Different methods are investigated in selecting and generating the appropriate microscope images for analysis of three-dimensional objects in quantitative microscopy. Traditionally, the ‘best’ focused image from a set is used for quantitative analysis. Such an objectively determined image is optimal for the extraction of some features, but may not be the best image for the extraction of all features. Various methods using multiple images are here developed to obtain a tighter distribution for all features.
Three different approaches for analysis of images of stained cervical cells were analyzed. In the first approach, features are extracted from each image in the set. The feature values are then averaged to give the final result. In the second approach, a set of varying focused images are reconstructed to obtain a set of in-focus images. Features are then extracted from this set and averaged. In the third approach, a set of images in the three-dimensional scene is compressed into a single two-dimensional image. Four different compression methods are used. Features are then extracted from the resulting two-dimensional image. The third approach is employed on both the raw and transformed images.
Each approach has its advantages and disadvantages. The first approach is fast and produces reasonable results. The second approach is more computationally expensive but produces the best results. The last approach overcomes the memory storage problem of the first two approaches since the set of images is compressed into one. The method of compression using the highest gradient pixel produces better results overall than other data reduction techniques and produces results comparable to the first approach.
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