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Lists commands for I/O operations. Most are self-explanatory. Noteworthy:
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Contains commands for organizing tracing views on screen, plus commands for displaying cached data used for auto-tracing.
This home tab aggregated widgets for tracing and frequent operations.
Enable Snapping checkbox If active (the default) the cursor snaps to the brightest voxel in its vicinity (Toggling shortcut: S). To accomplish this, SNT takes the cuboid of the specified dimensions (in pixels) centered on the current cursor position and searches quickly for local maxima in that neighborhood, moving the cursor to that position. The Z-plane in which the maximum was found is automatically selected if the "Z" parameter is greater than 0. Noteworthy:
- This feature assumes the signal is brighter than the background as typically found in fluorescent images.
- If multiple maxima exist (e.g., when the signal is saturated), it snaps to their centroid.
- To streamline the computation: XYZ dimensions are constrained to even numbers and limited range.
- Snapping occurs in 2D (i.e., in the active plane) if Z=0.
- The XZ, ZY views are synchronized when 3D snapping is active (i.e., Z>0).
Enable A* search algorithm By default, SNT uses the A* search to automatically trace paths between two manually selected points. To manually place nodes in a path, toggle this feature off. Note that it is also possible to enable other algorithms through the installation of SNT add-ons. See Tubular Geodesics for details.
Hessian-based analysis (Toggling shortcut: H) A quick way to improve the quality and efficiency of the pathfinding is to enable this feature, in which paths are computed after filtering the image for tube-like structures. Upon such filtering, SNT will use a measure of Tubeness at each point of the image to define the best path through it, based on eigenvalues and eigenvectors of the Hessian matrix. The later can be used to infer the likelihood that a point in the image belongs to a tube-like structure. This concept is also known as vesselness or neuriteness.
Hessian options (Specified in the gear drop-down menu) may be chosen Manually (if you already have a quantitative understanding of the image) or Visually (generally more intuitive). Hessian analysis requires two parameters to be specified:
- Sigma Controls the spatial scale of the filter. A lower sigma (σ) will capture small scale structure but is more sensitive to noise. A larger σ will consider larger scale structures but is less sensitive to local shape characteristics. It is generally advised to pick a value which reflects the average radius of the structures to be traced. It corresponds to the standard deviation of the 3D Gaussian kernel used to smooth the image prior to Tubeness filtering
- Maximum The maximum pixel intensity on the Tubeness image beyond which the cost function for A* search is minimized.
If you choose Visually, the dialog will prompt you to click on a representative region of the image which has meaningful structure. Once you click there, a 9-square palette is generated showing increasing values of sigma (from top-left to bottom-right) applied to that region of the image. The palette is an image stack which can be scrolled through with the mouse wheel.
|Choosing sigma and max. parameters visually for hessian analysis from a representative region in the image (orange box). With saturated max, the cost function for A* search is equivalent across the signal. Also, note how lower sigma values cause the axon to be dimmer on the center and brighter on the edges, while higher sigma values eliminate detail from the structure.|
Experiment with different Adjusted max. values to find a setting which captures the structures you are interested in while taking care to avoid over or under-saturation. In the former case, the search might consider pixels outside of the tubular structures, giving a less accurate result. In the latter case, the search might take significantly longer, since each pixel will carry greater cost than is reasonable. It is usually safe to choose a value somewhere near the default. Once you have chosen this value, click on the square in the palette with the sigma that best fits your application (its border will turn green) then press "Apply". The parameters will be transferred to the interface and a Gaussian convolution will be run over the image, which may take some time depending on its size. Once it's done, pathfinding will occur on the Hessian (a.k.a. Tubeness) image.
Other options are also available in the gear drop-down menu:
- Cache All Hessian Computations By default SNT computes the Hessian matrix on a "compute-as-you-go" basis, which saves memory but increases the pathfinding time. If you have enough RAM, you can use this option to compute once the matrix and for the full image and cache the result, which in certain cases, can lead to almost-instantaneous searches. Note that this computation adopts the current values of sigma and max. reported in the widget, so you should adjust those before running this command.
