ZEISS Knowledge Base

This modules enables the acquisition of time series (time-lapse) images with definition of intervals between images, total acquisition duration and number of time points. Time series can be started and stopped manually, at fixed times, after a waiting period or by input (trigger) signal, to analyze images already acquired or change the experiment parameters. The experiment size is limited only by free space on the hard drive and the images are acquired at maximum possible speed.

Acquiring Time Series Images

To set up Time Series experiments, you need to have licensed this functionality and activated it under Tools > Toolkit Manager. For LSM systems, this functionality is part of the system license.
You have set up a new experiment with at least one defined channel and adjusted focus and exposure time correctly, see also Set up a new experiment and Set up multi-channel experiments.
You are on Acquisition tab.

In the Acquisition Dimensions section, activate Time Series .
The Time Series tool is displayed in the Left Tool Area.
Open the Time Series tool.
Set the Duration of your time series. You are able to select an interval (days, hours, minutes, seconds, milliseconds) or cycles (1-n), e.g. 10 cycles.
Set the Interval of your time series.
Click Start Experiment.

The time series experiment starts. You have successfully set up a time series experiment.

You can display the individual images via the Time slider on the Dimensions tab.

The shortest possible interval is calculated by performing a blind experiment. The camera exposure time, number of steps of a z-stack and the number of acquisition channels are taken into consideration in the calculation. Depending on the number of z-stacks and channels and whether long exposure times have been set, it may take some time to calculate the shortest time interval.

Use time series to acquire an image series consisting of a number of time points. Define, for example, the acquisition interval, the length of the experiment and other specifications to control the experiment.

Parameter		Description
Duration		Defines the minimum duration of your experiment with the slider or the input field. The dropdown list defines the number of cycles or the duration:
–	Cycles	Defines the duration in number of cycles.
–	ms	Defines the duration in milliseconds.
–	s	Defines the duration in seconds.
–	m	Defines the duration in minutes.
–	h	Defines the duration in hours.
–	d	Defines the duration in days.
As Long as Possible		Activated: The acquisition of the time series continues as long as possible, depending on the free disk space of the hard disk. For hard disks with more than 20 GB free disk space the acquisition always runs until 2 GB disk space is left. Example: An experiment using a hard disk with 250 GB of free disk space will run until 2 GB are left. The calculated/required disk space for the experiment is 248 GB. For hard disks with less than 20 GB free disk space, at least 10% disk space will be always left free. Example: An experiment using a hard disk with 15 GB of free disk space will run until 1,5 GB are left. The calculated/required disk space for the experiment is 13,5 GB.
Interval		Defines the interval from individual image to individual image in an image series. You can specify the interval to set the gap between individual time points in milliseconds, minutes, hours or days. For LSM it is possible to change the interval during time series acquisition with the parameter Interval Time.
Acquire with High Speed		Only visible for Lightfield 4D acquisition. Activated: Acquires the image with high speed. In this mode, the images from the camera are directly streamed to the hard drive (SSD) of the computer for maximal acquisition speed. This option is automatically activated by default after setting up Lightfield 4D channels if all channels are using the same filter position in the reflector changer. If you have defined two or more Lightfield 4D tracks, the exposure time for each channel is set the same. Updating the exposure time of one channel updates it for all defined Lightfield 4D channels. Note that Interval gets disabled.
Calculated Volume Rate		Only visible for Lightfield 4D acquisition. Displays the volume rate which is calculated based on the exposure time. The maximum possible value for a time series with one channel is 80 vps. In case you have defined multiple channels, the volume rate is divided among the individual channels.
The following parameters are only visible if Show All is activated:
Use Camera Streaming if Possible		Only visible in Camera/Widefield Mode. Activated: The software tries to use the free running/streaming mode of the active camera.
Use Burst Mode if Possible		Only visible in Camera/Widefield Mode. Activated: The system buffers acquisition data in the main memory. For more information, see Burst Mode.
Measure Speed		Not available for LSM acquisition. Checks whether the experiment can be performed using the interval which is set. If the interval is too small, the shortest possible value is defined automatically for the interval.
Start/Start Next Time Slice/Stop/Pause Begin/Pause End		Defines the Start, Stop and Pause conditions of your experiment. Select the parameters for the corresponding condition from the Mode dropdown list.
–	Manual	The experiment starts immediately when clicking Start Experiment in the Experiment Manager.
–	At Time of Day	The experiment is started, stopped or paused at the entered time. Enter the desired time in the spin box/input field below the dropdown list.
–	After Delay	The experiment is only started, stopped or paused once the length of time entered has passed.
–	On Trigger	The experiment is started, stopped or paused once a TTL signal has been received.
–	After Cycles	Only available for Pause Begin. The experiment is paused after the defined number of cycles. If this function is used for a LSM time series, the acquisition will no longer be executed by the real time electronics and will be markedly slower overall.
Interval Time		Only available for LSM. Interactively changes the interval during a running time series acquisition.
–	Interval	Displays the interval value which is taken over from the current set interval.
–	Trigger In	Assigns a TriggerIN signal to change the interval.
–	Trigger Out	Assigns a TriggerOUT signal for each interval change.
–	Add	Adds a new interval.
–	Delete	Deletes the selected interval.
Set Marker		Only available for LSM. Marks events during the time series acquisition.
–	Description	Sets the name of the marker.
–	Set	Sets a marker during time series acquisition.
–	Trigger In	Selects the TriggerIn signal for setting a marker.
–	Trigger Out	If necessary, assign a TriggerOUT signal for every set marker. Note that the TriggerOut signal for the marker function consists of a high spike followed by a 3 second long TTL pulse. Within the duration of the pulse a further low spike is delivered.
–	Add	Adds a new marker.
–	Delete	Deletes the selected marker.
Interactive Switches		Adds and configures switches that can be used to execute certain actions during your experiment. To add a new switch, click . The Switch button opens a dialog to configure an interactive switch, see Interactive Switches Section.

