If the hematoxylin or the eosin is too dark or too pale, the contrast between the nuclear staining and the cytoplasmic staining will be poor. Decreasing the time in the hematoxylin and/or eosin solution will decrease the intensity of the stain. Conversely, increasing the time will increase the intensity.
The time spent on differentiating solutions can also be adjusted. Increasing the time in the acid alcohol or the dehydrating alcohols will decrease the intensity of the hematoxylin and eosin, respectively. Decreasing the time spent on these will increase the hematoxylin's and eosin's intensity.
Considering these factors, the optimal times for the dyes and the differentiators must be determined to properly balance the nuclear and cytoplasmic staining.
To ensure good results, it is also essential to keep the reagents fresh by exchanging them with new reagents at regular intervals.
The top image shows a poorly stained slide with little contrast between the hematoxylin and the eosin. The bottom image shows a well-stained slide.
