- Detailed recognition of small complexes and organelles (such as mRNA complex, Mitochondria, Peroxizome etc.) to measure colocalization of these objects in yeast models
- Yeast cells segmentation is performed for both label-free or fluorescently tagged cells
- Colocalization measurement may be conducted under both live cell conditions or in fixed samples of yeast and other models
- Allows quantification and measurement on a cell-by-cell basis
This application is aimed for detection & quantification of colocalization and proximity events in yeast. The distance between two labeled organelles within a single yeast cell is measured. Automatic single cell recognition of yeast cells is implemented in this application to allow for accurate studies of per-cell colocalization events.
high quality and high-resolution images are requisite for colocalization assays, since they are based on the nanometric distance between two fluorescent foci within a single yeast cell. Furthermore, because yeast cells are roughly spherical, and most organelles in yeast move in three dimensions, image acquisition requires Z-stacks to gather 3D data in brightfield and multiple colors. The Hermes microscope’s capability to image rapidly ensures that all images are in sharp focus and not blurred due to sample movement during acquisition.
To meet the analysis requirement that yeast cells be identified and segmented using bright field illumination, we at IDEA added new image analysis algorithms specially designed to detect the yeast cell’s dark ring-structure.
Yeast serve as the primary model organism for molecular and systems biology, being a simple eukaryotic organism, easy to manipulate, able to cope with a wide range of environmental conditions and controls cell division similarly to human cells. Baker’s yeast, or Saccharomyces cerevisiae as it is also known, is among the best-studied experimental organisms. The maturity of yeast’s genetic and molecular toolbox has positioned it as the primary platform for development of many high-throughput technologies, including transcriptome, proteome, and metabolome screens. Yeasts play a significant role as a model organism for human disorders due to their conserved biochemical pathways that control key aspects of eukaryotic cell biology. These functional pathways drive cellular growth, cell division, cell trafficking, stress-response, and secretion, among others, all of which are known to be associated with various human pathologies. Yeast has also contributed to our understanding of cancers and neurodegenerative disorders and are also used extensively as a model organism in aging research.
Quantifying the localization of proteins, RNAs and complexes within cells, as applied with WiSoft® Athena’s yeast colocalization analysis application, can help determine their regulation and sites of action. mRNA and protein transport and localization have a key determinant of whether cells establish sub-cellular domains necessary for different activities, confer the molecular interactions required for protein function, and create functional organelles. Moreover, events that lead to the mislocalization of proteins can have dramatic consequences upon cell viability. Quantifying the distance between the trafficking object and the nearest intracellular organelles is therefore an important tool in exploring and understanding these important cellular mechanisms.
Yeast cells labeled for Mitochondria (red) and mRNA (green) imaged with Hermes imaging system using 40x/0.9NA objective
Analysis of yeast cells (blue outline), Mitochondria (red outline), mRNA complex (green outline) and colocalized events (cell marked using orange outline) with WiSoft® Athena dedicated analysis app are shown.