Current Version is 6.4.5, posted April 4th 2006

Index:

Version 6.4.5 (April 4th 2006)
Version 6.3.4 (September 30th 2005)
Version 6.2.1 (March 4th 2004)
Version 6.1.1 (October 11th, 2004)
Version 6.0 (July 5th, 2004)
Version 4 History
Version 3 History
Version 2 History

April 4th 2006 , Version 6.4.5 released
SUMMAY OF CHANGES SINCE 6.3.4

Bug fixes:

• Allowed FJ to run with a v8, v6, or v4 Site (FLS) License serial number.
• Added a unique error message in case the program times out while waiting for the
  Site License file to be downloaded from the Site License server.
• Fixed intermittent problem in Site License (FLS) validation, where a machine might
  sometimes run properly, and at other times complain that the machine
  could not be found in the given Site License.
• Fixed FLS authentiction lookup
• Enhanced "Notify" messages in cases where a valid Site License could not be found.
  
• ADDED SUPPORT FOR V8 Serial numbers and USB ThumbDrive dongles
• Fixed the ability to enter line breaks into Layout Editor annotations
• Fixed the bivariate display scaling of the event number parameter
• Find missing FCS files - now fixed for SMB networks. If the workspace is referring to missing files, there will only be one dialog per folder of FCS files - rather than every single missing file.
• No more hanging when the Tree Star web site is impossible to reach
• Can create derived parameters on Cyan-acquired data
• Workspace statistics don't disappear when a recalculate is performed
• Read all Coulter data files
• Cell cycle: constraining peaks to gate positions is now correctly saved
• Cell cycle: option to use pattern fill for graphs is correctly saved
• Annotation items in layouts are properly filled for batch outputs
• "FS LIN" and "SS LIN" parameters are now recognized as scatter parameters
• Improved performance and stability
  

September 30th 2005, Version 6.3.4 released
SUMMARY OF CHANGES SINCE 6.2.1

Important Notes:

Note: workspaces created with this version cannot be opened with older versions.

New Features.

New in 6.3.4 (cumulative changes since 6.2.1):

• Significant performance improvement noticeable when compiling large layouts or tables-particularly when hundreds or thousands of samples are involved!
• Fix text issues in dialogs on some computers running OS9.2.2.
• Summit 4.x files with incorrect data location specification keywords are now read in properly.
• The compensation definition window can now be resized to allow for long parameter names.
• Fixed: Memory was slowly being used up by doing large numbers of text boxes in the Layout Editor
• Fixed: Tables with negative numbers in the statistics will now show means & standard deviations.
• A number of optimizations and bug fixes were done to prevent crashes that occur during large computational demands; stability should be much better.
  
  
  General Operation:
• Site licensees are validated more efficiently
Preferences:
• There is a new preference to define the default sort order for samples in the workspace Workspaces:
• Workspaces are now read in and saved much more efficiently. Note: the first time you read in an older workspace, it may be much slower; as soon as you save in the new format, it will be considerably faster!
• Workspace sort criteria are now "persistent": if you add new files (or for templates), the files will be automatically sorted according to the last selected criteria. In addition, when you define the sort criteria, you can make them the default for all new workspaces.
• Workspace sort criteria can be individually set to be increasing or decreasing values.
• Unreferenced samples are now removed from workspaces (making templates much smaller).
• XML format has been modified (more similar to PC version). This format is still being updated and is not guaranteed to be consistent with future revisions of FlowJo.

Tables:

• Tables with keyword values that have <cr> or <tab> display properly
• You can copy a series of custom column heading names (from Excel or Word, separated by returns) into any column heading name – the entire series is copied to the subsequent table columns (hit return or tab after pasting). In this way, you can edit the column names in a different program and then copy them in all at once.
• The mean and std. dev. are filled in only for valid calculations (e.g., they are left blank for text columns)

Bug fixes:

