为进一步研究菌株耐药性、杀菌效果及其杀菌机制提供科学根据。     

Understanding molecular mechanisms for improving phytoremediation of heavy metal-contaminated soils

Heavy metal pollution of soil is a significant environmental problem with a negative potential impact on human health and agriculture. Rhizosphere, as an important interface of soil and plants, plays a significant role in phytoremediation of contaminated soil by heavy metals, in which, microbial populations are known to affect heavy metal mobility and availability to the plant through release of chelating agents, acidification, phosphate solubilization and redox changes, and therefore, have potential to enhance phytoremediation processes. Phytoremediation strategies with appropriate heavy metal-adapted rhizobacteria or mycorrhizas have received more and more attention. In addition, some plants possess a range of potential mechanisms that may be involved in the detoxification of heavy metals, and they manage to survive under metal stresses. High tolerance to heavy metal toxicity could rely either on reduced uptake or increased plant internal sequestration, which is manifested by an interaction between a genotype and its environment.A coordinated network of molecular processes provides plants with multiple metal-detoxifying mechanisms and repair capabilities. The growing application of molecular genetic technologies has led to an increased understanding of mechanisms of heavy metal tolerance/accumulation in plants and, subsequently, many transgenic plants with increased heavy metal resistance, as well as increased uptake of heavy metals, have been developed for the purpose of phytoremediation. This article reviews advantages, possible mechanisms, current status and future direction of phytoremediation for heavy-metal–contaminated soils.     

Determination of optimal rehydration,fixation and staining methods for histological and immunohistochemical analysis of mummified soft tissues

During an excavation headed by the German Institute for Archaeology, Cairo, at the tombs of the nobles in Thebes-West, Upper Egypt, three types of tissues from different mummies were sampled to compare 13 well known rehydration methods for mummified tissue with three newly developed methods. Furthermore, three fixatives were tested with each of the rehydration fluids. Meniscus (fibrocartilage), skin, and a placenta were used for this study. The rehydration and fixation procedures were uniform for all methods. The stains used were standard hematoxylin and eosin, elastica van Gieson, periodic acid-Schiff, and Grocott, and five commercially obtained immunohistochemical stains including pancytokeratin, vimentin, alpha-smooth-muscle-actin, basement membrane collagen type IV, and S-100 protein. The sections were examined by transmitted light microscopy. Our study showed that preservation of the tissue is dependent on the quality and effectiveness of the combination of the rehydration and fixation solutions, and that the quality of the histological and histochemical stains is dependent on the tissue quality. In addition, preservation of the antigens in the tissues is dependent on tissue quality, and fungal permeation had no influence on the tissue. Finally, the results are tissue specific. For placenta the best solution combination was Sandison and solution III (both fixed with formaldehyde) while results for skin were best with Ruffer I (using formaldehyde and Schaffer as fixatives), Grupe et al. (using formaldehyde as a fixative) and solution III (in combination with formaldehyde and Bouin fixatives). Ruffer II (using formaldehyde as a fixative) and solution III (in combination with Schaffer fixative) gave the best results for fibrocartilage.     

Yeast killer toxins,molecular mechanisms of their action and their applications

Killer toxins secreted by some yeast strains are the proteins that kill sensitive cells of the same or related yeast genera. In recent years, many new yeast species have been found to be able to produce killer toxins against the pathogenic yeasts, especially Candida albicans. Some of the killer toxins have been purified and characterized, and the genes encoding the killer toxins have been cloned and characterized. Many new targets including different components of cell wall, plasma membrane, tRNA, DNA and others in the sensitive cells for the killer toxin action have been identified so that the new molecular mechanisms of action have been elucidated. However, it is still unknown how some of the newly discovered killer toxins kill the sensitive cells. Studies on the killer phenomenon in yeasts have provided valuable insights into a number of fundamental aspects of eukaryotic cell biology and interactions of different eukaryotic cells. Elucidation of the molecular mechanisms of their action will be helpful to develop the strategies to fight more and more harmful yeasts.     

Determination of optimal rehydration, fixation and staining methods for histological and immunohistochemical analysis of mummified soft tissues.

