As discussed in our previous article on scientific camera electronic shutters, our camera range uses electronic shutters, meaning the sensor is continuously exposed with no mechanical shutter or aperture to block light to the sensor. These cameras continuously read out the sensor when in operation, and can produce an image upon request by software, which is the image you see on your computer.
In order to specifically determine when a camera produces an image, as well as controlling other factors such as exposure time, a trigger can be used to communicate with the camera. This article will cover the triggering of scientific cameras, and how triggers can be used to add precision and synchronization to your imaging, allowing different imaging components to communicate and control each other.
Triggers are typically either be produced by the camera itself (an internal trigger) or the camera can be triggered and controlled by another imaging device, typically a light source (an external trigger).
These internal and external triggers can also be categorized by their source, either produced in software or hardware. While software triggers are produced by imaging software or camera control drivers such as our PVCAM software, hardware triggers are typically electrical impulses. At Teledyne Photometrics, we use BNC coaxial cables (shown in Fig.1) for advanced hardware triggering, which sends a voltage signal in order to trigger the camera (where 0V is off, and a signal of 5V is on), known as a transistor-transistor logic (TTL) pulse.
These hardware trigger cables are often used to connect the camera to a light source so that one can control the other, the benefit here is that cameras can be synchronized with light sources, motorized stages, and/or other components of the imaging/microscopy system, to exactly determine when images are taken and how long exposures are, especially useful when using scanning or strobing light sources or with samples that rapidly bleach or decay.
Our BNC cables offer several methods for integrating with external hardware. Firstly, one end connects to the back of the camera for trigger input/output (I/O) operations, and the other end of split into seven (see Fig.1), all of which have different effects on the camera:
- Trigger In, initiates exposure and/or acquisition
- Trigger Ready Out, status indicator of whether the camera can accept another trigger
- Read Out, status indicating if the camera is currently reading out or digitizing
- Expose Out 1, output for controlling light source 1
- Expose Out 2, output for controlling light source 2
- Expose Out 3, output for controlling light source 3
- Expose Out 4, output for controlling light source 4
These cables are used with our Prime and Kinetix CMOS families and allow either the camera to be controlled by another imaging device (IN) or allow the camera to control other imaging devices (OUT). Both these input and output trigger modes are described in the next sections.
Input Trigger Modes
Using the Kinetix family as an example, there are six different trigger modes that allow control of the camera, one internal, three hardware triggers, and two software triggers, described below in that order.
This is the default mode using internal software triggering. The start of imaging is initiated in software, and each frame captured is controlled by the internal timing generators in the camera. Exposure and camera settings are set in the software prior to acquisition. This mode does not synchronize with other imaging hardware and requires them to be controlled independently through the software.
This is similar to Internal Mode but requires a hardware trigger from a trigger cable in order to start acquisition, allowing for higher precision of acquisition. Once the trigger signal is sent and acquisition is initiated, each capture is then controlled by the internal timing generators in the camera, with exposure and camera settings controlled by the software.
While Trigger-First Mode only requires a single hardware trigger to start acquiring frames, Edge Mode requires a hardware trigger for every single frame, with exposure and camera settings controlled by the software.
Similar to Edge Mode, Level Mode also requires a trigger for each frame, but the difference is that in Level Mode the hardware trigger is required to control the exposure time. This allows users the flexibility to control the exposure time for each frame. A TTL pulse is sent to the camera, and the length of time this pulse lasts will be equal to the exposure time, allowing for fine control over exposures for subsequent frames, shown in Fig.2.
Level Mode can even overlap frames, triggering subsequent frames while the initial frame is still reading out. This allows the camera to run at the maximum expected frame rate, but the rolling shutter of CMOS cameras must be paired with carefully controlled illumination to ensure there is no illumination contamination between frames. This means that for this mode, it would be best to have the same system controlling the camera exposure time as well as controlling the light source illumination cycle. This can be done with the Expose Out modes, which can output trigger signals from the camera to the light source.
Software Trigger-First Mode
This is a software trigger alternative to Trigger-First Mode, where the sequence of acquisition is started by a single software trigger, sent by the user via PVCAM, our software to communicate with Teledyne Photometrics cameras.
