细胞功能测定

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 ...

Coated vesicles are implicated in a number of receptor-mediated transport events within eukaryotic cells. In particular, they are required for the export of newly synthesized lysosomal enzymes from the trans-Golgi network, and at the plasma membrane they are responsible for receptor ...

Receptor-mediated endocytosis is an important mechanism by which the cell is able to take in macromolecules from the external environment and carry them through the necessary metabolic processes. Studies of the sequential steps of these events require methods of following the fate of the ...

The plasma membrane (PM) of polarized epithelial cells is composed of two physically continuous, but compositionally and functionally distinct domains: basolateral and apical. In hepatocytes, the basolateral domain includes the sinusoidal front, which faces the space of Disse and ...

The membrane organelles of an organized tissue, such as rat liver, that will always form closed vesicles, irrespective of the mode of homogenization, are the tubular membranes of the smooth and rough endoplasmic reticulum. Almost without exception, these vesicles retain their normal ori ...

Continuous-flow electrophoresis (CFE), which separates particles on the basis of surface charge density, should be regarded as an adjunct to centrifugation rather than as an alternative for membrane fractionation. The equipment is expensive and has been used for relatively specia ...

This fraction is defined broadly as the material that will sediment at about 3500g for 10 min from a postnuclear supernatant. It contains the nuclei that failed to sediment at 1000g for 5–10 min, the largest mitochondria, and very few other organelles, such as lysosomes and peroxisomes.