We developed a new computerized CCD video light microscopy system for obtaining three-dimensional (3-D) RGB measurements of shape (with color texture) and growth of intact plants under various conditions and over a wide magnification range. We used a modified shape-from-focus algorithm that incorporated a linear regression operator for 3-D reconstruction of images of petunia seedlings, which have a coarse texture. This algorithm was unsuitable for processing images of specimens with glossy texture (e.g., plant cells), but this drawback was mitigated by illuminating the cells in a checked pattern.The algorithm also reduced incorrect range estimates, which are due to defocused areas around the object's edge in original images. Within only a few minutes, the presented system can yield a series of original color images and reconstruct 3-D images incorporating color texture. All operations of our new system are carried out automatically, and the results are displayed by using in-house developed software that allows observation of the reconstructed 3-D image from any direction on the upper side. At angles from 10 to 80* to the objective's face, the surface area can be estimated to within an error of 5%.
The shape-from-focus algorithm, a passive method, was modified for microscopic measurement of 3 D shape and texture mapping of intact petunia seedlings with coarse surface.First, both max-min and least-squares (linear regression) operators newly developed to estimate the focus measure index were compared with the sum-modifiedLaplacian operator proposed by Nayar and Nakagawa.In the new operators a deformed mask was used for shortening of calculation time. Also, RGB full color images as well as gray image were used as a series of original 2-D images measured at consecutive focused planes by a computerized light microscope. As a result, the focused range image, i.e. 3-D shape estimated using the least-squares operator and RGB images was the best of all. This combination was effective for estimating the focused range in the case of not only the coarse texture but also the texture with large gradient in gray level such as the edge. Secondly, a focused color image was interpolated from a series of original images using the range image. Also, wire frame and texture mapping images were made using the range and focused color images.As a result, it was possible to make a clear color image of the seedling.Furthermore, it made easy to observe 3 D color images of the seedling from any direction by operating a mouse.
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