The correct order is dehydration, embed in wax, cut into sections, staining
Explanation:
There are certain proceedings needed to obtain stained sections of vegetable or animal tissues for their microscope observations.
These steps are:
Obtention of the material: The tissue is cut to an adequate size.
Fixation: When tissues are extracted from the organism, they suffer autolysis and putrefaction, so they need to be fixated in order to keep their cells in the best state possible. Fixation involves preserving the original morphological and molecular characteristics of the tissue. Fixation avoids autolysis, putrefaction, distortion, and retraction of cells and the tissue that could affect its volume or morphology.
Dehydration. Once fixated, the fixator must be eliminated and the tissue is dehydrated by using a gradual series of solutions with alcohol in ascendant concentrations. Dehydration must be gradual to avoid tissue deformation.
Inclusion. To obtain thin cuts that can be observed under the optic microscope, the tissues must be included in a consistent, firm substance, that might be either hydrophilic or hydrophobic. A hydrophobic medium is paraffin wax, that provides hardness and plasticity.
Cut. The tissue included in wax must be cut in slides or sections thin enough to allow the diffusion and penetration of light. A microtome is used to perform these cuts. When using paraffine for tissue inclusion, the cuts are about 5 to 20 micrometers of thickness.
Stain. Once the cuts are performed, paraffin wax must be eliminated. This can be done by using an organic solvent. Then the tissue must be stained. Hematoxylin and Eosin are the most common dyes. Animal tissues in general do not have any natural color, so they need to be stained to be observed.