• reading newest Coulter data files
• Cell cycle models that constrain to a region--constraint is saved after closing & reopening
• Cell cycle platform option to display pattern filled models is properly saved
• Annotation items for Layout Graphs are properly built in batch reports "FS LIN" and "SS LIN" are recognized as forward and side scatter parameters
• Fixed bug that could cause crash during gate computation while other tasks were happening-improved stabililty!
• When directing a batch output directly to the Printer, the page information is properly initialized without having to go through the "Custom" settings button.
• Fix: Preferences correctly reads layout color palette for overlay colors
• Fix: Bug that could cause rare occasional crashes when reading files
• Fix: When choosing to sort the workspace, the current sort criteria are displayed
• Templates that refer to compensated parameters now add all gates & statistics
• Renaming a gate will no longer cause a Layout (referring to that gate) to crash
• Opening graph windows no longer occasionally reset the Layout iteration value
• Cycling graphs (next/previous) no longer cause a rare crash that happens when gate frequencies were shown
• Modifying a series of gates that are all group-synchronized no longer causes a rare crash
• $SPILLOVER keyword values are properly supported
• Allow dragging gates from one workspace to another if the destination sample is selected
• Crash no longer occurs if changing a graph while an outlier plot is computed
• Selecting a graph from a sample not in the current group does not make that sample temporarily appear in the workspace window
• Double clicking multiple samples in the control definition list for an overlay graphic now sets all selected samples as controls
• Grid lines are displayed in histogram overlays
• Histogram overlays will not occasionally display parameter names for the Y axis label
• Boolean gate specifications are no longer case-sensitive
• The Layout batch Preview will not crash if the display is too large to show on-screen
• The line color for boxes drawn around text is correct in batched layouts
• Summit data files with badly-formatted $SPILL keyword (Summit version 4.1) are now read in (an error message is shown) but no compensation parameters are added. These files can be compensated the usual way in FlowJo, using appropriate compensation controls.
• FlowJo does not crash if you try to delete the only layer listed in the Legend Item for a Layout Graphic.
• Batch layout generation: if Iterate by Nth (n > 1), the resulting layout's discriminator is properly set.

Known issues:

• Files with long names (>31 characters) will not be automatically re-found when moved to a new location.

Important Notes:

The format of the workspace file has changed. Workspaces saved in 6.2b3 and beyond cannot be read by older versions of FlowJo.

New Features

Graphs and Gating:

• There is a new preference to control the default naming scheme for new Quadrant gates (you can choose to have only "Q#", where the number increments for each new quadrant set; you can choose to add the stain names; and, as before, you can choose whether or not to ask for a prefix to the gate name).
• The Backgating analysis now shows a legend of gates applied for each graphic, to better illustrate what is being shown. In addition, the last graphic in the window is no longer a backgating analysis (as there as no useful information for that level), but rather shows a simple overlay of the gated population.
• There is a new gating tool to create two gates that bisect a histogram; while tracking with the mouse with this tool active, the graph window will show live statistics (like the quadrant tool).
• A new menu item, "Show defining gate", allows you to quickly access the graph that defined any selected subset.
• Contour plots can no longer be drawn as unsmoothed.
• Log contour plots do not allow you to select "show outliers", which was meaningless for this display.
• Kinetics analyses now show the threshold value used for calculations in the kinetics window.
• Grid lines for linear parameters are now drawn to match major axis tick marks.

Data compensation and transformations:

• Corrects Summit Compensation matrices (for all versions of Summit).
• FlowJo now supports the FCS3.1 keyword "$SPILLOVER" as specified in the proposal.
• The compensation matrix name (if FlowJo was used to create the compensation) is now stored in a keyword ("FJ_CompMatrixName"),
• The compensation definition window has new buttons to calculate and close; to clear all positive & negative gates from the boxes; and has better selection of gates in the boxes.

General Operation:

• FlowJo keeps statistics of how many different kinds of operations you perform (number of files analyzed, gates created, events read): Choose "About FlowJo", and click on the disclosure triangle near the bottom of the window.
• All "Confirmation" dialog windows have button names that better explain what the two choices are (instead of just "Yes" or "No").

Layouts:

• When editing the attributes of platform graphics in the Layout Editor, you can now set additional options such as the colors and size of the graph.
• Scrolling large layouts is much quicker.
• You can now change fonts for individual layout items.
• For overlay legends, the cursor will change when targeting the color or line style controls to aid in identifying where to click.
• Generation of Batch Layouts was made more efficient (faster).

Tables:

• Tables can be saved as XML (for import into LIMS systems, for example). Choose "Save As XML" while viewing a completed table.