During an excavation headed by the German Institute for Archaeology, Cairo, at the tombs of the nobles in Thebes-West, Upper Egypt, three types of tissues from different mummies were sampled to compare 13 well known rehydration methods for mummified tissue with three newly developed methods. Furthermore, three fixatives were tested with each of the rehydration fluids. Meniscus (fibrocartilage), skin, and a placenta were used for this study. The rehydration and fixation procedures were uniform for all methods. The stains used were standard hematoxylin and eosin, elastica van Gieson, periodic acid-Schiff, and Grocott, and five commercially obtained immunohistochemical stains including pancytokeratin, vimentin, alpha-smooth-muscle-actin, basement membrane collagen type IV, and S-100 protein. The sections were examined by transmitted light microscopy. Our study showed that preservation of the tissue is dependent on the quality and effectiveness of the combination of the rehydration and fixation solutions, and that the quality of the histological and histochemical stains is dependent on the tissue quality. In addition, preservation of the antigens in the tissues is dependent on tissue quality, and fungal permeation had no influence on the tissue. Finally, the results are tissue specific. For placenta the best solution combination was Sandison and solution III (both fixed with formaldehyde) while results for skin were best with Ruffer I (using formaldehyde and Schaffer as fixatives), Grupe et al. (using formaldehyde as a fixative) and solution III (in combination with formaldehyde and Bouin fixatives). Ruffer II (using formaldehyde as a fixative) and solution III (in combination with Schaffer fixative) gave the best results for fibrocartilage.     

Protein array staining methods for undefined protein content, manufacturing quality control, and performance validation

Methods to assess the quality and performance of protein microarrays fabricated from undefined protein content are required to elucidate slide-to-slide variability and interpolate resulting signal intensity values after an interaction assay. We therefore developed several simple total- and posttranslational modification-specific, on-chip staining methods to quantitatively assess the quality of gel element protein arrays manufactured with whole-cell lysate in vitro protein fractions derived from two-dimensional liquid-phase fractionation (PF2D) technology. A linear dynamic range of at least 3 logs was observed for protein stains and immobilized protein content, with a lower limit of detection at 8 pg of protein per gel element with Deep Purple protein stain and a field-portable microarray imager. Data demonstrate the successful isolation, separation, transfer, and immobilization of putative transmembrane proteins from Yersinia pestis KIM D27 with the combined PF2D and gel element array method. Internal bovine serum albumin standard curves provided a method to assess on-chip PF2D transfer and quantify total protein immobilized per gel element. The basic PF2D array fabrication and quality assurance/quality control methods described here therefore provide a standard operating procedure and basis for developing whole-proteome arrays for interrogating host-pathogen interactions, independent of sequenced genomes, affinity tags, or a priori knowledge of target cell composition.     

A comparison of survey and focal follow methods for estimating individual activity budgets of cetaceans

Activity budget data are essential for determining behavioral responses to physiological and ecological variables. Yet, few studies are available to investigate the robustness, accuracy, and biases of the methods used to estimate activity budgets for cetaceans. In this study, we compare activity budgets of 55 adult female bottlenose dolphins in Shark Bay, Australia derived from two methods: surveys (n = 6,903) and focal follows (n = 1,185, totaling 2,721 h of observation). Activity budgets estimated from survey data differed in all behavioral states compared to focal follow data. However, when controlling for temporal autocorrelation, only time spent socializing and time spent traveling remained disparate between the methods. To control for biases associated with assigning group‐level behavior to individuals, we also compared survey and focal follow activity budgets for lone females. Here we found differences between methods in time spent foraging and traveling regardless of whether we controlled for temporal autocorrelation, which suggests detection biases likely play a role in explaining differences in activity budget estimates between the two methodologies. Our results suggest that surveys are less representative of individual‐level activity budgets, and thus, when individual‐level knowledge about behavior is needed, focal follows are preferred.     

Comparison of various staining methods for the detection of Cryptosporidium in cell-free culture

The complete development of Cryptosporidium in host cell-free medium first described in 2004, represented a significant advance that can facilitate many aspects of Cryptosporidium research. A current limitation of host cell-free cultivation is the difficulty involved in visualising the life-cycle stages as they are very small in size, morphologically difficult to identify and dispersed throughout the media. This is in contrast to conventional cell culture methods for Cryptosporidium, where it is possible to focus on the host cells and view the foci of infection on the host cells. In the present study, we compared three specific and three non-specific techniques for visualising Cryptosporidium parvum life-cycle stages in cell-free culture; antibody staining using anti-sporozoite and anti-oocyst wall antibodies (Sporo-Glo™ and Crypto Cel), fluorescent in-situ hybridization (FISH) using a Cryptosporidium specific rRNA oligonucleotide probe and the non-specific dyes; Texas Red, carboxyfluorescein diacetate succinimidyl ester (CFSE) and 4,6′ diamino-2-phenylindole dihydrochloride (DAPI). Results revealed that a combination of Sporo-Glo™ and Crypto Cel staining resulted in easy and reliable identification of all life-cycle stages.     

