微生物学技术

Natural-killer (NK) cells are lymphocytes that do not express the CD3 T-cell receptor but do express the CD16 (FcγRIII) and CD56 (isoform of NCAM) in humans or NK1.1 antigen in certain strains of mice. NK cells display spontaneous lytic activity but do not exhibit immunological memory. NK cells are im ...

Natural-killer (NK) cells play an important role in recognizing and eliminating virally infected and transformed cells. To study this process, convenient assays for NK-cell function are required. Conventional NK-cell activity assays measure the release of 51Cr from prelabeled ta ...

This chapter provides a protocol for the large-scale purification of adenovirus type 2 and 5 virions and the soluble major coat protein hexon. The purified virus particles remain intact and are suitable for vector, vaccine, or structural studies and can also be used as seed stock for further rounds ...

All adenoviruses (Ads) sequenced so far encode a single endopeptidase of the cysteine class, named adenain. The Ad2 adenain is a 204-residue, nearly inactive monomer, which is activated during virus maturation by an 11-residue cleavage fragment of capsid protein pVI. This chapter describ ...

This method is designed to measure rates of transcription from adenoviral promoters as a function of the concentrations within infected cells of the promoter(s) of interest. The latter parameter is assessed by quantification of viral DNA by hybridization of membrane-bound DNA followi ...

Entry of the adenovirus (Ad) capsids during the early stages of infection is a multistep process that includes initial attachment of the virus capsid to the cell surface followed by internalization of the virus into early endosomes. The Ad fiber protein, a complex of three apparently identical ...

Members of the family Adenoviridae have been isolated from a large variety of hosts, including representatives from every major vertebrate class from fish to mammals. The high prevalence, together with the fairly conserved organization of the central part of their genomes, make the adeno ...

Adenoviruses exhibit considerable intraserotypic genetic variability. Restriction enzyme analysis of the adenoviral genome is currently the most widely used procedure for the characterization of adenovirus isolates and has been extensively used for molecular epidemi ...

Adenoviruses (Ads), like other DNA tumor viruses, have evolved specific regulatory genes that facilitate virus replication by controlling the transcription of other viral genes as well as that of key cellular genes. In this regard, the E1A transcription unit contains multiple protein d ...

Here we describe a collection of methods that have been adapted to produce highly efficient nuclear and cytoplasmic extracts from adenovirus-infected HeLa cells. We describe how to produce extracts from virus-infected cells and how to analyze RNA splicing in vitro using T7 RNA polymerase ...

ELISAs offer excellent specificity and, once fully optimized, sensitivity that rivals that of bioassays. The major variables that need to be experimentally determined when developing an ELISA are the optimal number of fresh cells required per well, the optimal antigen concentrations ...

Chemokines are primarily low molecular mass proteins that are produced and usually released by a wide variety of cell types. Differential chemokine responses can be excellent early markers of immune dysfunction, allowing clinical intervention prior to expression of full blown unde ...

Immunoglobulins are a heterogeneous group of proteins. It naturally follows that the strategies for purifying them are diverse and numerous. A good knowledge of their respective physiochemical properties will obviously make the task easier. The choice between using polyclonal and ...

Specific allergen immunotherapy is an effective treatment for IgE-mediated allergic disease and involves T- and B-cell mediated events. IgE receptors on the surface of antigen-presenting cells facilitate the presentation of allergens in the presence of specific IgE antibody res ...

The genus Helicobacter currently comprises over 15 different species with members described as microaerophilic gram-negative spiral rods with a G + C content of 35–38%. The most medically important Helicobacter is Helicobacter pylori, a human pathogen that has been causally linked to t ...

Lactococcus lactis strains are the principal organisms used in cheese-making, and their performance is of vital importance to the quality of the cheese. A goal for the dairy industry has always been to improve strains of L. lactis and to stabilize beneficial traits. In recent years, genetic engin ...

The streptococci are a diverse genus of bacteria consisting of commensal and pathogenic organisms of human and veterinary origin, as well as industrially important species. Although many of the species are naturally transformable, several species do not enter a competent state. The abi ...

Diversity—the variability carried by the amino acid sequences of a synthetic antibody library—can be generated by synthetic degenerate oligonucleotides. One can experiment with different diversity designs in the variable domains of light and heavy chains (VH and VL) to generate ant ...

Antibody-based immunologic reagents are useful for identifying, isolating, or eliminating cells with particular characteristics related to different diseases. Phage display is a highly valuable technique for antibody selection related to this purpose. In brief, a diverse gro ...

Recently there has been an increased use of microarray technology as a tool to determine the presence of functional genes in a population of hard to culture communities (e.g., soil, extreme environments). A functional gene array (FGA), or GeoChip, uses probes to screen for specific functional ge ...