Preferences:

• Preferences reorganized (a 'Platforms' preferences panel added)
• The preference for copying the FCS Comment keyword into the workspace annotation was removed
• A new button clears all memorized auto-answers for confirmation dialog windows

New preferences to define:

• 1. Default statistics that are added to layout editor when Cell Cycle platform is dragged in (and the 'auto-create statistic table for platforms' preference is enabled).
• 2. Default statistics that are added to layout editor when Proliferation platform is dragged in (and the 'auto-create statistic table for platforms' preference is enabled).
• 3. Default columns to be created whenever a new overlay legend is created in the layout editor.
• 4. Default colors and graph types for Backgating analysis (these can be set either through the Preferences dialog, or by selecting the checkbox before closing a Backgating window).
• 5. The default scheme by which to name new Quadrant gates (you can choose any of 'Q#', where the # increments for all new quads for a subset; the stain names (e.g., CD3+CD4-); a prefix that is asked for when the quads are created).

Bug fixes.

• Correct sorting of file names that have the format 'name.#.#' (like abc.040602.001)
• Line color of histogram was sometimes forgotten when the graphic info dialog was opened in Layout Editor
• Distribute horizontally in Layout Editor works even if items are too close together
• Selecting 'About FlowJo' no longer puts application in Demonstration mode
• Layout items now correctly show cell counts after batching
• Resizing Proliferation window properly draws the model histograms
• Copy table from Multisample compare platform fixed
• Inserting keywords for specific samples in Layout Editor Text chooses correct sample (was one off)
• Occasionally, 10.e-4 would be displayed as 10.e-3 for frequency in Workspace
• Correctly displays time parameter for BD files using the 'TimeTicks' keyword
• Copying from layouts no longer clips the right or top edge of certain graphs
• Fix Copy/Paste of a single textbox in the Layout Editor creates a new text box, not a picture of a text box
• Group-synchronized gates don't crash if more than 1 is simultaneously changed (like quadrants)
• Layout text boxes with a line width of 'none' will not show the box line in printouts
• Fixed obscure bug where rarely a Layout textbox could lose the last statistic in its text
• Layouts with statistics tables won't crash during batching
• Open Graph Sets will properly tile windows (and resize if necessary) on the screen
• Holding down 'option' key while selecting a sample in the 'go to sample' menu button on graph windows will advance all open graph windows to that sample
• Fixed a memory allocation bug that could cause apparently random crashes
• Reading old layouts with legend overlays will now properly initialize the legends instead of leaving them blank
• Changing the threshold value for a Kinetics analysis forces an immediate recalculation
• Thick lines in the layout editor are better handled (selection and arrows)
• Avoid page breaks in the Layout Editor will not cause a graph and its annotation or legend to be put on different pages
• Create group window: Removing a criterion clears it; complex formulas doing comparisons on string values now work
• Clicking escape while drawing a gate now cancels the gate creation
• Selection of gate constraints in Cell Cycle platform was corrected
• Histogram gate lines in layout editor (and exported graphs) don't extend beyond edge of graph
• Adding a statistic to a group based on a parameter that has a ')' in its name no longer crashes
• Putting commas in custom table column names no longer alters the formatting of table headings
• Dragging a PICT file into the Layout Editor now refreshes the view immediately
• Tables dragged into the Layout Editor now automatically refresh statistics when gates are moved
• Double-click timing to open graphs from the workspace window restored to proper functioning
• Changing a gate does not reset the Layout Editor iteration to 'off'
• Only the 17th parameter in a file could not be compensated: now it can!
• Scrolling while entering table editor custom headings improved
• Fixed: Changing a gate does not reset the Layout Editor iteration to 'off'
• Fixed: You can again edit custom column headings in Layout Editor statistic Tables
• Fixed: Scrolling of window while editing custom column headings in Table Editor
• Fixed: You can now compensate the 17th parameter in a data file
• Fixed bugs that crept up in beta versions:
• Contextual menu for gates restored
• Group-based synchronizing multiple gates (like quads) fixed

September 10th 2004, Version 6.1 released
SUMMARY OF CHANGES SINCE 6.0

Important Notes:

Workspaces saved in FlowJo v6.1 cannot be directly opened with earlier versions of FlowJo.  However, you can save a workspace as XML (File->Save As XML...); that file can then be opened in FlowJo v6.0.  Please see notes on XML under "Workspace" below.