Experiment Conditions Section

If you define times as time series acquisition conditions, these apply once for the entire experiment. This also applies to experiments that use the Experiment Designer.

The following functions are only visible if the Show All mode is activated:

You can define the Start, Stop, Pause Begin and Pause End conditions for your experiment. Start Next Time slice can only be used with trigger functionality of LSM 990. Select the parameters for the corresponding condition from the dropdown list:

Parameter	Description
Manual	The experiment is started immediately when clicking on the Start Experiment button in the Experiment Manager.
At Time of Day	The experiment is started, stopped or paused at the entered time. Enter the desired time in the spin box/input field to the right of the dropdown list.
After Delay	The experiment is only started, stopped or paused once the length of time entered has passed.
On Trigger	The experiment is started, stopped or paused once a TTL signal has been received.
Interval Time	Only available for LSM. Use this function to interactively change the interval during a running time series acquisition. Use the + button to add a new interval value which is taken over from the current set interval. Add another interval first before changing the current interval. Highlight the desired interval during the experiment to change the interval before the next image frame is scanned or assign a TriggerIN signal to change the interval. You can always assign a TriggerOUT signal for each interval change. The change of the interval is also marked in the time series acquisition. The marker is visible next to image (2D view, Gallery view, MeanROI) when the change occurred.
Set Marker	Only available for LSM. Use this function to mark events during the time series acquisition. Predefine the marker by typing in the marker name into the editing box. Either click Set or use TriggerIN to set a marker during time series acquisition. You can also assign a TriggerOUT signal for every set marker. The Set button becomes active when the acquisition is running. The marker is visible next to image (2D view, Gallery view, MeanROI) when the marker was set.

Interactive Switches Section

The following functions are only visible if the Show All mode is activated:

Here you can add and configure switches that can be used to execute certain actions during your experiment.

Left-click on a new or existing switch to open the dialog in which you can configure the button parameters:

Parameter	Description
Name	Here you can enter a name for the button.
Description	Here you can enter a description for the button.
Color	Activated: Shows a colored line at the left edge of the switch.
Color Selection	Opens the Color Selection dialog. Here you can select a color for the line at the left edge of the switch.
Action	Here you can select one of the following actions. This action will be executed when you click on the button: None Set Interval As Fast as Possible Trigger Hardware Setting Jump to previous block Jump to next block Jump to block #
Delete Switch	Deletes the selected switch.

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Time Series

Acquiring Time Series Images

Time Series Tool

Experiment Conditions Section

Interactive Switches Section

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