Here you will find software downloads and product support materials for our cellular imaging and analysis portfolio, including high content imaging, high content analysis and microscopy.
Limitations of Hamamatsu ImagEM "Real-time Image Processing" Features
The Hamamatsu ImagEM cameras (C9100-13 and C9100-14) include real-time image processing features. This technical note discusses the limitations of some of these features when used for fluorescence imaging.
Background Subtraction and Shading Correction
The "real time background subtraction" and "shading correction" tools on the ImagEM work by processing a single frame. These tools are safe to use for any type of acquisition as they do not depend on successive frames being related.
Spot Noise Reducer
The "spot noise reducer" tool works by comparing each frame to previous ones and eliminating small bright signals that only appear in a single frame. This tool does not work well where successive frames bear little or no relation to each other - it is very likely that spurious events will either not be rejected, or that small bright features will be incorrectly eliminated.
Unfortunately, many acquisition protocols involve moving or reconfiguring hardware between frames and the "spot noise reducer" is therefore not suitable for use with these protocols. In practice this means that this feature can only be used for single channel, single Z protocols where the camera is running at maximum speed.
The "recursive filter" tool reduces image noise by calculating a weighted rolling average of each frame against previous frames. Like "spot noise reduction", this tool does not work well where successive frames bear little or no relation to each other. Depending on the exact protocol, this tool may create artefacts including:
- Digital "bleed-through" between channels
- Blurring of Z stacks
- Reduction of intensity where images captured with the shutter closed are averaged against images captured with the shutter opened
Like "spot noise reduction", the "recursive filter" tool should only be used for single channel, single Z protocols where the camera is running at maximum speed.
High Content Analysis
Using Openlab Layer User Data To Set Properties Within Volocity
The "Set Volocity Data" task can be used to add data to an Openlab layer which will be read as a property in Volocity.
Using the Auto Contrast Function with Volocity
If you drag a stack of grayscale images into a Volocity library, it is possible to adopt and then 'Auto Contrast' them all. However, when a volume is made from these images, the resultant XYZ view looks corrupted, with regions in the image that are expected to be dark appearing bright and vice versa. This happens because the Auto Contrast function modifies the LUT (look up table) of an image and maximises the contrast based on the data in that particular image. Therefore, if Auto Contrast is applied to an image stack, each image is likely to have a different LUT applied.
A volume made from images enhanced in this way would be expected to show enhancement in the XYZ or 3D view. However, when a volume is created in Volocity, any LUT applied to the first image in the stack is applied as the LUT for the whole volume. It is likely that the LUT applied to the first layer in the stack will not be appropriate for the other layers in the stack. The volume thus created will not therefore look as you would have expected from viewing the individual enhanced layers. However, you can correct the problem by applying Auto Contrast again, this time to the volume. During Auto Contrast of a volume, the data of the whole volume is taken in to account to generate the appropriate LUT.
Problems can be avioded altogether by only applying Auto Contrast to the volume and not to the individual images which make up the volume.
Volocity 3D Rendering - How To Export a 2D View
Position the volume in the 3D rendering view to achieve the view required.
Select "Capture Snapshot" from the View menu. This will add a single 2D image to the library which represents the view at that moment. That image can then be exported from the library in any of the supported formats.
Capture Snapshot also operates on the Image view, adding a folder containing 4 images to the library.
Volocity Alignment Tool
The Volocity Alignment Tool is designed to compensate for rotational and translational shifts between the two-dimensional planes that make up a volume.
When performing rotational correction the alignment module will search for correlation +/- 30 degrees from the origin.
The Alignment Tool searches for correlation between slices in the data to match features and establish where each slice should be in relation to the next. For alignment to be successful features must be represented in neighboring slices ie the slices need to be similar.
The Alignment Tool will not be able to perform on structures which are, or approximate to, spheres since they show rotational symmetry which makes feature matching impossible.
During rotation of images anti-aliasing is applied which has a smoothing effect on the data.
Alignment operates on individual volumes, and volume sequences, not 2 dimensional data. Select volume or volumes, or channels within a volume sequence, to be aligned and select "Align Volume" from the "Tools" menu.
The original data is not overwritten, a new image or image sequence is created with the suffix (aligned).
There is no benefit to be gain by repeatedly aligning the same data.
The XY dimensions of an aligned volume will be the same as the original volume, areas of image that have been shifted outside the original boundary to allow alignment will be cropped from the result.
Volocity for Windows: Dongle Installation
It is possible for Windows computers to fail to recognise the Volocity dongle if the dongle is plugged in before Volocity is installed. In this circumstance:
Finish the Volocity installation. The steps outlined below will not work if the installation program has not been run at all.
For Windows XP and 2000:
Right click "My Computer".
Chose "Properties" from the menu.
Click "Hardware" tab.
Click "Device Manager" button.
The Device Manager shows a list of installed hardware.
Look for an icon with a yellow exclamation mark next to it called "Unknown USB Device" or similar. This will most likely be under "Universal Serial Bus Controllers"
Click this icon to select the device in the list, then from the toolbar at the top of the window click the 'Uninstall' button to remove it. OK the dialog.
Click the Computer icon at the top of the hardware list. Again from the toolbar at the top of the window click the "Scan for hardware changes" button.
Windows should then redetect the dongle and install the correct drivers.
Volocity LE 2.5 and Higher
The feature content of Volocity has increased with Volocity 2.5. In order to continue to offer Volocity LE free of charge changes to the features available have had to be made. Some Volocity LE users may prefer not to upgrade to version 2.5 to continue using features available only in version 2.0.1 or below.
The following are no longer available in Volocity LE:
- Export as QuickTime movie.
- Export as AVI movie (Windows only).
- Image enhancement using the "Levels" tool.
The following features new to Volocity 2.5 are not available in Volocity LE:
- Image View - Extended Focus.
- Image Sequence View - Timeline.