细胞技术

In the 20 yr since the discovery of proteins whose levels oscillate during the cell cycle in marine invertebrate embryos (1), the study of cyclins and their cognate protein kinases has revealed a wealth of information on how eukaryotic cells control cyclical functions connected with cell prol ...

It is becoming apparent that an increase in the rate of protein degradation represents one mechanism for the regulation of key proteins during the cell cycle (1). The importance of protein degradation in the control of cell proliferation is elegantly illustrated by the regulated synthesis a ...

Immunofluorescence is one of the most widely used techniques to study the localization of transcription factors, proteins, and structural components of nuclear architecture and cytoarchitecture. High-resolution in situ immunofluorescence approaches permit assessme ...

This technique is used to visualize sites of active transcription in a permeabilized cell and does not require radiolabeled molecules (e.g., Uridine). Nonradioactive ribonucleic acid (RNA) precursors (e.g., 5-bromouridine-5′-triphosphate ) are used and can be detected by using fluo ...

The resolution that can be attained in assessment of the intranuclear localization of cellular proteins is dependent on specificity of the antibodies. Primary antibodies should be well characterized. We recommend testing antibody specificity by immunoblotting. In general, mon ...

Only a handful of tagged molecules have been used successfully in vivo to monitor gene expression. The most commonly used are β-galactosidase (β-gal) and green fluorescent protein (GFP). Both molecules pose limitations with in vivo detection. When using GFP, autofluorescence may be encou ...

The electrophoretic mobility shift assay (EMSA) determines sequence-specific protein-deoxyribonucleic acid (DNA) interactions (e.g., transcription factors with cognate regulatory elements). This assay is based on reduced electrophoretic mobilities of protein/DNA ...

Mapping DNase I hypersensitive sites can often localize the control regions in a eukaryotic gene. It is generally believed that chromosomal regions with loose or more open conformation are sensitive to DNase I cleavage. By comparing the patterns of hypersensitive sites obtained by diges ...

Ligation-mediated polymerase chain reaction (LM-PCR) is used for high-resolution genomic footprinting. This technique was originally developed to study in vivo protein-deoxyribonucleic acid (DNA) interactions at regions of genes important for transcriptional regula ...

Apoptosis, or programmed cell death, refers to both the initiation and execution of the events whereby a cell commits suicide. This process is important in development and its deregulation is found in many diseases (1–6), including cancer (6–10). Apoptosis is distinct from other ways in which ce ...

Malignant tumors are composed of cells that have lost their proliferation control and multiply independently of a physiologic need for increase in cell number in the tissue. Cells that have lost these proliferative controls are often referred to as “transformed.” Transformation of a mam ...

Alkylating agents are a series of potentially carcinogenic compounds that are able to introduce lesions into deoxyribonucleic acid (DNA; ref. 1). For their electrophilic nature, alkylating agents have an affinity for the nucleophilic centers of DNA and are monofunctional (have one re ...

Apoptosis plays a critical role in important biological processes such as morphogenesis, tissue homeostasis, and elimination of genomic damaged or virally infected cells and of self-reactive clones from the immune system (1). Although apoptosis is important during normal develo ...

γ Radiation is an electromagnetic radiation with wavelengths in the range of 0.1–100 pm. Like all forms of electromagnetic radiations, the γ ray has no mass and no charge and interacts with material by colliding with the electrons in the shells of atoms. Irradiation with γ rays is very penetrating, and t ...

The genera of the retroviridae comprise oncoretroviruses and lentiviruses (1). Oncoretroviruses can only infect dividing cells, as they require the breakdown of the nuclear membrane to access the cellular chromosomal deoxyribonucleic acid (DNA; ref. 2). Conversely, lentivirus ...

Ultraviolet (UV) radiation is mutagenic in a wide variety of organisms and is a major source of physical deoxyribonucleic acid (DNA) damage. The mutations caused and the biological consequences of exposing cell cultures to UV light have been extensively studied and have given many insights ...

With this method, antibiotic selection of the infected cells gives rise to a countable number of colonies after 7–10 d (1–4). Other reporter genes are based on green fluorescent protein (GFP; ref. 5) or the Escherichia coli lacZ gene, which encodes for β-galactosidase for X-gal staining (4,6). GFP-ex ...

The assessment of the viral titer is an important step in optimizing and reproducing working conditions for the gene transduction for various mammalian cell types. The assessment of the viral titer is an important parameter to determine the maximum number of target cells that can be infected f ...

Immunocytochemistry is the most diffuse technique to visualize and localize specific biochemical components in cell compartments and tissues. With this method the antigens are tagged by antibodies that can be visualized with appropriate markers attached directly (direct met ...

The fluorescence labeling technique is a method with a high degree of specificity and sensitivity and is often chosen as a tool in the study of protein expression and subcellular compartments (1). Recently, a large number of fluorescent dyes with distinct fluorescence excitation and emiss ...