- Flush Cached Data Removes cached data from memory
- Load Precomputed "Tubeness Image" This (advanced) option prompts you to load a file of a Tubeness image generated elsewhere. This is useful if e.g., you want o experiment algorithms from other software packages, or your own. Note that loaded image is assumed to have compatible dimensions with the image currently being traced.
- Show Cached "Tubeness Image" Displays the Tubeness data currently cached. Useful for debugging purposes. Also mirrored in . ›
Tracing on Filtered Image
This is one of SNT's most advanced features. Hessian-based analysis provides a convenient and easy way to perform auto-tracing on pre-process data in which voxels defining the traced structure are scored by their likelihood of belonging to a tube-like structure. However, it is just one approach for "tubeness" classification. What if your data requires different filtering?, or you want to experiment with other approaches?, or the perfect pre-processing algorithm for your images is not yet available in ImageJ? Tracing on Filtered Image is the answer to these questions: It allows you to feed SNT with pre-processed data on which the A*star search will operate. Here are two specific usages for this option:
- Frangi Vesselness filtering For certain datasets Frangi filtering (Frangi et al. 1998) is more effective than Tubeness at enhancing tube-like structures in the image. However, it is more computation intensive, and thus, less suitable to be adopted by the "compute-as-needed" approach used in Hessian-based analysis. Thus, one can import a pre-computed Frangi-filtered image using this option.
- Hessian-analysis at multiple scales Let's consider a structure formed simultaneously by very thick and very thin processes. To trace it one would need to adjust continuously the sigma parameter while tracing. Instead, to trace structures of variable diameters more effectively, the Hessian image associated with the second sigma could be loaded as Filtered Image.
- Adopting probability maps Pre-classified images (using e.g., machine learning) could also be loaded here.
Importantly, this option can be toggled at will, during tracing. Indeed, one can ping-pong between filtered image and tracing image simply, by pressing I, the shortcut for the Trace on filtered Image checkbox.
Filtered Image options can be specified in the gear drop-down menu, including commands for Loading/displaying the image file and the powerful Generate Filtered Image command:
See the Generating Filtered Images walk-through for more details.
Filters for Visibility of Paths
By default, the entire path is projected onto the current Z-slice. This is useful to see how much has been completed and gives a sense of the overall structure of the reconstruction. However, SNT provides 3 additional visibility options for paths:
- Only selected paths (hide deselected) Only show paths that have been manually selected in the Path Manager or with the G key (⇧ Shift+G to select multiple paths).
- Only nodes within X nearby Z-slices Only highlight nodes within X number of Z-slices on either side of the current slice. The projected skeletons of all paths remain visible.
- Only paths from active channel/frame If tracing on a multichannel image or an image with a time axis, only show paths from the active channel or frame.
Any combination of these options may be toggled simultaneously.
Default Path Colors
By default, finished paths are colored by their selection status (only selected paths can be edited, or extended). The default colors are Green (selected paths) and Magenta (deselected). Default colors can be customized by pressing the respective button in the widget and using the CMYK color chooser. For customizing unconfirmed and temporary paths, see the Colors option in the UI Interaction widget.
Enforce default colors (ignore color tags) If active, SNT will force all paths to conform to the default "Selected" and "Deselected" color buttons. Any custom color tags will be ignored until the option is toggled off.
This tab aggregated widgets for advanced settings.
If tracing on a multi-dimensional image (i.e., one with multiple data channels and/or a time axis), a particular channel/frame can be loaded into the views by selecting each and pressing the "Reload" button.
- Overlay MIP(s) at X% opacity Overlays the Maximum Intensity Projection of the image "over" the image canvas at the specified opacity. Overlaid projection is only used as a visualization aid and is ignored by the auto-tracing algorithms. It is rendered using the LUT of the channel currently being traced. To reload the overlay (e.g., in case the image being traced changes during a tracing session) toggle the checkbox twice.
