Showing papers in "Methods in 2003"

TL;DR: The print-tip loess normalization as mentioned in this paper is a well-tested general purpose normalization method which has given good results on a wide range of arrays and can be refined by using quality weights for individual spots.

TL;DR: A collection of protocols to culture MCF-10A cells, to establish stable pools expressing a gene of interest via retroviral infection, as well as to grow and analyzeMCF- 10A cells in three-dimensional basement membrane culture are provided.

TL;DR: A library of bacterial strains corresponding to roughly 86% of the estimated 19,000 predicted genes in C. elegans is constructed, and this library is used to perform genome-wide analyses of gene function.

TL;DR: The benefits and pitfalls of some of the most commonly used protocols, focusing on stable isotope labeling with amino acids in cell culture (SILAC), are reviewed and it is speculated that SILAC will soon be a key tool of quantitative proteomics.

TL;DR: Current used protocols that have allowed us to apply VIGS to the identification of genes required for disease resistance in plants are described and the underlying general principles also apply when Vigs is used in the analysis of other aspects of plant biology.

TL;DR: A transgene containing inverted repeats separated by a functional intron such that mRNA produced by the transgenes is predicted to form loopless hairpin RNA following splicing, and the presence of the intron spacer greatly enhances the stability of inverted-repeat sequences in bacteria, facilitating the cloning procedure.

TL;DR: The advantages of generic gene-silencing vectors such as the pHANNIBAL and the pHELLSGATE series are described and evaluated and the protocols for their efficient use are provided.

TL;DR: An overview over recent intracellular FCS applications and a practical guide for investigators, who are seeking to integrate FCS into live cell imaging to obtain information on particle mobility, local concentrations, and molecular interactions.

TL;DR: Fluorescence correlation spectroscopy extracts information about molecular dynamics from the tiny fluctuations that can be observed in the emission of small ensembles of fluorescent molecules in thermodynamic equilibrium and provides access to environmental parameters such as local oxygen concentrations, pH, or viscosity.

TL;DR: It is demonstrated that thick tissues including muscle and bone can be imaged and sectioned through several hundred micrometers of depth by laser-scanning second harmonic generation (SHG) microscopy and found that SHG provides molecular level data on radial and lateral symmetries that GFP constructs cannot.

TL;DR: The system development for the image acquisition for one- and two-photon excitation FRET microscopy and the precision FRET (PFRET) data analysis algorithm that is developed to remove spectral bleed-through and variation in the fluorophore expression level (or concentration) for the donor and acceptor molecules are described.

TL;DR: A set of assays are described that allow the analysis of tight junctions to determine their integrity and functional state and contribute to the maintenance of cell surface polarity by forming a fence that prevents apical/basolateral diffusion of lipids in the outer leaflet of the plasma membrane.

TL;DR: The methods currently in use in the laboratory to isolate and immunoprecipitate native chromatin from cultured cells, and to isolation and analyze immunop Recipitated protein and DNA are outlined.

TL;DR: The experimental design and analysis of FRAP experiments are examined to allow a quantitative assessment of the relative influences of different molecular interactions on the steady-state distribution and protein function in vivo.

TL;DR: The ChIP assay currently used in the lab is presented and the various ways to optimize this assay for one's own use are discussed.

TL;DR: Robotic detection and tracking methods were developed to recognize and characterize the behavior of single molecules in recorded video sequences that enabled us to measure the kinetics of photobleaching and lateral diffusion of membrane-bound molecules.

TL;DR: Here it is explained how to prepare and use Drosophila embryo lysates to dissect the mechanism of RNAi, which can be recapitulated in vitro in lysate of syncytial blastoderm Dosophila embryos.

TL;DR: Methods are presented for detecting differential expression using statistical hypothesis testing methods including analysis of variance (ANOVA), and methods for multiple testing correction and their application are described.

TL;DR: A modified crossl linking procedure that includes a protein-protein crosslinking agent in addition to formaldehyde is reported that has successfully mapped Rpd3 binding sites in vivo.

TL;DR: The mechanisms that mediate leukocyte transmigration and BBB disruption, as well as tissue culture models that are used to study leukocytes trafficking, are the focus of this review.

TL;DR: This article discusses plasmid design strategies and vaccine regimens, which can elicit immune responses that differ in magnitude, quality, and balance of cellular and humoral responses from those elicited by single components and thus provide further enhancement for DNA immunizations.

TL;DR: Dermal immunization methods described in this article target the epidermis, the dermis, or both and include the removal of hair, the circumvention or modification of the stratum corneum layer of the epidersmis, and the addition of DNA or amplification of DNA signal.

TL;DR: This system provides a much needed tool for automated digitization and reconstruction of 3D neuronal morphology that reliably captures detail on spatial scales spanning several orders of magnitude, that avoids the subjective errors that arise during manual tracing with existing digitization systems, and that runs on a standard desktop workstation.

TL;DR: This review focuses on the application of AFM for quantitatively characterizing the structural and thermodynamic properties of protein-protein and protein-nucleic acid complexes.

TL;DR: In this article, the authors show that there is a relationship between the detection of vaccine-induced T-cells by cytokine-based assays and clinical responses, and that these assays become increasingly relevant in clinical practice to suggest issues of assay validation and quality control.

TL;DR: A brief account of the theory and measurement of ROA is given and the ROA spectra of a selection of proteins, nucleic acids, and viruses are presented which illustrate the applications ofROA spectroscopy in biomolecular research.

TL;DR: The rationale, methodology, and useful hints for performing T-cell epitope mapping using ELISPOT analysis are described, and typical results of such an analysis are reviewed.

TL;DR: This article focuses on providing the detailed procedure to construct genetic adjuvant plasmids and prepare DNA vaccines formulated with conventional adjuvants, and offers a practical guide for the procedure of intramuscular DNA injection.

TL;DR: The dynamic organization of eukaryotic genomes in cell nuclei recently came into the focus of research interest and appropriate labeling procedures as well as cell chamber systems and important controls for live cell microscopy are described.

TL;DR: The relevance of the assay-based identification of regulated genes was confirmed in a series of breast carcinomas in which EPSTI1 was highly upregulated compared with normal breast, taking advantage of a three-dimensional hydrated collagen gel approach.