Abstract
We have developed an imaging method designated as correlative light microscopy and block-face imaging (CoMBI), which contributes to improve the reliability of morphological analyses. This method can collect both the frozen sections and serial block-face images in a single specimen. The frozen section can be used for conventional light microscopic analysis to obtain 2-dimensional (2D) anatomical and molecular information, while serial block-face images can be used as 3-dimensional (3D) volume data for anatomical analysis. Thus, the sections maintain positional information in the specimen, and allows the correlation of 2D microscopic data and 3D volume data in a single specimen. The subjects can vary in size and type, and can cover most specimens encountered in biology. In addition, the required system for our method is characterized by cost-effectiveness. Here, we demonstrated the utility of CoMBI using specimens ranging in size from several millimeters to several centimeters, i.e., mouse embryos, human brainstem samples, and stag beetle larvae, and present successful correlation between the 2D light microscopic images and 3D volume data in a single specimen.
Highlights
Light microscopy has been used in biology to perform morphological analysis since era of van Leeuwenhoek in the 17th century[1]
This block size is sufficient to include most specimens for light microscopy encountered in the biological laboratory, as microscopy specimens are generally smaller than a glass slide with a short side length of 26 mm or a cryostat specimen holder with a diameter of 30 mm
The correlation between 2D and 3D data was achieved for specimens smaller than a millimeter by scanning electron microscopy (SEM) combined with an automatic tape-collecting ultra-microtome (ATUM)[8]
Summary
Light microscopy has been used in biology to perform morphological analysis since era of van Leeuwenhoek in the 17th century[1]. Some laboratories have developed apparatus for obtaining serial block-face images from frozen blocks or paraffin-embedded tissue, and reconstructed 3D images from various specimens, such as mouse and zebrafish embryos[3], rat knee joint[4], whole adult mouse[5, 6], and equine ovary[7]. These types of apparatus are not widely used. Successful correlations between 2D light microscopic data and 3D volume data of serial block-face images within single specimens are shown
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