Hematoxylin: 30 seconds–10 minutes; depends on concentration and pathologist preference
Tech Tip: The main difference between the 2 most common types of hematoxylin is concentration: Mayer’s = 1gm/L and Gill’s = 2gm/L.
If the hematoxylin incubation time is too long or too short, then the following may occur: Dissatisfied pathologist difficulty distinguishing cellular morphology.
If the hematoxylin is exposed to light, then the following may occur: Dissatisfied pathologist oxidation creating crystalline precipitates that are observed under the microscope; these crystalline structures can be distracting and, in some instances, make accurate diagnosis difficult.
Dehydration and Mounting:
Dehydration: graded alcohol (3–10 minutes) and xylene or xylene substitute (5–15 minutes)
Mounting Media: non-aqueous (permanent chromogens) vs. aqueous (non-permanent chromogens)
Tech Tip: Non-permanent chromogens like AEC are soluble in alcohols. It is important to air-dry these slides and use an aqueous mounting media.
No or inadequate dehydration can result in: Cloudy appearance water is not fully removed creating a hazy appearance to the tissue; cellular morphology can also be affected.
Use of the wrong dehydration method or inappropriate mounting media can result in: Weak staining non-permanent chromogens can fade if used with alcohols or with non-aqueous mounting mediums.