Nonetheless, high-throughput systems are in urgent demand to support studies of neural morphologies at larger scale and more detailed level, as well as to enable research on non-human primates (NHP) and human brains. Brain-wide imaging with single-cell resolution provides unique advantages to access morphological features of a neuron and to investigate the connectivity of neuron networks, which has led to exciting discoveries over the past years based on animal models, such as rodents. The SEM is too small to show error bars.Ī deep understanding of the neuronal connectivity and networks with detailed cell typing across brain regions is necessary to unravel the mechanisms behind the emotional and memorial functions as well as to find the treatment of brain impairment. (g) Speckles: fluorescent foci of phospho-Histone2AX induced by 2 Gy of irradiation in human U2OS cells disappear at timepoints as the cells recover. Totals do not equal 100% due to slight overlap between compartments. (f) Protein localization: the mean intensity of NFκB staining in the cytoplasm and the nucleus is shown in response to TNFα in human MCF7 cells (top). Wild-type human HT29 nuclei that stain positively for phospho-histone H3 tend to be smaller than phospho-H3-negative cells (right). (e) Phospho-protein content: human HT29 cells treated with RNAi reagent against Polo kinase have an increased percentage of nuclei with high phospho-H3 staining compared to wild-type cells, consistent with a mitosis-stalled phenotype (left). elegans, only the boxed region of interest was analyzed. The histone content is decreased by roughly half in each daughter nucleus after division. elegans) content is shown near each nucleus in arbitrary intensity units. cerevisiae) or GFP-histone H2B (HeLa and C. (d) Chromatin content in time lapse movies: GFP-histone H4 (S. RNAi-targeted samples were also as expected for Aurora kinase B (polyploid), Mad2 (fairly normal cell cycle distribution), String (4N-enriched), and Anillin and Cyclin A (both polyploid). The cell cycle distributions are as expected, with the 2N peak being predominant in the wild-type human sample, whereas most wild-type Drosophila nuclei are known to have 4N DNA content. ![]() (c) DNA content in cell populations: measurements are shown for human HT29 cell populations (1 image for each RNAi condition, left) and for Drosophila Kc167 cell populations (1,750 images for each RNAi condition were combined, right). (b) Cell size: CellProfiler's cell area measurements are comparable to those of a Coulter particle counter for Drosophila Kc167 cells, for wild-type (no dsRNA) and RNA-interference induced samples. Example images and CellProfiler outlines for these cell types are shown in e. For images of Drosophila Kc167 cells with various genes knocked down by RNAi (right), the two researchers' counts varied by 16% and CellProfiler's counts were within 17% of their average. (a) Cell count: for a set of 6 images of wild-type human HT29 cells (left), two researchers' counts varied by 11%, and CellProfiler's counts were within 6% of their average. Validation of CellProfiler for many cellular phenotypes.
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