实验基础

Phosphoinositide 3-kinases (PI 3-kinases) are an evolutionarily conserved family of lipid kinases that have attracted much attention over the past 10 years or so (reviewed in ref. 1). Three PI 3-kinase classes have been defined on the basis of primary structure, regulation, and their in vitro li ...

Interleukin-2 (IL-2) was first discovered and characterized as T-cell growth factor (1). It is responsible for the growth of T cells and thus plays an important role in the proper functioning of the immune system. IL-2 has a high affinity receptor consisting of an α-, β-, and γ-chain. The γ- chain is shared with a n ...

The discovery of green fluorescent protein (GFP) as a naturally fluorescing protein and its subsequent modification to allow easy detection with standard fluorescent equipment such as the fluorescence-activated cell sorter (FACS) has revolutionized our ability to detect the pr ...

An understanding of the structure and function of cytokines requires the availability of milligram to gram amounts of highly purified and biologically active cytokines. A variety of expression systems have been used to produce recombinant proteins, including Escherichia coli, Pi ...

Interleukins are soluble peptide molecules that mediate the interaction between immunocompetent and haematopoietic cells and between the immune and neuroendocrine systems (1). They are produced by a variety of activated cells and exert their biological activities by binding to s ...

The purpose of the gene array techniques is to simultaneously analyze the expression or characteristics of a large number of genes (1–3). These techniques can be used to compare gene expression in various cells or in the same cells following various treatments, to analyze function of multiple ge ...

This chapter describes several methods for recognizing apoptosis in tumor cells following infection with a replication-deficient adenovirus expressing the tumor suppressor gene p53. We include cytotoxicity assays and assays of apoptosis, including DNA-nucleosomal DNA fr ...

Oncolytic adenovirus (Ad) vectors belong to a new class of cancer therapy agents that destroy cancer cells as part of the virus’s lytic infectious cycle. In this chapter we describe an immunocompetent, semi-permissive cotton rat tumor model to evaluate the safety and efficacy of oncolytic Ad v ...

The enteric adenoviruses of subgroup F (Ad40 and Ad41) pose some special problems of cultivation, as they cannot be readily passaged in many of the cell types used to propagate the more commonly used subgroup C serotypes (Ad2 and Ad5) and there is no standard plaque assay. Methods to propagate Ad40 in com ...

Oncolytic adenoviruses (Ads) are promising candidates for cancer therapy. However, current animal models to evaluate these vectors have substantial limitations. Because Ad replication is generally species-specific, oncolytic Ads are usually examined in immunodeficient ...

A critical step in working with adenovirus (Ad) and its vectors is the accurate, reproducible, sensitive, and rapid measurement of the amount of virus present in a stock. Titration methods fall into one of two categories: determination of either the infectious or the particle (infectious plus n ...

Ribozymes are small catalytic RNAs with the ability to reversibly cleave covalent bonds in RNA in the complete absence of protein (1). With the exception of the RNA component of bacterial RNase P, presently characterized ribozymes catalyze a one-time intramolecular self-cleavage in the ...

Ribozymes are recognized as useful tools for the manipulation of genes because of their high specificity and the fact that they act without influencing the expression of genes that are unrelated targets (1,2). To date, many successful experiments with intracellular ribozymes have been re ...

Hammerhead ribozymes are short catalytic RNA molecules possessing endoribonuclease activity. The enzyme’s specificity is mediated by variable flanking sequences complementary to the RNA target substrate (1,2). Cleavage occurs via transesterification 3′ to a nucleotide tr ...

Influenza A virus infection causes acute respiratory disease in humans and in various animal species, notably pigs, horses, seals, as well as chicken and water fowl. In this study, we present evidence that the multiplication of the RNA virus Influenza A can be inhibited by intracellular express ...

Bovine leukemia virus (BLV) is a retrovirus structurally related to human T-cell leukemia virus-I and -II and simian T-cell leukemia virus. It infects principally B-lymphocytes (1), causing a persistent expansion of B-cells, termed persistent lymphocytosis, in approx 30% of infected ca ...

Lyssavirus infection, better known as rabies, in animals or in humans, is characterized by a clinical picture of viral encephalitis. According to the WHO estimations, it is responsible for more than 50,000 human deaths each year. The diagnosis of lyssavirus infection in animals, which may be res ...

Measles, mumps, and rubella (MMR) virus infections are common during childhood throughout the world. Measles and mumps viruses belong to the Paramyxoviridae family with an RNA genome of negative polarity and a simrlar overall viral structure at the molecular level (1,2). Rubella virus is a me ...

Parvovirus B 19 was discovered at the Virus Reference Laboratory in 1975 by Cossart and colleagues (1). First found in healthy blood donors, parvovirus B19 infection usually manifests as erythema infectiosum (fifth disease) in children. In adults the infection can cause acute arthritis. T ...

Both epidemiological and experimental evidence suggests sexual activity contributes to the etiology of cervical cancer. Searches for a responsible agent have pointed to an association with human papillomavirus (HPV). These viruses are nonenveloped, approx 55 nm in diameter, con ...