细胞功能测定

The basement membrane that separates an epithelium or other parenchymal tissues from the connective tissue has been postulated for a long time to be of epithelial origin. Because of difficulties in interpreting results obtained mostly by autoradiographic labeling at the light micro ...

Understanding the function of ECM and cell-matrix interactions in mammalian development has reached new levels of sophistication with the introduction of gene knockout technology. Indeed, two of the chapters in this volume provide detailed methods for producing mice with deletio ...

The polyamines putrescine, spermidine, and spermine are essential for mammalian cell growth, �differentiation, and cell death and have important physiological roles in all tissues. Many of the properties of polyamines that can be demonstrated in vitro are common to all three molecules ...

Immunofluorescence (IF) microscopy is indisputably a key tool to study the location of proteins and to visualize intracellular structures in immobilized cells and in spread nuclei. In yeasts, much has been learned using fluorescent tags as labels employing the original methods descr ...

The observation of damage to chromosomes and of alterations to normal cell cycle progression were early findings in radiation biology (1–6) and provided a strong impetus for the elaboration of the causative basis for understanding deoxyribonucleic acid (DNA) repair in all its manifest ...

Two-dimensional gel electrophoresis (2DE) is a key analytical method for investigating bacterial �proteomes. The relatively simple genomes of many bacteria combined with only limited post-�translational modifications of bacterial proteins mean that a significant propo ...

The image analysis part of gel-based proteome research plays an important role in the overall success of the experiment. The main purpose of software-assisted 2DE gel analysis is to detect the protein spots, match them between gels within an experiment, and identify any differences in protein ...

Two-dimensional electrophoresis (2DE) immunoblotting combines the resolving power of 2DE and the selectivity of antibodies, allowing the selection of individual or sets of proteins from the total proteome. It is essential when immunoblotting membranes that reproducible 2DE ge ...

Chemical modification reactions play an important role in various protocols for mass-spectrometry-based proteome analysis; this applies to both gel-based and gel-free proteomics workflows. In combination with two-dimensional gel electrophoresis (2DE), the addition of “t ...

The proteome of the cell is at the frontier of being too complex for proteomic analysis. Organelles provide a step up. Organelles compartmentalize the cell enabling a proteome, physiology and metabolism analysis in time and in space. Protein complexes separated by electrophoresis have be ...

The degree of protein diversity and dynamic range within organisms means that even the simplest proteome cannot be captured by any single extraction and separation step. New techniques have focused on major protein classes often under-represented in proteome analysis; low abundance, ...

This chapter describes the technical improvements of the two-dimensional electrophoresis pattern resulting of an optimized pH range in the first dimension. Various types of pH gradients are available. Different strategies can be applied in order to select the pH ranges for the explora ...

With the development of the Internet, a growing number of two-dimensional electrophoresis (2DE) databases have become available (60 in 2009, for a total of 425 image maps). By linking the two components constituting 2DE databases, gel images and protein information, the active hypertext li ...

Protein identification is a key aspect in the investigation of proteomes. Typically, in a 2-DE gel-based proteomics analysis, the spots are enzymatically digested and the resulting peptide masses are measured, producing mass spectra. Peptides can also be isolated and fragmented with ...

Identification and characterization of proteins are ultimately the goal in proteomic analysis. In order to identify a protein trypsin is commonly used to digest protein into peptides which can be analyzed by matrix-assisted laser desorption ionization mass spectrometry (MALDI- ...

In this protocol, we describe an approach in which two-dimensional electrophoresis (2DE)-separated proteins are guanidinated in-gel prior to enzymatic cleavage. In contrast to previously described techniques, this procedure allows the extracted tryptic peptides to be N-term ...

Two-dimensional electrophoresis (2DE) is an excellent technology for the analysis of complex protein mixtures, but it has drawbacks, such as hardly detecting very hydrophobic proteins. Shotgun protein analysis is one of the major technologies used to compensate for the weaknesses of ...

The Drosophila embryonic ventral epidermis has served as a unique tissue for the genetic analysis of patterning. Two types of epidermal cells are easily distinguished: those that secrete short, thick hair-like structures called denticles and cells that only secrete smooth cuticle. De ...

The site-specific recombinase FLP is used in Drosophila to precisely manipulate the genome, in particular, to eliminate gene function by mitotic recombination and to activate transgenes in discrete populations of cells. These approaches are already part of the standard tool kit for stu ...

With the live imaging of embryos, the dynamics of developmental processes, such as tissue remodeling, cell morphogenesis, and, campus protein dynamics can be observed and quantified. This has greatly improved the mechanistic understanding of biological processes. Here we describe ...