生物信息学技术

Dielectrophoresis is a phenomenon which can be exploited to provide significant quantitative electrophysiological data in a range of biochemical setting, from oncology to drug discovery. This chapter seeks to elucidate those applications and the electrophysiological phen ...

Phospho-specific flow cytometry, or phospho flow, measures the phosphorylation state of intracellular proteins at the single cell level. Many phosphorylation events can be analyzed simultaneously in each cell, along with cell surface markers, enabling complex biochemical sig ...

Intracellular cytokine staining (ICS) is a popular method for visualizing cellular responses, most often T-cell responses to antigenic or mitogenic stimulation. It can be coupled with staining for other functional markers, such as upregulation of CD107 or CD154, as well as phenotypic ma ...

In the second edition of this series, we described the use of cell tracking dyes in combination with tetramer reagents and traditional phenotyping protocols to monitor levels of proliferation and cytokine production in antigen-specific CD8+ T cells. In particular, we illustrated how tr ...

The biotechnologist faced with the challenge of producing significant quantities of recombinant protein will endeavor to identify a host cell for protein production that will synthesize authentic, biologically active proteins at relatively high yields. Due to the complex foldi ...

Commercially exploitable compounds are being produced using modern biotechnology for use as food additives, chemotherapeutic agents, and pesticides. Traditionally, animal testing has always played an important role in the safety evaluation of such agents. However, financial ...

Glycosyl-phosphatidylinositol (GPI)-anchored proteins bound to the outer surface of cell membranes (1,2) can be selectively released in a soluble form by the action of a highly specific bacterial enzyme, phosphatidylinositol-phospholipase C (PI-PLC) (3,4). This chapter descri ...

Although most integral membrane proteins are bound to the lipid bilayer by a hydrophobic polypeptide transmembrane domain, a small functionally diverse group of proteins is uniquely anchored to the plasma membrane by the covalent attachment of a complex phospolipid anchor to the carb ...

Developing protein-based therapeutic agents requires both product quality and consistency to be maintained throughout the development and implementation of the production process. Differences in host cell type, the physiological status of the cell, and protein structural co ...

Heterogeneity of cell culture-produced glycoproteins often results from the presence or absence of a few sugars found on the terminus of glycoprotein oligosaccharides. Variability in bioprocess factors can potentially lead to variability in this oligosaccharide heterogen ...

The Enzyme-Linked Immunosorbent Assay (ELISA) and Western Blot analysis have been workhorse techniques for the analysis of protein levels and state, such as phosphorylation. The ELISA is also useful for measuring the affinity of a molecule for its ligand. The disadvantage of these techni ...

In flow cytometry, the quantitation of fluorophore-tagged ligands and receptors on cells or at particulate surfaces is achieved by the use of standard beads of known calibration. To the best of our knowledge, only those calibration beads based on fluorescein, EGFP, phycoerythyrin and allo ...

Multicolor flow cytometer assays are routinely used in clinical laboratories for immunophenotyping, �monitoring disease and treatment, and determining prognostic factors. However, existing methods for quantitative measurements have not yet produced satisfactory re ...

Recent advances in biotechnology have resulted in cytometers capable of performing numerous correlated measurements of cells, often exceeding ten. In the near future, it is likely that this number will increase by fivefold and perhaps even higher. Traditional analysis strategies ba ...

Initial approaches to cytometry for classifying and characterizing cells were based on microscopy; it was necessary to collect relatively high-resolution images of cells because only a few specific reagents usable for cell identification were available. Although flow cytomet ...

Over the past decade, there has been a rapid growth in the number of BSL-3 and BSL-4 laboratories in the USA and an increase in demand for infectious cell sorting in BSL-3 laboratories. In 2007, the International Society for Advancement of Cytometry (ISAC) Biosafety Committee published standards ...

In vivo biotinylation tagging, based on a method in which a protein of interest is tagged with a peptide that is biotinylated in vivo by coexpression of Escherichia coli BirA biotin ligase, has been successfully used for the isolation of protein–protein and protein–DNA complexes in mammalian c ...

Flow cytometry has been employed for the analysis of higher plants for approximately the last 30 years. For the angiosperms, ∼500,000 species, itself a daunting number, parametric measurements enabled through the use of flow cytometers started with basic descriptors of the individual c ...

A flow cytometer is an instrument that illuminates cells (or other particles) as they flow individually in front of a light source and then detects and correlates the signals from those cells that result from the illumination. In this chapter, each of the aspects of that definition will be described: ...

Cell type-specific expression of fluorescent proteins allows the purification of rare cells from complex �tissues by flow cytometry. This strategy is especially useful for molecular analysis of cancer cells because these cells can be effectively purified away from the noncancero ...