- Draw diameters in XY view Displays the stored diameter (if any) in the XY view for all existing nodes. Each diameter is drawn as a line segment with length = diameter, which is bisected by the orthogonal tangent vector to the path at that node.
- Apply zoom changes to all views If a zoom change is applied to any one of the XY, ZY or XZ views, apply the same change to the two other views if they are open. Since in ImageJ zooming may resize the image window, you can use to reset their positions ›
- Resize Canvas If using a display canvas to view reconstructions, reset its dimensions to the default. (Currently, this command is only available for display canvases, to resize an image go to IJ's command › ›
- Display ZY/XZ views If currently using the XY only view, display the ZY and XZ views as well.
- Confirm temporary segments If active, prompts for either confirmation or denial of whether or not to keep an unconfirmed path segment. If inactive, automatically confirms the path segment created on each left-click after starting a path (first click). Applies to both auto-traced and manually traced path segments. The following two settings are only toggle-able when this setting is active.
- Pressing 'Y' twice finishes path Finish a temporary path on two successive Y key presses.
- Pressing 'N' twice cancels path Discard a temporary or unconfirmed path, including the start node, on two successive N key presses.
- Colors Specifies how components should be rendered, including:
- Canvas annotations The label shown on the top-left corner of the views indicating the state of the UI ("Tracing Paused", "Choosing Sigma", etc.)
- Fills The pixels that have been reached by the Fill search
- Unconfirmed and Temporary paths.
- Path nodes rendering scale (default scale is inferred from current zoom level) Adjusts the radius of the circles representing path nodes. A path node is rendered as a circle centered at the XYZ coordinate of the point annotation.
- Activate canvas on mouse hovering If active, moving the mouse cursor over the any of the views automatically brings the view window into focus, allowing it to receive input.
- Skip confirmation dialogs If active, forgoes the Are you sure? prompt preceding major actions. Note that this option does not apply to irreversible actions such as deleting paths.
- Debug mode If active, logs detailed information about actions in the console.
- Preferences... Allows setting other options, namely:
- Whether the position of dialogs should be remembered across restarts
- Whether Gzip compression (lossless) should be used to reduce the storage footprint of ".traces" files.
- "Reset Preferences..." Allows the reset of all options to their defaults. A restart of SNT may be required for changes to take effect.
This tab aggregates widgets related to 3D interaction.
The Reconstruction Viewer is an advanced OpenGL visualization tool. For performance reasons, some Path Manager changes may need to be synchronized manually from RV controls. To open Reconstruction Viewer with the currently loaded tracings, press Open Reconstruction Viewer. For more information, see the Reconstruction Viewer wiki page.
Legacy 3D Viewer
The Legacy 3D Viewer is a functional tracing canvas but it depends on outdated services that are now deprecated. It may not function reliably on recent operating systems. For usage instructions, see Legacy 3D Viewer
SciView is IJ2's modern replacement for the Legacy 3D Viewer providing 3D visualization and virtual reality capabilities for both images and meshes. It is not yet available in SNT.
Right-clicking on any of the image views will bring up a menu with various editing tools. The corresponding keyboard shortcuts are shown to the right of each option.
- Select Nearest Path G or ⇧ Shift+G Will select the path closest to the mouse cursor.
- Fork at Nearest Node ⇧ Shift+⎇ Alt+Left Click Creates a fork point at the node closest to the mouse cursor. Once a fork point is made, the branch may be extended as described in Step-By-Step Instructions.
- Continue Extending Path Allows continued tracing of previously finished paths. Note only one path may be extended at a time. To extend a path: first select it, choose this option, then place additional nodes as shown in Step-By-Step Instructions.
- Pause SNT Waives all keyboard and mouse inputs to ImageJ, allowing you to interleave image processing routines with tracing operations. Note that if the image contents change while SNT is paused, the image should be reloaded so that SNT is aware of the changes. Tracing views are annotated with the SNT Paused label to indicate this state.
- Pause Tracing Disables tracing functions until this option is deselected. Tracing views are annotated with the Tracing Paused label to indicate this state.
