Sharpen

Delineate

This method enhances the edges of individual regions in an image. It corrects the halo effect and only affects edges.

Parameter	Description
Threshold	Enter the threshold value for edge detection using the slider or spin box/input field. The threshold value should correspond roughly to the gray value difference between objects and the background.
Size	Enter the size of the edge detection filter using the slider or spin box/input field. The value should correspond to the size of the transition area between objects and the background.

Enhance Contour

This method allows you to enhance contours in an image and emphasize regions in which gray values change. The function is suitable for visually emphasizing fine structures in an image.

Parameter		Description
Strength		Adjusts the factor for increasing edge enhancement.
Normalization		Defines how out-of-range pixel values are mapped. The calculated pixel values of the output image may be out-of-range and are mapped into the available range.
-	Clip	Values exceeding the pixel value range are set to the highest value available (white), values falling short of the pixel value range are set to the lowest value available (black). The effect corresponds to underexposure or overexposure. This means that in some cases information is lost.
-	Automatic	Normalizes the pixel values automatically to the available pixel value range. The highest resulting value is mapped to the maximum pixel value, the lowest resulting value to 0. As a result, the whole range of resulting pixel values is compressed evenly.
-	Wrap	If a resulting value is larger than the maximum pixel value of the image, the difference exceeding the maximum pixel value is added to 0. Similarly, if a resulting value is below 0, the resulting pixel value is the maximum pixel value minus the difference falling below 0.
-	Shift	Normalizes the output to the value "pixel value + maximum pixel value/2". As a result, all resulting values are mapped to the available value range. The middle value of the pixel value range remains constant. Values left and right of the middle value are changed progressively, so that values inside the pixel value range are changed only slightly. Values outside the pixel value range are changed strongly and mapped to the fringes of the pixel value range.
-	Absolute	Converts negative pixel values into positive values. Positive pixel values exceeding the maximum pixel value are set to the maximum pixel value.

Extended Depth of Focus

With this method, you can combine the sharp regions from the individual sections of a z-stack image to create a single sharp image. This enables you to display a considerably larger depth of field than is possible on a microscope.

Parameter		Description
Method
-	Wavelets	Uses a wavelet transformation to detect the sharpest areas in the images.
-	Contrast	For this method, the value is the difference between the brightest and the darkest pixel value within the “Kernel”.
-	Maximum Projection	Images with the brightest and darkest pixels are generated first. Of these images the image with the higher variance is used as the resulting image. Usually, best suited for fluorescence images.
-	Variance	Calculates the variance of the pixel values within the “Kernel”. Usually, best suited for bright field images.
-	Fast EDF	Uses a method for fast EDF to detect the sharpest areas in the images.
Z-Stack Alignment		This parameter should not be used for processing whole slide image scans. It is only required when processing images stemming from a stereo microscope. Not visible if you have selected Maximum Projection as Method. Selects if the z-stack image should be aligned before the calculation and with what quality level. If you acquire images with a stereo microscope, the images are displaced against each other. This displacement can be corrected. The higher the selected quality of alignment, the longer takes the calculation. Select No Alignment, if you acquire images with a compound microscope.
-	No Alignment	The z-stack image is not aligned before the calculation. Select this setting if the z-stack image has not been acquired using a stereo microscope.
-	Normal	Uses an alignment with high speed for normal image quality.
-	High	Uses an alignment with lower speed for high image quality.
-	Highest	Uses an alignment with the lowest speed with best image quality.
Process Tiles Separately		Only visible if you have selected Maximum Projection as Method. Activated: Processes each tile region separately.
Contrast Lengthscale		Only visible if Contrast or Variance is selected as Method. Sets the length scale (pixel units) at which the contrast for every slice is calculated. For images with small and sharp structures, a small contrast length scale provides good results. For images with larger structures with smoother edges, a larger contrast length scale is required.
Smoothing		Only visible if Contrast, Variance, or Fast EDF is selected as Method. Sets a value for smoothing. This parameter helps you to bridge smaller regions with missing contrast information, i.e. image regions with no structures.
Reconstruction		Only visible if Contrast or Variance is selected as Method. Defines the amount of image area with little contrast that needs to be reconstructed by an algorithm. If an image contains larger regions with little or no contrast information, the algorithm has to interpolate image information at a larger extent.
Decay Flicker Correction		Only visible if Contrast or Variance is selected as Method. Activated: Equalizes the brightness and intensity of pixels over the entire z-stack. During the acquisition of fluorescence images, bleaching can lead to a difference in pixel brightness, also called decay. When acquiring an image with a lamp, flickering can lead to fluctuations in pixel intensity. This option tries to correct both effects through the equalization over the entire stack.
Pixelwise Iteration		Only visible if Fast EDF is selected. Larger values lead to a smoother heightmap. Use the slider to adjust the desired value. Default values (depending on the selected preset): Preset = Default: 2 Preset = Small Structures: 2 Preset = Medium Structures: 3 Preset = Large Structures: 3
Depthwise Kernel Size		Only visible if Fast EDF is selected. Larger values lead to a more robust heightmap/image, especially for areas with low contrast. Should only be used by small depth of focus step sizes. Default values (depending on the selected preset): Preset = Default: 1 Preset = Small Structures: 0 Preset = Medium Structures: 2 Preset = Large Structures: 2
Resize Ratio		Only visible if Fast EDF is selected. Sets the scaling factor. Typically, smaller values are faster. Default Value: 0.25 0.25 0.5 1.0

Unsharp Mask

By using this method, you can increase the impression of sharpness in an image and consequently obtain an image which shows structures with more contrast. The function enhances contrast especially for smaller structures.

Parameter		Description
Strength		Enter the strength of the Unsharp Masking using the slider or spin box/input field. The higher the value selected, the greater the extent to which small structures are enhanced.
Sigma		Adjusts the sigma value derived from Gauss filtering. Reduces noise in an image. Each pixel is replaced by a weighted average of its neighbors. The neighboring pixels are weighted in accordance with a two-dimensional Gauss bell curve.
Color Mode		Select the desired color mode from the drop-down list.
-	RGB	Calculates the sharpness for each color channel individually. The color saturation and the color of structures may be changed, and color noise may occur.
-	Luminance	Only calculates the sharpness on the basis of the brightness signal detected. This mode does not show any color noise and changes the color saturation accordingly.
Threshold Mode		Here you can select a setting from the drop-down list for calculating the boundary between the sharpened image regions. It is only effective if the value for the Threshold Low parameter is not equal to 0 or the value for the Threshold High parameter is not equal to 100.
-	None	No adjustment takes place.
-	Binary	The transition follows the threshold values.
-	Linear	Calculates a linear course.
Threshold Low		Enter the lower threshold value using the slider or spin box/input field. This determines the lower limit from which existing contrast structures are changed.
Threshold High		Enter the upper threshold value using the slider or spin box/input field. This prevents structures in the image already featuring high contrast from being further enhanced unnecessarily.
Clip to Valid Bits		Activated: The value range of the gray/color values of the output image is adjusted to the value range of the input image.