实验仪器使用技术

Transfection of specific elements of DNA into cultured mammalian cells allows for the analysis of a range of functional and toxicologic mechanisms. At the heart of this technique is the ability to promote the uptake of DNA into actively growing cells, and to detect and analyze the expression of the g ...

Transcription factors are induced in neurons to alter gene expression to adapt to brain injury caused by exposure to neurotoxins, neurodegeneration disease, or mechanical damage. These DNA-binding proteins bind to specific recognition sequences in the promoter region of genes to re ...

Over the course of an organism’s life, cells divide, grow, differentiate, and die. For many years cell death has been recognized as significant in normal neuronal development. More recently, interest has grown in the mechanisms that regulate both cell death and cell survival in neurons during ho ...

The patch-clamp technique has revolutionized the understanding of ion channel physiology (Hamill et al., 1981; Neher and Sakmann, 1976). This technique facilitates the measurement of currents and voltage in cells under voltage-clamp and current-clamp conditions. Cells can be patch ...

One of the basic tenets of experimental science has been the search for techniques that give relevant information from intact systems while causing minimal effects. This concept is particularly important in the physiological branch of the biological sciences. In general, reduction ...

This chapter describes methods for the study of ion channels and transporters by recording from membrane macropatches. While investigators have made use of many different cell types for such experiments, we focus here on studies of these proteins expressed exogenously in Xenopus oocyt ...

One of the main challenges for a better understanding of signaling mechanisms in the normal and diseased central nervous system (CNS) is to unravel the molecular basics of function on the cellular and systemic levels. Several important cellular processes, such as proliferation, differe ...

The technique of patch clamping can be seen in retrospect as a combination of two separate lines of development that both originated in the 1960s and 1970s. The classical biophysics of the nerve impulse had by then been established in the squid giant axon using a combination of (1) voltage clamping with ...

Patch clamping facilitates directly controlling the voltage of a cell and thus has been acknowledged to be the gold standard for assessing ion-channel physiology in academic and industrial laboratories. The method was developed 30 years ago by (1976), and its various different configur ...

The extracellular patch voltage clamp technique has facilitated studying the currents through single ionic channels from a wide variety of cells. In its early form (Neher and Sakmann, 1976), the resolution of this technique was limited by the relatively low resistances, which isolated the ...

The patch-clamp recording technique measures ionic currents under a voltage clamp and was designed to study small patches of membrane in which near-perfect control of the transmembrane voltage can be readily achieved. Today, this technique is most frequently used to examine currents ac ...

The ability to record the real-time movement of a protein is quite enthralling; in effect, this is what we achieve when we record the activity of an individual ion channel. Clearly we do not “see” the amino acids that make up the ion channel undergo the conformational changes that must occur, but we observe the ...

Combined electrophysiological and fluorometric recordings have proven to be a powerful tool, especially in the field of neurobiology. With this combination it became possible to overcome three major limitations of pure electrophysiological measurements. First, high-reso ...

Patch-clamp recording is a powerful approach to monitoring membrane electrical activity with high temporal resolution. However, the spatial resolution of patch-clamp recording in a distributed structure such as a neuron or a brain slice is limited by the fact that each electrode records ...

Brain slices have become an integral part of synaptic and cellular physiology since the pioneering studies of Henry McIlwain (Li and McIlwain, 1957; Yamamoto and McIlwain, 1966). The hippocampal slice preparation was first brought to the United States from Per Anderson’s lab, initially by T ...

The patch-clamp technique, introduced by Erwin Neher and Bert Sakmann (Neher and Sakmann, 1976) in the mid-1970s facilitated recordings of currents through single-ion channels in living cells, advancing the ability to study the membrane function of excitable cells, such as neurons. Ion ...

The whole-cell patch-clamp recording technique (Marty and Neher, 1995) is nowadays a standard method for studying electrophysiological properties of the cellular membranes and synaptic inputs. This technique has been applied mainly to in vitro preparations such as culture cells, d ...

“The determination of the three-dimensional structures of protein molecules showed for the first time in detail the construction of the molecular ‘machines’ of the life cycle. If we want to learn how these ‘machines’ work, it is not sufficient only to know their construction. We actually have to see ...

The development of molecular cloning techniques over the past two decades has generated a powerful arsenal of tools for the study of gene structure and function. It is now possible to synthesize complementary DNA (cDNA) copies of mRNAs from virtually any source and to propagate these cDNAs in vit ...

In vitro translation systems are extremely important tools for studying protein biosynthesis. These systems have been most useful in revealing the intracellular sites of synthesis of numerous proteins, the nature of cotranslational proteolytic cleavage events, the process of c ...