The development and standardization of testing methods for genetically modified organisms and their derived products

As the worldwide commercialization of genetically modified organisms (GMOs) increases and consumers concern the safety of GMOs, many countries and regions are issuing labeling regulations on GMOs and their products. Analytical methods and their standardization for GM ingredients in foods and feed are essential for the implementation of labeling regulations. To date, the GMO testing methods are mainly based on the inserted DNA sequences and newly produced proteins in GMOs. This paper presents an overview of GMO testing methods as well as their standardization.     

抗污损海洋天然产物的开发及其作用机理研究进展

首先对近年发表在学术期刊Biofouling上的一篇关于抗污损化合物的综述做一简短总结。其次,突出介绍了对无脊椎污损生物附着和变态分子水平的调控机制的研究近来的进展。旨在给那些从事生物污损和抗污损技术研究的科研人员提供一定的帮助。     

稻田水生昆虫的采样技术方法与应用

稻田水生昆虫在稻田生态系统功能实现中发挥着重要的作用。本文基于作者对稻田生态系统中水生昆虫多样性研究,系统地介绍了稻田水生昆虫的生态位分布、定量研究与定性研究的采集技术方法与流程,为更好地开展稻田水生昆虫研究提供技术支撑。     

Antiproliferative effects of selective adenosine receptor agonists and antagonists on human lymphocytes: evidence for receptor-independent mechanisms

The effects of standard adenosine receptor (AR) agonists and antagonists on the proliferation of human T lymphocytes, unstimulated and phytohemagglutinin-stimulated human peripheral blood lymphocytes (PBL), and Jurkat T cells were investigated. Real-time PCR measurements confirmed the presence of all four AR subtypes on the investigated cells, although at different expression levels. A_2A ARs were predominantly expressed in PBL and further upregulated upon stimulation, while malignant Jurkat T cells showed high expression levels of A₁, A_2A, and A_2B ARs. Cell proliferation was measured by [³H]-thymidine incorporation assays. Several ligands, including the subtype-selective agonists CPA (A₁), BAY60-6583 (A_2B), and IB-MECA (A₃), and the antagonists PSB-36 (A₁), MSX-2 (A_2A), and PSB-10 (A₃) significantly inhibited cell proliferation at micromolar concentrations, which were about three orders of magnitude higher than their AR affinities. In contrast, further investigated AR ligands, including the agonists NECA (nonselective) and CGS21680 (A_2A), and the antagonists preladenant (SCH-420814, A_2A), PSB-1115 (A_2B), and PSB-603 (A_2B) showed no or only minor effects on lymphocyte proliferation. The anti-proliferative effects of the AR agonists could not be blocked by the corresponding antagonists. The non-selective AR antagonist caffeine stimulated phytohemagglutinin-activated PBL with an EC₅₀ value of 104 μM. This is the first study to compare a complete set of commonly used AR ligands for all subtypes on lymphocyte proliferation. Our results strongly suggest that these compounds induce an inhibition of lymphocyte proliferation and cell death through AR-independent mechanisms.     

Molecular analysis of Hsp70 mechanisms in plants and their function in response to stress

Studying the strategies of improving abiotic stress tolerance is quite imperative and research under this field will increase our understanding of response mechanisms to abiotic stress such as heat. The Hsp70 is an essential regulator of protein having the tendency to maintain internal cell stability like proper folding protein and breakdown of unfolded proteins. Hsp70 holds together protein substrates to help in movement, regulation, and prevent aggregation under physical and or chemical pressure. However, this review reports the molecular mechanism of heat shock protein 70 kDa (Hsp70) action and its structural and functional analysis, research progress on the interaction of Hsp70 with other proteins and their interaction mechanisms as well as the involvement of Hsp70 in abiotic stress responses as an adaptive defense mechanism.     

Methods of selective staining of polyacrylamide gels for -SH groups and cadmium

A new and very sensitive procedure for analysis of polyunsaturated phospholipids is described. The method involves resolution of lipids by high-performance thin-layer chromatography, conversion of lipids to a fluorescent derivative, and quantitation by densitometric scanning. The formation of the fluorophore is dependent on the presence of at least two double bonds in an acyl chain. As little as 20 pmol of phosphatidylinositol can be determined with a precision of better than 5%.