This panel is used to quickly modify color and brightness of the image using Look-Up Tables (LUTs). Simply drag the left or the right triangle to adjust the input intensity range. Lift or drop the yellow point to adjust the Gamma parameter. Double-click into the graph area to zoom the histogram so that the “high” and “low” limits are distinguishable. For more details about LUTs, please see LUTs - Non-destructive Image Enhancement.

Enable/Disable LUTs

If turned on, LUTs are applied to the current image.

Keep Auto LUTs

Click this button to run the Auto LUTs permanently on the live image (see below).

Auto LUTs

This button adjusts the white slider position of all channels automatically with the purpose to enhance the image reasonably.

Reset LUTs

Discards all LUTs settings and turns LUTs OFF.

Settings

Opens the Auto Scale Settings where the Low and High fields determine how many of all pixels of the picture are left outside the sliders when Auto LUTs are applied (0-10%). Check Use Black Level to ensure that the black slider is influenced by the auto scale functions.

Pixel saturation indication

If turned on, all pixels in the image with values reaching maximal values are highlighted red.

In Color / Mono

Switches the single channel image between the color and monochrome view mode.

Zoom

Use mouse wheel to zoom the graph. This button jumps to the last zoomed position.

Tip

Double-click in the graph to zoom the input intensity range.

ROI Statistics

This panel shows statistics - Mean (Min, Max) - for the drawn ROI.

Draw Rectangle

Click and drag in the image to draw a rectangular ROI. To move the ROI, click on its center and move it around. Drag its edge to resize it.

Draw Circle

Click and drag in the image to draw a circular ROI. To move the ROI, click on its center and move it around. Drag its edge to resize it.

Draw Polygon

Click points in the image to define the polygonal ROI. Use the secondary mouse button or a double-click to place the last node of the polygon and close the object. To draw a free-hand shape, hold down the primary mouse button. To move the ROI, click on its center and move it around.

Reset

Clears all ROIs in the image.

Measure

Measures the intensity of the whole image or inside the drawn ROI(s) on the selected channel and shows the measured data in the table above. The intensity measurement against the image dimension (Z-Stack, Timelapse) is displayed in the graph below. Select the measured channel from the drop-down menu on the left.

Auto Refresh

If checked, this function automatically measures intensities right after ROI changes are made.

Intensity Profile

This panel is used for interactive intensity measurements. An arrow is shown in the image indicating the profile cross section. This cross section intensity is shown in the graph. The profile line can be resized by dragging its ends by mouse arbitrarily. It is always a straight line. Context menu over the arrow allows you to Reverse Direction of the arrow, edit Line Properties (color, width and style), change the Intensity Profile Properties (described below) and to choose which channel is used for the measurement.

Draw Free Line

Click and drag directly in the image to draw a profile line.

Free Line

Insert a profile line which can be freely positioned.

Vertical Line

Inserts a vertically fixed profile line.

Horizontal Line

Inserts a horizontally fixed profile line.

FWHM

This tool measures the Full Width at Half Maximum (FWHM) value on the given graph range.

1. Click into the graph to define the maximum.

2. Click again into the graph to define the minimum.

3. Half of this interval defines the value in which the graph width will be measured.

Figure 29. 

Note

If the minimum is defined in a different part of graph (e.g.: different peak), the width is measured on the peak where the maximum value was defined.

Auto FWHM

Measures the FWHM value automatically once this button is clicked.

Realtime Auto FWHM

Measures the FWHM value continually on the live image.

Vertical Autoscale

Zooms the graph so that both minumum and maximum values are visible.

Reset

Resets any zoom changes.

Settings

Opens the Intensity Profile Properties dialog window where you can set the graph interpolation type and the width of the line neighbourhood used for the visualization of the mean/max intensity values.

Measurements

This panel enables the user to perform simple measurements on the current image.

Pointing Tool

Use this tool for selecting measurement objects in the image.

Simple Line

Line length is measured. Click and hold the primary mouse button over the starting position and drag to the ending position and release the button. Confirm the object by the secondary mouse click.

Polyline

Polyline length is measured. A polyline consists of several line segments. Use the primary mouse button to draw the line segments. Each mouse-click creates a new node. Complete the total length definition by the clicking the secondary mouse button.

Polygon

EqDiameter of a polygon is measured. Draw a polygon by clicking the primary mouse button inside the image. The secondary mouse button or a double-click will place the last node of the polygon and close the object. To draw a free-hand shape, hold down the primary mouse button. Confirm the object by the secondary mouse click.

Autodetect Polygon

EqDiameter of a detected object is measured. The system automatically detects a homogeneous area around the clicked point.

