Variation in the intensity of the hematoxylin and/or eosin in different areas of the same slide (as seen in the image on the right) could be caused by a variety of reasons:
- The tissue varies in thickness. When cutting the tissue the sections should be of uniform thickness.
- Solutions may be too low. The volume of all the reagents must completely cover the tissue to ensure that the entire section is exposed to all of the reagents. This includes the water rinses.
- The duration of the water rinses following the differentiating and bluing steps must be adequate to halt the reactions and to avoid carry over from one reagent to the next.