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Traptavidin, a streptavidin mutant with about tenfold lower 'off' rate for biotin than streptavidin itself, has increased mechanical strength and thermostability. It should find use in a diversity of applications in which the dissociation of streptavidin can be a limitation.
A microfluidic device containing a suspended microchannel resonator capable of measuring the mass of microscopic objects with femtogram resolution allows determination of bacteria, yeast and mammalian cell growth rates in less than one cell cycle by repeated measurement of the buoyant mass of single growing cells.
Robust and accurate quantification of human tumor tissue proteomes is made possible by combining the tissue sample with an 'internal standard' mixture of five relevant, stable isotope–labeled cell lines, followed by mass spectrometry analysis.
A theoretical and experimental treatment of fitting methods for localizing the centers of diffraction-limited spots is presented. Use of an analytical point spread function shows that maximum likelihood fitting is superior to both unweighted and weighted least squares Gaussian fitting.
An iterative algorithm implemented on a graphics processing unit determines maximum likelihood estimates of the positions of isolated fluorophores at a rate of 105 localizations per second and allows real-time generation of super-resolution images with high precision.
A conditional gene expression system in Caenorhabditis elegans is reported. It should permit the generation of temperature-sensitive alleles for most genes.
Comparing the haplotypes of a few randomly microdissected chromosomes to a full genome-wide haplotype allows one to determine the long-range haplotype of chromosomes for which only one copy was captured.
A side-view endoscope permits the imaging of large fields of gastrointestinal and respiratory mucosa at high resolution in the mouse. The approach is applied to imaging changes during inflammation and tumor progression in the living mouse.
Fast protein interaction dynamics can be observed in cells via a fluorescence correlation spectroscopy–based method. Endogenous bait proteins are captured via quantum dots, and their interaction with fluorescently tagged prey proteins is monitored with high temporal resolution.
By stochastically sampling cells in groups of ten, the authors identify transcriptional programs with strong cell-to-cell expression differences thus allowing them to study endogenous heterogeneities in single cells.
Using a GFP complementation assay to tag effectors of the type-III secretion system in gram-negative bacteria allows localization of the effectors in the host cell in the course of bacterial infection.
A Toxoplasma gondii strain that efficiently secretes Cre recombinase into infected host cells permits the deletion of genes specifically in the infected cells. It should facilitate the study of host-pathogen interactions in vitro and in vivo.
Using DNA microarray–derived gene expression data from complex tissues and the relative frequencies of cell types in the tissue as input the algorithm csSAM finds cell type–specific differentially expressed genes.
Protein dynamics can be studied in single living cells by time-resolved fluorescent imaging of the unfolding of a fluorescence resonance energy transfer (FRET) probe—labeled protein as fast temperature jumps are applied.
A Rosetta full-atom framework, called fragment assembly of RNA with full-atom refinement (FARFAR), allows the de novo structure prediction and design of noncanonical RNA motifs with near-atomic resolution.
Chemically inducible dimerization probes selectively target proteins to the surface of specific organelles or tether organelles to each other, thus allowing precise spatiotemporal analysis of signaling events.
By subdividing a charge-coupled device (CCD) array into subgroups using a digital micromirror device and offsetting exposure times, temporal pixel multiplexing allows simultaneous high-speed and high-resolution imaging using a single CCD. This imaging modality allows 250 Hz microscopic imaging of fast cellular responses with a 10-Hz 1.3 megapixel camera