New Features:

General

• Version Locking.  You can now "lock" a version of FlowJo on a given computer (which will apply to all users on that computer).  This lock ensures that no other version of FlowJo will be used for data analysis on that computer.  This feature is useful for laboratories who have validated a given version of FlowJo and want to ensure that no analysis is done by a different version.  To "Lock" a given version, choose the Lock command under the file menu, enter an Administrator username and password, and then follow the instructions.  Locking can be performed in a password-protected fashion (if desired).  NOTE: "Locking" a FlowJo will not prevent versions 6.0 (or 4.x) from launching, because they cannot check for this feature.  However, these older versions of FlowJo cannot read workspaces saved by this (or future) versions of FlowJo, so there is no risk of analysis by the wrong version.  NOTE: Because the "lock" feature will apply to all users on that computer, you must have Administrator privileges.  When you choose "Lock", you will be asked by the Macintosh operating system to authenticate a Username with Administration privileges.

Workspace

• Complex Group Definitions.  Sample Groups can now be defined based on arbitrarily complex combinations of keyword information.  In the Group Definition Dialog, you can convert current keyword restrictions to a formula, and continue to extend this formula to be as complex as you wish!  Formulas can contain complex string manipulations, so you can search for substrings within keyword values, etc.
• Opening sets of graphs.  Through the preferences, you can now define one or more "Graph Sets".  These are predefined sets of graphs (with specific X and Y parameters defined for each).  Any defined graph sets will appear in the contextual menu when you click on a sample or subset; choosing that Graph Set will cause one window for each defined parameter pair to be opened for that sample.  This allows you to predefine combination of parameters to view simultaneously for each subset.  (Use "option-Next" or "option-previous" to move all graph windows forward to the next or previous sample simultaneously).
• Copying Boolean gates will now notify you if the subsets on which they depend are not found in the destination.  In addition, if you delete a subset on which a Boolean gate depends, you will also be notified that this is the case.
• FlowJo will now read badly-formatted legacy Cytomics FCS files.
• XML Workspaces:  Versions 6.x give you the option to save as XML (and can read such workspaces as well).  At this time, Tree Star does not guarantee that the XML format will be compatible with future versions of FlowJo; we are in the process of evolving this format to be consistent with the PC version.  If you have suggestions or questions regarding the FlowJo XML format and/or future plans, please contact us!

Statistics

• You can now calculate the standard deviation ("SD") on any parameter.
• When adding a statistic calculated on a parameter (e.g., mean, CV, SD), you can choose to add that statistic to all parameters, all fluorescence parameters, or all compensated parameters at once, by choosing those in the parameter menu of the statistics dialog.
• The order of the statistics in the Statistics Dialog window was changed to group them better.

Layouts

• Printing:  For batched outputs (either direct to printer, or printing from the Preview output), you now have the option to use the original pagination as defined in the Layout Editor.  This means that you can print batch reports where each panel crosses page boundaries—for example, if you spread the page boundaries such that each iteration of the batch is on 3 pages, you will get 3 pages times the number of samples in the group for your output.  To choose this option, select "Use Layout's Pagination" where offered.

• Batched layouts are no longer grouped after being created.
• Keywords are better handled in layouts (particularly, for batched layouts and/or templates).
• When entering a keyword into a text box, you can select which sample for which to display the value (and single text boxes could have keywords from multiple different samples).
• You can now set the text style for axis labels differently from axis numbers (see Preferences); you can make the axis text as large as you wish and it will no longer be "clipped" (see Bug fixes).

Graphs

• There is a new contextual menu popup for when you click on a gate in a graph window that allows you to perform operations on the selected gate.
• The "Create Manual Gate" interface has been extended to allow you to create a series of gates that are adjacent (non-overlapping) across the entire range of the displayed parameters.  The series can be specified to step by equal channel numbers, equal intensity levels, or equal percentile levels.

