Rapid tissue histology using multichannel confocal fluorescence microscopy with focus tracking

In histology, hematoxylin and eosin (H&E) is widely used as a gold standard because it can provide precise morphological information on various tissue types (1). Hematoxylin stains nuclei blue or purple, and eosin stains cytoplasm and extracellular matrix pink (2). Since the tissue being examined is made into a slide to be viewed with a bright-field microscope, H&E histology generally requires physically sectioning tissue to around 5 to 10 μm thick. Paraffin sectioning is routinely used and regarded as the gold standard for H&E histology. Although paraffin sectioning provides clear visualization of tumor tissue, it requires up to days to fix and embed the tissue in paraffin (1). Instead of embedding the tissue, freezing before sectioning can be used to acquire rapid results (3) known as frozen histology. This cryo-sectioning allows for histology results in 30 to 40 minutes compared to days required for paraffin sectioning (4-7)