Software Edge Mode
This is a software trigger alternative to Edge Mode, where every new frame has to be started by an independent trigger sent by the user via PVCAM.
Output Trigger Modes
Again using the Kinetix family as an example, there are five different hardware trigger modes that allow the camera to control aspects of the imaging process, such as the light source.
First Row Mode
In this mode, the trigger signal from the camera is high only when the first row of a frame is being exposed. This means the length of the trigger from the camera is equal to the exposure time for the first row, with exposure time set in software. Due to the negligible gaps between frames, this means First Row mode trigger signal is mostly high and only drops to zero during first row readout, with lasts for only a few microseconds. First Row mode can be used as a timestamp for frames during time-lapse, or to initiate other actions that are connected to the beginning of image frame capture.
All Rows Mode
This mode is similar to First Row mode, except the trigger signal from the camera is high only when all rows on the sensor are exposing. This is useful for controlling external light sources, such that the signal detected at the camera starts and finished at the same time across all rows. This provides a Pseudo-Global Shutter imaging method, where all rows of the sensor detect signal using the same start and stop times. The duration of the trigger is equal to the time between the start of the last row’s exposure and the end of the first row’s exposure, so the timing of sequential exposures is altered in order to maintain the desired exposure time.
Rolling Shutter Mode
This mode is similar to All Rows mode, but has the goal of maintaining a set framerate (while All Rows mode will change framerate to maintain the exposure time). If the framerate and the exposure time input into the software are not greater than the take taken to readout the whole sensor, then the trigger signal won’t be sent. This mode also provides Pseudo-Global Shutter imaging from the camera and light source.
Line Output Mode
This mode is used for synchronization purposes when using our Programmable Scan Mode. Line Output mode creates a rising edge for each row that the rolling shutter readout mechanism of the sensor advances. For more information, see our tech note on Programmable Scan Mode.
First Row, All Rows, Rolling Shutter and Line Ouput triggering modes are summarised in Figure 3.
Any Row Mode
Any Row triggering mode is a bit different, as it forces the camera out of ‘overlap’ operation, where one frame can be exposed while the previous frame is still being exposed and read out (as seen in Fig.3). Any Rows mode is shown in Fig.4, where it creates distinct gaps between frames, not found in other trigger modes. The trigger signal is high when any row is exposing, and so is equal to the time between the start of the first and last row’s exposure. Maximum camera frame rates are not achievable with Any Row Mode, but it does avoid frame overlap if this is desired.
Our Prime and Kinetix families of CMOS cameras have four independent trigger output signals, as seen from the BNC trigger cables we use (Fig.1) which have four outputs. This enables the camera to have hardware control over light sources that cycle through different excitation wavelengths during acquisition, such as a multi-wavelength LED light source that rapidly switched between different colors to excite different fluorophores.
The enabled outputs are cycled within each frame. For example, if two outputs are enabled, the output trigger signal alternates between outputs 1 and 2. If all four outputs are enabled, the sequence would look similar to Fig.5.
Sequenced Multiple Acquisition Real-Time Streaming, or SMART Streaming, is an exclusive Teledyne Photometrics camera feature that enables our cameras to capture a continuous sequence of images while cycling through a maximum of 16 pre-programmed exposure time values. This results in very high frame rate imaging while maintaining the correct exposure level for each fluorophore.
The maximum exposure time per frame is 10 seconds, in keeping with SMART Streaming’s high frame rate benefits. By combining Multiple Output Triggers with SMART Streaming, it is possible to control the exposure time of each output independently, as seen from Fig.6. This is much faster than using software-based methods to control the timing of illumination devices
Triggering allows for finer control over camera exposure and acquisition and can pair the camera with other imaging hardware, such as light sources, for synchronization during advanced imaging experimentation. If imaging with a motorized stage, multiple light source wavelengths, samples that are highly sensitive to light, or advanced camera scanning modes such as Programmable Scan Mode, then using our external hardware triggering cables allows for modulation and more control over your imaging.