New Preferences

• Gates and Graphs preferences were split into separate panels.
• A preference can be specified for how FlowJo exports histograms as graphs.  The default is to export as polygons, which is useful for changing their characteristics; this works well in programs like PowerPoint.  Deneba's Canvas does not interpret Mac PICT polygons properly; thus, changing this preference will cause FlowJo to export histogram graphs as connected lines.  This makes it show up properly in Canvas (and other programs), but the histograms cannot then be filled in those programs (exporting of filled histograms will still work from FlowJo in all cases).
• Graph Sets can be defined in order to simultaneously open multiple graphs; these sets can be chosen from the contextual menu when you click on a sample or gate in the workspace.
• The text style for layout editor graph axes labels can now be defined differently than that for the axes numbers—you can have large axes labels!
• The location of FlowJo's Cache folder for transferring data from remote servers can now be specified to be either in the User's directory (default) or in the local shared directory.  The latter is preferable for computers where User directories are kept on remote computers.
• The Preference for Hiding or Moving "clipped" axes numbers has been removed (see bug fixes below).

Miscellaneous

• You can now zoom in and out of graphics in the SciBook.
• Workspaces saved with many SciBook entries now read in faster.
• A new option for Groups can be set ("Allow stain mismatch") that forces the group to add analyses and gates even if the stain names for the parameters are different than for the original sample's analyses.
• FlowJo now corrects the improperly formatted compensation matrix in Summit v4.0 data files.
• When files are moved and need to be found by FlowJo, you now have the option to ask FlowJo to perform an exhaustive search of all mounted disks for the data file (this may take several seconds).
• The text color for Print output headers and footers can be set in the Preferences.
• New quadrant gates will ignore the Preference for gate annotation position; the frequency will always be placed in the graph corners.
• The "Edit Annotation" menu item has been removed (annotations can be edited by simply double-clicking on them in the workspace).
• If you misidentify a file that has been moved, FlowJo will show you the name of the file before asking you again to locate it.
• Layouts will now show at least one additional page, horizontally or vertically, when scrolling, to allow you to better place new objects.
• A new Preferences lets you use the Bi-exponential/Logicle transformation on uncompensated height parameters ("-H") in addition to the area parameters. Click on the button: Preferences->Workspaces->DefineÉ (32 bit options).

Bug fixes.

Layout Editor

• Axes labels and numbers are no longer clipped in the Layout Editor or upon export to other programs. Finally!  You can now have axes labels & numbers as large as you want!
• Layout Statistics Table tool doesn't crash if there are no samples in the workspace
• Overlaid histograms with auto-scaling have exactly correct Y axis values (previously, these "relative" numbers were off by 4%).
• Text boxes in the Layout Editor can have braces in them again "{}".
• Gate frequencies are not duplicated underneath the graph (after ungrouping an exported layout).
• Printing a specified range of pages from the Layout View now works (previously, would print all pages).

Workspace

• Double-clicking in the workspace sometimes didn't open a graph; this now works.  (Although if you double-click too fast, it may not).
• You can now delete a keyword value in the workspace and leave it blank.
• The Time parameter is properly retained on saved and re-opened workspaces.
• Italicized text in the workspace and Table output is better displayed on-screen (not clipped).
• A crash could occur if graphs and statistics were selected in the workspace, and the contextual menu "Open graphs" was selected; this was fixed.
• Workspace mousing (click & drag, etc) is more responsive.
• Save As Template does not crash when the preference for incrementing workspace versions is selected.
• Occasional mis-sorting (or poor sorting) of samples fixed.
• No more unnecessary recalculations of subset gates that occurred in some circumstances (despite a Preferences setting of "Only recalculate when needed").
• Concatenating events from different 32-bit data files (e.g., DiVa data) is now handled properly.

Graphs & Gating

• Autogating will now place the resulting gate properly in windows that are not the standard size.
• Autogating tool is not active until the data has been loaded and displayed.
• Autogating will occur on the correct parameters after axes are changed.
• Boolean gates will immediately recalculate upon creation, if the Preferences are set for "immediate recalculate."
• Boolean gates work more reliably when copied from sample to sample.
• FlowJo will not crash if you choose to create Boolean gates on a group of selected items including Statistics.
• Possible bug causing crashes during gate adjustment fixed?

Platforms

• Contextual menu for graphs & gates no longer appears for Platforms (causing a crash)
• Cell cycle and Proliferation platforms no longer crash if something happens to invalidate them during their computation (like a parent gate moving).
• Cell Cycle and Proliferation: Copy table to text will always work now.