Count

Click inside the image to insert counted points. A red cross is inserted to the image after each click. Right-click to the image to finish counting.

Clear Overlay

Deletes all measurement objects from the image.

Clear All

Deletes both the measurement objects and the measurement table.

Export Data

Exports the results table into a .csv file.

Live

Displays the live camera image according to the camera settings (the button changes to Freeze when the live image is shown).

Smart Live

The Smart Live function can be enabled in the Configure... menu. It works only on Fluorescence experiments. It checks the hardware status, including stage movement and changes in the light power. Then it captures all channels if a hardware change affecting all channels has been made beforehand, such as a movement in XY or Z. However, if a change affecting only one channel has been made (e.g., a change in the power of the associated laser line), the other channels remain unchanged. Once the experiment channel(s) are captured, the light is deactivated. This ensures the convenient access to a multichannel live image without causing bleaching to the sample.

Capture

Acquires a single image from the camera.

Auto Focus

Performs the fastest possible auto focus. There are two focus ranges available, short and long. By default, the short range is used. Hold the Shift key and click on the Auto Focus button to use the long range ().

The criterion for focus detection is dynamically changed based on the selected light path (modality).

FLUO

A criterion suitable for fluorescence samples.

BRF

Suitable for brightfield samples. Brightfield auto focus works based on the values set in Devices > Service Settings .

Cells.ai Criterion

An AI-based algorithm suitable for brightfield cell samples. A specific hardware configuration must be active for this criterion to be available. To switch between the brightfield criterions, use the context menu over the Auto Focus button. The Enable Context Menus option must be selected in the Configure... menu.

The AI neural network has been trained and is intended to be used with the following cell lines:

• A431

• BSC_1

• CHOK_1

• Cos7

• Day1_iPS_neuro

• Day4_iPS_neuro

• Day7_iPS_neuro

• Hela

• HepG2

• HT29 J774_1

• Neuro2a

Note

The Z lens piezo in the system functions with the auto focus only when the register key (HKEY_LOCAL_MACHINE\SOFTWARE\Laboratory Imaging\Misc\DeviceManagerv2\NosepiecePositionForObjectivePiezo) indicates the nosepiece position where the lens piezo is installed and when it aligns with the current nosepiece position. Please create the registry key “NosepiecePositionForObjectivePiezo” as “DWORD(32bit)”. Set the value in hexadecimal, Position #1 = “0”, Position #2 = “1” ... Position #6 = “5”.

Autosignal

Automatically finds the optimal camera exposure and light power using an AI algorithm. For brightfield the function adjusts acquisition parameters to utilize the full dynamic range of the camera. For Fluorescence the function adjusts parameters to achieve the required image quality by prioritizing the Signal-to-Noise Ratio (SNR). This SNR-based optimization requires a supported GPU. If a supported GPU is not detected, a warning will be displayed once after clicking the Autosignal button, and the system will fall back to behavior utilizing the full dynamic range.

Autosignal.ai has two options for different algorithm strategies:

Live Sample

Tries to be more sensitive to the sample, e.g. by limiting the maximum light power.

Fixed Sample

Prefers to use more light with short exposures.

To switch between Live Sample and Fixed Sample algorithm, go to the Configure... menu.

Deshading

(requires: Local Option)

Turns on the shading correction. The Define Automatically/Define Manually button appears in the experiment area. Click on the arrow to switch between these two modes and then click on the button itself to define the shading. The definitions are also available in the context menu over the Deshading button from where you can open the Shading Correction Panel (Shading Correction with the Local Option module).

Distortion Correction

(requires: Local Option)

This function is applied on large images only where it corrects the geometrical image distortion to eliminate duplicated structures in overlapping image tiles. It is closely described in Distortion Correction.ai. Click on the arrow next to the icon to Calibrate Distortion Correction.ai or click Distortion Correction.ai for information on how to adjust the live image for the large image acquisition and proper distortion correction.

Auto All

Performs Auto Focus and Autosignal in one operation on all channels.

ND Acquire

There are two types of ND acquisition available:

Define/Run ND Acquisition

Opens the standard ND Acquisition window (Combined ND Acquisition).

Define/Run ND Acquisition Wizard

Runs the Addons > ND Acquisition > ND Acquisition Wizard command used for a step by step ND acquisition.

The selected acquisition type is opened by clicking on the ND Acquire button itself. Switch between the two acquisitions using the the secondary mouse button over the ND Acquire button if Enable Context Menus is checked in the Configure... menu or switch it directly in the menu.

Large Image

Runs the Addons > Large Images > Scanning Wizard command used for a step-by-step large image acquisition.