- Sholl Analysis at Nearest Node ⇧ Shift+⎇ Alt+A Runs the Sholl Analysis plugin found in . Note the Center parameter, which sets the center point of the analysis, is left out as this value is given by the selected node. › ›
Pressing Edit Path with a single path selected will activate Edit Mode, allowing use of the menu options under the Edit Path option. When Edit Mode is active, moving the mouse cursor along the path will activate the nearest node and synchronize the current Z-slice to the location of that node. Note that the ability to create new paths is temporarily disabled when in Edit Mode.
- Reset Active Node Clears the active node from the cursor.
- Delete Active Node D or ⌫ Backspace Permanently removes the active node from the path.
- Insert New Node At Cursor Position I Inserts a new node at the cursor position. The inserted node is placed between the active node and its parent.
- Move Active Node to Cursor Position M Moves the active node to the cursor position.
- Bring Active Node to Current Z-plane B Moves the active node to the active Z-plane. Note that the translation is only done in Z. XY positions are unchanged.
- Connect To (Start Join) Allows two existing paths to be merged or joined.
The Path Manager dialog displays all existing paths in a hierarchical structure (tree), where one path is "primary" (path 0) and all other paths (paths 1...N) are children of the primary path. The dialog also contains several menus with various editing, tagging, refinement/fitting, filling and analysis options. Paths can be searched by name and/or tags in the text filter, with more sophisticated search capabilities in the Advanced Filtering Menu.
- Delete Removes selected path(s) from the Path Manager.
- Rename Renames the selected path.
- Make Primary Makes the selected path the primary path, moving it to the top of the tree. Note that this will alter the reconstruction topology.
- Disconnect Disconnects the selected path from all of its connections (Undoable operation)
- Merge Merges the selected paths (at least two) into one. Note the starting node of path i is merged to the endpoint of path i+1
- Specify Radius Assigns a constant radius to all the nodes of selected path(s). This setting only applies to unfitted paths and overrides any existing values.
- Ramer-Douglas-Peuker Downsampling Given an inputted maximum permitted distance between adjacent nodes, performs Ramer-Douglas-Peucker downsampling on the selected path(s).
Assigns tags to Paths. Tags are organized in the following categories:
- Type Type of neurite compartment (Axon, (Basal) Dendrite, etc.), as used by the SWC format. It is also possible to pair each type with a color tag by enabling the respective option in ›
These tags are considered to be essential annotations and all paths are assigned the Undefined-type tag when created. For this reason they are not eliminated by the Remove All Tags command.
- Color A preset swatch color, a custom one chosen from the color chooser. Note that is also possible to assign mapping LUTs using the command. ›
- Image Metadata Information on hyperstack position details (e.g., channel or frame on which a path was traced).
- Morphology Morphometric properties, such as Path length, Path mean radius or branch order.
- Custom Ad-hoc comments.
Note that only SWC-type tags are preserved across restarts when saving traces in the SWC format. All others require data to be saved in SNT's own .Traces format.
See Fitting in the Step-By-Step Instructions wiki page.
- Fill Out Begins the filling process for selected paths. For detailed instructions see Filling in the Step-By-Step Instructions page.
This menu contains several options which provide quick ways to analyze and visualize numerical properties of paths.
- Color Coding Assigns color codes to paths based on the chosen metric.
- Color by Drop-down menu containing the metrics which inform the color mapping.
- LUT Drop-down menu containing the LUTs (Look Up Tables) that define the color palettes. The LUTs are those that come packaged with ImageJ. The selected LUT is displayed in the color bar directly underneath.
- Rec. Viewer Color Map If active, opens an instance of the Reconstruction Viewer with the selected paths color coded with the selected LUT.
- Rec. Plotter Color Map If active, open an instance of the Reconstruction Plotter with the selected paths color coded with the selected LUT.
- Remove Existing Color Coding Removes existing color coding from the selected paths.
|After choosing metric: Branch Order, LUT: Ice, showing the Reconstruction Viewer with the color-mapped OP_1 tracing.|
- Distribution Analysis Plots a histogram of the selected metric.