Miscellaneous

• The "Application Lock" contact information and hint information are no longer switched for new Lock commands.
• After batching a layout, the graphics are properly reset to the "off" iteration state (previously, they were reset to show the first sample in the current group rather than their original state).
• Layout axes will now correctly adopt the color set by the Preferences.
• Layout overlay graphs will obey the specified background color (including transparency).
• Gate trees added by groups (i.e., when you add samples to a workspace) would not add an entire tree if any gate was based on parameters with stains that did not match the original; now only the mismatched gates are not added (unless the group option is to allow mismatches--see new feature above)
• Coulter ".HST" files are properly read and loaded.
• Cell cycle: The remove debris, remove doublets no longer crashes
• If a Boolean gate was opened without first opening the other gates, then the Boolean gate could show the wrong events.
• Fixed crash that could occur on very first use of autogate tool on first dotplot graph.
• Fixed: Opening graph sets places graph window below the menu bar.
• Fixed: Contextual menus in the workspace window show the platforms & Boolean gate options.

July 5th 2004, Version 6.0 released
SUMMARY OF CHANGES SINCE 4.6.1

Important Notes:

• Workspaces saved in version 6.0 cannot be opened with FlowJo v4.6

Major New Features (click here for more information)

Autogate tool. Point… click… gate. It shouldn’t be any harder. And, with FlowJo’s new Autogating tool, it can’t get easier! FlowJo’s autogating tool has an almost prescient knowledge of where you want the gate—an effect made possible by the instantaneous feedback of where the gate will be as you mouse over the graph! Choose the new autogating tool in the graph window, or simply hold down ctrl-option as you move the mouse over any bivariate graphic. (Note: if you hold down the mouse button as you click to create the gate, you can then track the mouse to inflate or deflate the gate to size it with even more precision!)

SciBook. The SciBook is a place where you can store pictures, text, tables, or custom annotations. The SciBook is stored with the workspace; you can use it as a permanent record of your analyses, as a temporary holding place for outputs, or just to keep notes for yourself. You can paste graphics, text, or tables from anywhere in FlowJo into the SciBook (the Graph window, Platforms windows, Layout Editor, Batch Layout View, and Table View all have buttons that will directly copy their contents into the SciBook). Contents in the SciBook do not change-therefore, if you change a gate, delete a sample, or make other modifications, the SciBook contents will not be modified. Once in the SciBook, you can copy into other programs, print, or simply just view. The SciBook gives you an interface to add comments about each item that you insert, and records the date and time that the item was inserted.

The SciBook is meant to evolve; we look forward to your comments regarding which features you use the most, what features you would like to see in the future, and what you do (and don't) like about it!

Backgating Analysis. Validating gate positions is one of the most important aspects of multiparameter Flow Cytometry. FlowJo now provides you with a tool to view the effect of every gate on your final gated population! In any graph window for a subset that is gated more than once, you can choose “Backgating Analysis…” under the Graph menu (or, click on the Backgating button that appears near the top right of the window). FlowJo shows you a window where each level in the gating hierarchy is displayed, with the final gated population overlaid at each level. This overlay is a backgating overlay: in other words, it shows what the final gated population would look like without that level’s gate! Therefore, you can quickly scan to see if any one of your gates is positioned incorrectly. The backgating display can be printed, copied, or saved in the SciBook (see below). Note that the Layout Editor can also show a version of the backgating analysis, by choosing “Show Ancestry” and selecting the “Backgating” option for a graph

Boolean gates. You can now create a gate which is a Boolean combination of any existing gates (i.e., "and", "or", and "not" combinations). Boolean gates are "live", meaning that they will reflect any changes to the gates on which they depend. FlowJo can also automatically create a series of Boolean gates that represent all combinations (plus and minus) of a set of gates. For example, if you have a sample which was stained on different parameters for IL2, TNF, and IFNg, you can create a positive gate for each of the three cytokines (for example, a histogram gate). Then use FlowJo's Boolean Combination to create the 8 gates that represent all combinations of expression of the cytokines (e.g., IL2+TNF+IFNg+, IL2+TNF-IFNg-, etc.). To create or edit Boolean gates, choose the parent subset(s) and select the menu item "Boolean gates..." under the "Platforms" menu.

