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Obtaining clear images from a Z-stack has been a challenge for microscopists. The SharpStack add-on module for Image-Pro Plus and Image-Pro Discovery meets the challenge by extracting clear, sharp images from a stack of hazy planes.

Unlike other deconvolution solutions, SharpStack integrates seamlessly with image capture, 2D processing, analysis and reporting within the world's most popular image analysis software, Image-Pro Plus, as well as with the new Image-Pro Discovery.

With SharpStack, nearest neighbor, no neighbor, and inverse algorithm functions are employed to easily sharpen one or all planes from a Z-stack.

Animation of raw and deconvolved Z-stack images of a pollen grain. On the right, pollen has been deconvolved using SharpStack's nearest neighbor algorithm. (Animation features are part of Image-Pro.)

The Inverse Filter is a one-step non-iterative approach based upon inverse filtering theory. The deterministic blurring as a convolution of the image with the point spread function can be modeled. In the frequency domain a convolution transforms into a multiplication of the Fourier transform of the sample with the optical transfer function. The optical transfer function (OTF) is the Fourier transform of the point spread function. The inverse filter then accomplishes image restoration by dividing the Fourier transform of the image by the OTF.

The Nearest Neighbor algorithm works by deconvolving one image slice at a time. It uses information from image slices that reside above and below the image slice that is being processed. The precise increment slice position from the measured slice is user selectable. If the slices are chosen judiciously, this approximation will produce results very close to the inverse filter method but takes much less time.

The No Neighbor method uses the information from each single slice to construct a 2D PSF. This is the fastest but may not be as representative of the sample as the other methods.

Inverse filter, nearest neighbor, and no neighbor algorithm functions are all included in SharpStack.