- Measurement - Drop-down menu with the available metrics.
|Choosing Branch Order for the measurement.|
- Measure Shows a table of summary statistics for selected paths. If measuring multiple tracings, the table can be sorted by column.
- Convert to ROIs Allows conversion of paths to ROIs (Regions of Interest). Creates an instance of the ROI Manager
- Convert Drop-down menu specifying the compartment to convert.
- View Drop-down menu specifying which view to overlay the ROIs.
- Impose SWC colors
- Adopt path diameter as line thickness
- Discard existing ROIs in ROI Manager
|Converting paths to ROIs with the default parameters.|
- Plot Profile Uses the pixel intensity values under point annotations to generate a pixel intensity vs. distance plot for selected paths.
|Intensity profile for Path 0 of OP_1.|
- Skeletonize Outputs a binary image that is a topographic skeleton, ie, it generates an empty (zero-filled) image of the same dimensions of the one being traced, then paints a pixel at each node coordinates following the topographic rules of bitmap skeletons in which fork points, tips and slab voxels are determined by voxel connectivity.
- Roi filtering
- Convert only segments contained by ROI
- Run "Analyze Skeleton" after conversion Runs the AnalyzeSkeleton plugin on the skeletonized output image.
- Roi filtering
|Converting paths to a topographic skeleton with default parameters.|
- Save as SWC Exports selected paths as an SWC file. Note the paths to be exported must include a primary path (i.e., one a the top level in the Path Manager tree).
The text filter allows paths to be searched by any characters in the path name string and by color. The name string includes all tags.
- Up Arrow Button Finds next occurrence of the entered phrase.
- Down Arrow Button Finds previous occurrence of the entered phrase.
- Balloon Button Highlights all occurrences of the entered phrase.
- Text Filtering Contains options for advanced text filtering
- Case Sensitive Matching treats uppercase and lowercase characters as distinct.
- Enable Wildcards Allows the use of the question mark (?) and asterisk (*) wildcards.
- "?" Matches exactly one character. For example, the pattern 123? will match 123 and 1234, but not 12345.
- "*" Matches zero or more characters. For example, doc* matches doc and document but not dodo.
- Loop After First/Last Hit Cycles back to the first/last matched phrase upon pressing the Up/Down Arrow Button at the end of the list of matches.
- Display No. of Matches Displays the number of phrase matches in the bottom left corner of the Path Manager dialog.
- Replace Replaces all occurrences of the matched phrase in the path name string for currently selected paths.
- Clear History Erases the previously searched phrases in the text filter drop-down menu.
- Tips & Shortcuts
- Case-insensitive by default. Wildcards ? (any character), and * (any string) can also be used.
- Press the Highlight All button or Ctrl+A to select all the paths filtered by the search string.
- Press and hold Ctrl while pressing the up/down keys to select multiple filtered paths.
- Press the up/down keys to find the next/previous occurrence of the filtering string.
- Uncheck Display No. of Matches to improve search performance.
- Color Filters Allows filtering of paths by color code. Custom colors may be selected by right-clicking an empty swatch, which will bring up the CMYK palette. The chosen color is temporarily saved in that swatch.
- Morphology Filters Allows filtering of paths by select morphological properties.
- Branch Order Filters for paths of branch order in the inputted range. Note that the paths of interest must have Branch Order tags. To tag paths with their centrifugal branch orders, use . See › › Tag for more information.
- Length Filters for paths of length within the inputted range. Note that the paths of interest must have Length tags. To tag paths with their length, use . See › › Tag for more information.
- Mean Radius Filters for paths of mean radius within the inputted range. Note that the paths of interest must have Mean Radius tags. To tag paths with their mean radius, use . See › › Tag for more information.
- No. of Nodes Filters for paths with node count within the inputted range.
- SWC Type Filters for paths with the selected SWC type tags. Note that the paths of interest must have Type tags. To tag paths with SWC types, use . See › Tag for more information.