Derived parameters. You may now create a derived (computed) parameter that is any algebraic combination of existing parameters. FlowJo allows you to specify arbitrarily complex formulas, and includes a wide variety of mathematical functions that you can use to transform the data. This platform can be used in FRET calculations to create a parameter that reflects the efficiency of resonance energy transfer.

Layout Editor: New Statistics Table. FlowJo now renders tables of statistics in a beautiful and highly customizable way in the Layout Editor. You can copy tables (either by dragging them from the Table Template Editor to use with Batch iteration, or directly from a completed table). Or, you can create them from scratch, adding your own statistics and keywords and other information. Statistics Tables in the Layout Editor come in two flavors: (1) Each row is from a single sample: this is like the output of the table editor, where statistics for subsets or the whole sample are all represented on the same row; (2) Each row can have subsets from the same sample: this allows you to view a number of statistics (e.g., MFI) on many subsets from a single sample in column format. Note that you can copy Layout Statistics Tables directly to spreadsheets (as tabular data) or to graphics programs (as their graphical representation).

Layout Editor: Overlay legends. The legends for overlays in the Layout Editor can now be customized to show statistics, keywords, or other pertinent information from each subset that is overlayed in the graphic. Simply double click on the legend to specify what should be shown.

Layout Editor: Iterate by "nth" sample. Sometimes you just want to do batch iteration where the order in which you collected the samples defines how you want the output to appear-for example, every three tubes comes from a single sample, where the first of the three is a control, and the other two are two different panels. FlowJo lets you easily specify such a design by using the "Iterate by nth" option. (Choose "Iteration options" under the layout menu). In this example, you would iterate by "every 3rd sample", telling FlowJo to group tubes into samples of 3. This significantly simplifies batch operations. All new Layouts are created with the “Iteration by nth” (n = 1, i.e., every sample) by default.

Templates: Improved functionality. This "Iterate by nth" Layout Editor feature also means that Template Workspaces that contain batched layouts will work well. A batched layout is now automatically defined to have the appropriate iterator (and discriminator) values; when you batch a layout over every sample and save it as a new layout, the new layout is automatically defined to iterate by "nth", where "n" is the number of tubes in the original batch.

In addition, sample identification is significantly improved in FlowJo. This means that Template workspaces will fill out layout graphs as expected-FlowJo tries very hard to overcome any ambiguities that might arise because of multiple samples fitting the criterion for a single graphic.

Other New Features.

General

• The FlowJo icon is properly displayed on all workspace files (that have names ending in ".jo", ".ws", or ".flowjo").
• There are new “Help” menu items to provide quick links to documentation and contact information.
• Lists of FCS Keywords are sorted better, with user-defined keywords appearing first, and standard FCS keywords arranged in better order.

Workspace

• The workspace auto scrolls to show the sample corresponding to the front-most graph window.
• The number of samples in a group is now shown in the first column after the name, not aligned with the "Statistic" column.
• Template workspaces are now saved with the default extension of “.jot”.
• While editing a keyword value, you can paste a selection (copied from another program) that has a series of values (tab- or carriage-return-delimited, as from Excel). FlowJo will fill out successive rows with these values.
• You can now ctrl-click (or click-and-wait) on analyses in the sample list to show a menu of actions appropriate for the selected items.
• The column resizing button has been moved from the top of the workspace window to the middle bar.
• $PnE keywords (log/linear scaling of parameters) can now be edited.
• When sorting on keyword values, FlowJo looks to see whether a keyword value is numeric, alphabetic, or mixed, and sorts appropriately. For mixed values, FlowJo will sort “A-2” before “A-10” (even though this is not alphabetical); it recognizes the presence of both strings of letters and numbers and treats them independently when determining sort order.

Graph Window

• Graph windows now use the "ctrl-click" (or click-and-wait) to show a menu to change the graph attributes (graph type, options, etc.), similar to the Layout Editor graphs.
• Cloning a graph window (“+” button, or “Open New Window” menu) now creates a new window that has exactly the same settings.
• You can specify (for dot plots only) the number of events to display, on a graph-by-graph basis.

