Below you see the pictures, longer structures in B.POINT: shows the importance of a kinetic readout, how it completes the story of cytochalasin D’s effectsThese results were replicated 2 more times at Essen and agree with reports from the literature.(for an assay of 80% power to show a 10% change in neurite length) The intra-assay variability was quantified using “non….on polydlysine” plated at 8k cells/wellError bars represent standard deviationCoefficient of variation remains well under 10%.96 well plate view demonstrates low variability.A corresponding power analysis demonstrated that N=6 with 9 images per well is suggested.In these graphs of three different timepoints you see that “NeuroTrack quantitation….in a high content imager.”
#Neurotracker neurite quantification software#
They utilized their corresponding software to measure neurite length and compared the results with the ZOOM analysis. These plates they took to the University of Michigan to image in their Image Xpress Micro. Scientists at Essen performed assay validation in an experiment where they took plates of cells at three different timepoints, imaged in ZOOM, and immediately fixed, immunostained with Beta-tubulin, the standard for labeling neurites in fluorescence.This slide summarizes the flow of information in a NeuroTrack assay-the phase images of the cells can be made into time lapse movies, and the masks automatically produce data of the metrics just described.This is a slide showing the NeuroTrack process– From the phase images, it generates the user-customized mask, and from this mask quantitative data is produced in real-time.NT masks these very fine primary neurites with the same precision as large, thick neurites.A good processing definition on NeuroTrack can be made in under 10 minutes. HCI systems in comparison have much more complex, time consuming processes. There are just enough parameters to customize to the user’s cell type, but it’s not an overwhelming process. Really there is a minimal amount of user involvement needed to create a very good mask like the one you see here. These are non labeled E18 rat cortical neurons plated on poly-D-lysine at 3 different timepoints post-plating1.Very important fact that NT is compatible with such a wide range of models, as they have very different morphology. This is the alternative protocol NeuroTrack (NT) provides scientists.Step 1: Select cells.POINT: Labor intensive process, complex, plenty of troubleshooting involved, and all for data that comes from only a single timepoint!Essen combined the capabilities of ZOOM and the talent of the programming engineers to improve the system for measuring neurite dynamics.Generally paraformaldehyde used-not a user friendly chemical.Long time required for incubation with antibodies.
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Neurite dynamics have an important role in….What are neurite dynamics?A term that includes neurite outgrowth, in which a neuron extends processes to create neural networksHere you see the stages of outgrowth, in which axons and dendrites formNeurite is a non-specific term including both axons and dendritesIn addition to outgrowth, loss of neurite length is included in neurite dynamics, and is also important to studies of development and degenerative diseasesNote: NeuroTrack does not distinguish between axons/dendrites.
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