Correct Image options are used to correct systematic errors that occur during data collection. The three basic errors are caused by photobleaching, inconsistent illumination intensity, and CCD defects.
By default, softWoRx automatically applies these options to images during the deconvolution process (see More Deconvolution Options).
If you are analyzing raw images (images that are not deconvolved), use the Correct options tool to apply these options to the images before you process them. Applying correction options is especially important when you are performing quantitative analysis on images that contain multiple Z sections or Time series.
Open Correct Image by choosing Process | Correct from the softWoRx main menu.
Use |
To... |
Normalize Intensity |
Control whether to use intensity normalization for the image correction. The default is on. |
Use Photosensor |
Control whether to use photosensor readings for intensity normalization. The default is on. If this button is off and the normalize correction button is on, the image correction program normalizes intensities based on a best-fit polynomial, rather than photosensor readings. The best-fit polynomial method is a reliable alternative to the photosensor method.
Note If an image contains invalid photosensor readings, the image correction program automatically switches to the polynomial method. In this case, the switch is recorded in the corresponding "log" file. |
Correct Bleaching |
Correct photobleaching. For situations where image intensity changes are due to the specimen and not photobleaching, turn this option off. The default is on. |
Replace Z-Lines |
Use the Z-Line correction. This is designed to detect CCD defects by searching for unusual intensity profiles along the Z-axis. For example, a "dead" CCD element appears as a straight, dark line along the Z-axis, and a "hot" CCD element appears as a straight, bright line along the Z-axis. Voxels identified as Z-line errors are replaced with the average of their neighbors. The default is on. |
Smooth Z-Lines |
Smooth the stack along the Z axis. The default is on. |
Camera Intensity Offset |
Specify a Camera Intensity Offset parameter. Conventional CCD cameras will exhibit an offset in the approximate range of 50-100 counts, whereas EMCCD cameras may be in the many hundreds. This value can be measured by acquiring a bias image at the target camera gain and noting the image mean value.
Note The bias image will change with the particular experiment variables, such as binning, exposure, and camera gain. Therefore, it is essential that you measure the Camera Intensity Offset under the same conditions in which you are imaging. |
Pass Waves Unprocessed
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Process certain channels while passing others to the output file untouched. This may be useful when an image is collected with both bright-field and fluorescence channels. Select the toggle buttons for any waves you would like to pass unprocessed. |
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You can specify options for running the Correct Image process. The following options are available.
Use |
To... |
Log File |
Specify the name of the file that contains the informational record of the Correct Image process. |
Command File |
Specify the name of the command shell script that runs the process. |
Run Options |
Specify when the job runs. There are three options: Run Now (runs immediately after choosing the Do It button, Run Later (creates a command shell script to be manually executed at a later time, or Add To Queue (puts the job at the end of a list of commands to be executed in order by the Queue Manager). |
Show Output Log |
Show the progress of the process when the check box is activated. |
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