Platforms

• Compensation: When editing a compensation matrix, you can now copy it (to the clipboard) or print it.
• Cell cycle: There is a new preference to choose a Pulse Area parameter as the default for new analyses, and new preferences to control the default modeling characteristics.
• Cell cycle: The cursor coordinates are now shown in the window.
• Proliferation: The modeling algorithm has been improved to better find good fits. (The formula for the model has not changed, only the approach to the fit).
• Derived Parameters: Older derived parameters for Ratios, Gain/Offset are now automatically converted to Algebraic derived parameters with the correct formula shown.
• Derived Parameters: You can double click a previously-defined algebraic parameter to edit it.

Layouts

• Overlay Legends: You can now click to select any row in the legend; press the "delete" key to remove it from the overlay; drag to re-order the overlay.
• Show ancestry plots: You can choose to make all ancestry graphs show at equivalent size, rather than scaling them to fit in a single row or column.
• Text boxes: If you select a text box and choose "Copy", then the contents of the text box are put on the clipboard so you can paste them into other applications.
• Arrows: If you create a line with arrow heads, you can now specify the shape of the arrow head to customize its appearance.
• Graphs: You can now specify the number of events to display (dot plots only) on a graph-by-graph basis.
• Graph Item Annotation: The dialog window in which you can specify custom contents for a graphic item annotation box now has a menu to choose keywords to insert into the annotation text. Also, graphic item annotations can be edited for multiple graphs simultaneously from the "multi-item attribute editor" window.
• Batch Iteration Dialog: The dialog window that appears when you create a new Batch output has been made easier to understand; the iteration controls have been removed from this window (to specify iteration options, you must choose them via the "Iteration options" menu item).
• Text boxes, annotations: In addition to specifying FJML commands for items like statistics ("<Stat ...>") and Keywords ("<Keyword ...>"), you can also specify information about any given subset using the "Property" command. Properties that can be displayed include: "Comment" (any comment in the workspace for that subset), "GateName" (the name of the subset), "SampleName" (the name of the sample), and "Path" (the full gating path for that subset). For example, "<Prop SampleName>" will be translated into the subset's sample name.

Tables

• Editing column names: While editing a column name, you can paste a selection (copied from another program) that has a series of values (tab- or carriage-return-delimited, as from Excel). FlowJo will fill out successive rows with these values.
• Hold down the option key while clicking on the "Copy to clipboard" button and a transposed version of the table is copied onto the clipboard.

New Preferences

• Pulse-area parameter should be the default for Cell cycle analyses.
• Newly-created histogram overlays in the Layout Editor should be scaled to a fixed value by default (rather than scale-to-max).
• Specify the default line styles for histogram overlays.
• Default graph window size for newly-created graphs (brought back from Version 3.x of FlowJo!).
• Text traits for gate frequencies shown in graphs in the Layout Editor
• Text traits for headers and footers in printouts
• Specify the default statistic that is added to the Table Editor when a gate is dragged to the Table.
• Specify whether axis numbers, on graphs that are near the edge and would appear clipped as a consequence, should be moved slightly or hidden.
• Cell cycle platform preferences can be defined.
• Proliferation platform model display preferences can be defined.

Bug fixes

• Suggested output file names are chosen better.
• Sample Information window can be resized to show keyword values better.
• Deleting a sample from the workspace will close its graph window.
• Create Value Series (while editing a keyword value) now obeys the maximum number of values to fill in, and better remembers the previous values you used for it.
• Cell cycle and Proliferation analyses immediately recompute a model if a parent gate changes.
• Proliferation: If create a constraining gate to position the undivided peak, you can now select it without having to close the window first.
• Layout graph items with oversized text (from the preferences) no longer randomly resize themselves. However, large text will still be clipped if it is too big. (You can choose to hide ticks or axis numbers to provide additional space for label text).
• Cell cycle: Remove apparent constraints that allowed positioning of peaks by absolute values (this was never a possibility).
• Cell cycle: Fixed synchronous peak constraints for DJF model.
• Cell cycle: copying analyses to groups or other samples retains the preferences for how models are displayed
• Workspace: Fixed editing a keyword value for workspaces with >1000 samples.
• Kinetics: Fixed an issue that could cause a crash for files with corrupt data at the end of the time series.
• Layout Editor: Multiple graphs dropped in from the workspace are now positioned better (so as not to overlap).
• Workspace column resizing no longer occasionally resizes the window height.