Configure...

Opens the Acquisition Settings window where settings for the Live, Auto Focus, Autosignal and ND Acquire buttons are adjusted. User can enable or disable context menus and tool tips, and customize which buttons are displayed on the toolbar.

Save

Saves the current image.

Save As...

Opens a standard dialog for saving image files.

Open

Displays a standard opening dialog for opening image files.

Each opened image is shown as a tab. Click on the tab to activate the image or click X to close it.

The navigation tools at the bottom of the image are the same for both layouts and are described here: Control Bar. The top toolbar buttons are described here.

Undo

Moves one step back through the history of changes.

Redo

Moves one step forth through the history of changes.

Auto Calibrate

Run the live image and click this button to automatically calibrate your image using the current hardware setup.

Mouse XY

This feature requires a motorized XY stage. If turned on, the mouse cursor on a live image changes and you can control the XY stage by dragging the live image. Before using this function, it is recommended to perform automatic calibration.

Split Components

Turns on a special view, where color channels of the image are displayed separately (tiled). This mode is supported in the modes Main View, Slices View and Volume View.

Main View

Default viewing mode.

Show Slices View

This viewing mode displays orthogonal XY, XZ, and YZ projections of the image sequence. It requires a Z or T dimension.

Volume View

This view mode creates a 3D model of the acquired object. It requires a Z dimension.

Click Open Volume Options to adjust options for the 3D model. See also Volume Viewer.

Show Tiled View

This view mode displays frames of the selected dimension arranged one next to other. It requires a Z, T or XY dimension. One or two dimensions can be viewed at a time.

Show Maximum Intensity Projection

This function analyses all frames of one dimension and picks pixels with the maximum intensity values. These pixels are used in the resulting image. It requires a Z or T dimension.

Show Scale

Displays a line scale which can be moved around the image using the mouse. Context menu over the scale enables the user to Burn Scale into the image permanently, move it to the Default Scale Position, Lock Scale Position or adjust Scale Properties ( View > Layers Properties > Scale Properties).

Tip

Use / to show or hide the scale.

Show Graticule

Displays a graticule layer over the image for quick and approximate measurements. Graticules behave like adjustable floating rulers. User can simply align a graticule with the measured object and read the distance value. Click on the line and drag it to move the graticule. Context menu over the graticule enables the user to move the graticule to the center (Move to Center) and adjust the Graticule Properties.

Tip

Use the * key shortcut to show or hide the graticule layer.

Show Binary

Displays or hides the binary layer in the image.

Tip

Use Ctrl + Up/Down to decrease/increase the binary layer transparency.

Show ROIs

Displays/hides any ROIs in the image. The ROI Statistics panel is shown below the LUTs control.

Show Intensity Profile

Displays/hides the intensity profile in the image. The Intensity Profile panel is shown on the left.

Automatic Deconvolution

Runs automatic deconvolution on the current image.

See: Automatic Deconvolution.

Create EDF Focused Document

Creates a new simple focused image from an ND file. See Extended Depth of Focus.

Show EDF Anaglyph

Enables/disables the anaglyph 3D view. New options appear in the toolbar - you can define the position of the first frame (Lowest/Highest), Style (the color combination used for anaglyph image), Effect (Descend into the Screen / Rise from the Screen) and Z-zoom (define the percentage).

Show EDF 3D Surface View

Opens the 3D surface view of the image. See Applications > EDF > Show Surface View .

Show EDF Z-Profile

Opens the EDF Z-profile panel with the Z-profile graph of the image. See View > Analysis Controls > EDF Z-Profile .

Show EDF Z-Map

Creates a new image displaying the EDF Z-map. See Applications > EDF > Show Z-Map Image.

Compare View

Arranges the two last viewed images next to each other and turns on their dimension and zoom synchronization.

Full Screen

Hides all screen elements such as menus, toolbars, etc. and maximizes the image. Only some basic tools for controlling the image stay displayed. Hit Esc to return to the main view.

Fit to Screen

Adjusts the zoom factor to view exactly the whole image as big as possible.

Zoom 1:1

Adjusts the zoom factor so that one pixel of the image matches one pixel of your monitor.

Zoom

Select a zoom factor from the drop-down menu.

Single Image

Displays one image at a time.

Tile to Columns

Arranges all images as tiles ordered to columns.

Tile to Rows

Arranges all images as tiles ordered to rows.

Synchronize Views

If turned on, the views of all currently opened images are synchronized (Z position, zoom, etc.). See also Synchronizer.

Show Tiles

Opens a panel with all currently opened images shown as thumbnails. Click on an image thumbnail to activate the image.