Fixing and Staining specimens
By microscopy the specimen to be viewed is first fixed with formaldehyde, or alcohol embedded in wax and before being mounted on a glass slide cut into thin sections with a microtome and viewed under the microscope. Subcellular organelles cannot be distinguished readily under the light microscope without first staining the specimen with a chemical, like eosin or hematoxylin. In a specimen the location of an enzyme can be revealed by cytochemical staining by using a substrate which is converted into a colored product by the enzyme.
In standard light microscopy the specimen to be checked is first fixed having a solution containing formaldehydeor alcohol usually. These compounds denature proteins and, introduce covalent cross-links between amino groups on adjacent molecules , in the case of formaldehyde which stabilize protein-nucleic and protein-protein acid interactions. Then The fixed specimen can be embedded in paraffin wax or a resin and cut into thin sections (0.5-10 μm thick) by using a microtome. On a glass slide each section is mounted and then positioned on the movable specimen stage of the microscope. The sevral subcellular constituents (mitochondria, nucleus, cytosol, etc.) absorb regarding the same degree of visible light, making it hard to distinguish them under the light micro- scope without first staining the specimen. Various chemical stains bind to biological molecules; for instance, hematoxylin binds to the basic amino acids lysine and arginine in proteins, and eosin binds to acidic molecules (like DNA and the side-chains of the amino acids and glutamate and aspartate). Another way of visualizing particular structures within cells is cytochemical staining in which an enzyme catalyzes the production of various molecules of a localized, colored reaction product from a colourless precursor. In the light microscope the colored product may then be seen where the enzyme is present. For instance, peroxisomes may be visualized by using a cytochemical stain for catalase.