细胞技术

Human vitamin D receptor (hVDR) belongs to the superfamily of steroid receptors. The receptors are nuclear transcription factors that regulate gene expression in response to binding of their specific ligands. According to present knowledge, the molecular mechanism of vitamin D acti ...

The green fluorescent protein (GFP) from the jellyfish, Aequoria victoria, converts blue light to green fluorescence when expressed in intact cells and transgenic animals, and has proven to be a powerful tool for biological and medical research. This chapter describes the application of ...

The development of techniques that allow defined alterations of the mammalian genome have dramatically increased the possibilities for elucidating functions of specific genes in the context of a whole organism. Although some of these techniques have been applied to several mammal ...

The yeast two-hybrid system, originally developed by Fields and Song (1), is a sensitive genetic assay for the detection of protein-protein interactions. The system exploits the fact that eukaryotic transcriptional activators contain separable functional domains for DNA-bind ...

Nuclear hormone receptors belong to a large family of structurally related proteins that include the steroid, retinoic acid, and vitamin D receptors. These receptors function as ligand-activated transcription factors and regulate complex programs of gene expression involved in ...

Measurement of the intracellular Ca2+ concentration (i) in populations of cells is an excellent tool to complement population measurements of other cell parameters, but the usefulness of this approach is limited by several problems, not the least of which is temporal averaging. In popula ...

Many commercial organizations currently use the Fluorometric Imaging Plate Reader (FLIPR�: Molecular Devices, Sunnyvale, CA) to conduct highthroughput measurements of intracellular Ca2+ concentration (see Chapter 7 ), taking advantage of its rapid kinetics, reliability, a ...

The Fluorometric Imaging Plate Reader (FLIPR�; Molecular Devices, Sunnyvale, CA; see Fig. 1) has made a significant contribution to drug discovery programs in the pharmaceutical industry since the first commercial instruments were introduced 9 yr ago (1). The key advantage of FLIPR over co ...

Voltage-sensitive Ca2+ channels (VSCC) play a central role in an extensive array of physiological processes. Their importance in cellular function arises from their ability both to sense membrane voltage and to conduct Ca2+ ions, two facets that couple membrane excitability to a key intra ...

Hormones, neurotransmitters, chemoattractants, and growth factors all elicit intracellular responses on binding to cell surface receptors by activating inositol phospholipid-specific phospholipase C (PLC). Activated PLC catalyzes the hydrolysis of phosphatidyli ...

In general, the action of intracellular second messengers, 3′:5′-cyclic adenosine monophosphate (cAMP) and 3′:5′-cyclic guanosine monophosphate (cGMP), are terminated by cyclic nucleotide phosphodiesterase (PDE). In most mammalian tissues, PDE exists in multiple forms that di ...

The specificity, homogeneity, and availability of large-batch production of liposomes with natural lipids and synthetic lipids have made them an extremely useful tool for the study of diverse cellular phenomena, as well as in medical applications. In many cases, however, the success of the ...

Several investigations, carried out in either artificial or cellular models and using a variety of techniques (1–3), confirmed the prediction of Singer and Nicholson (4) about the presence of domains in biological membranes, that is, of zones where the concentration of the components and the ...

Liposomes are synthetic mimics of cellular membranes and represent an experimental system widely used for more than 30 yr in the field of biochemical research involving lipids. Liposomes typify a hallmark in the reductionist’s approach to biology: by controlling the nature and mole frac ...

Current experimental evidence suggests that the merging of two closely apposed lipid bilayers to form one continuum is mediated by specific fusion proteins. The dissection of the molecular pathways eventually leading to membrane merging can be accomplished by various approaches, i ...

In 1990, the first federally approved clinical trials for treatment of a genetic disorder by gene therapy began under the leadership of R. Michael Blease, W. French Anderson, and their colleagues at the National Institutes of Health. This technique involves the identification of required DNA ...

Enveloped animal viruses infect host cells by fusion of viral and target membranes. This crucial fusion event occurs either with the plasma membrane of the host cells or with the endosomal membranes (1). Fusion is triggered by specific glycoproteins in the virus membrane (2) and involves a range of ...

The isolation of photosynthetically active chloroplasts is the starting point for many plant metabolic studies as diverse as carbon assimilation, electron flow and phosphorylation, metabolite transport, and protein targeting. Whatever the subsequent use of the chloroplast ...

Perhaps the most striking feature of bacterial membranes is their multifunctional nature (1,2). Bacterial cytoplasmic membranes, for example, catalyze the reactions of respiratory and photosynthetic electron transfer and associated energy transduction, and contain nume ...

The isolation of membranes from cultured mammalian cells poses a number of problems. First and foremost is the problem of homogenization, which is particularly difficult with suspension culture cells, as opposed to those that grow as an adherent monolayer. The choice of subcellular fract ...