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The page below is a sample from the LabCE course Troubleshooting Guidance for Hematoxylin and Eosin (H&E) Stain. Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

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Adjusting for Coloration

Of course, getting everyone to agree on what is the best stain can be a challenge. It may be easiest to program whatever staining is being used to accommodate several protocols with slightly different times, so that the pathologists have some options. I also create a simple code system so that I know what the stain times are, but the pathologists will have to respond based only on what they see.
When adjusting coloration, it is best to keep it simple. The following adjustments often work well:
  • Hematoxylin +/- 30 seconds
  • Eosin Y +/- 15 seconds
Typically, it is helpful to only make one adjustment at a time. These times are just enough to make subtle changes. You can be more aggressive if needed. If the hematoxylin is way too light, then certainly increase the time by one minute or more. For tweaking, however, the times above tend to work well.
REMEMBER: Sometimes simply adjusting either the hematoxylin or eosin may be enough to make the other brighter or lighter, depending on what you are looking for.