Theoretical and Practical Aspects of Routine H&E Staining (Online CE Course)

(based on 93 customer ratings)

Author: Anita Buchiane, HT(ASCP) QIHC
Reviewer: Carla Shoffeitt, MSM, HT(ASCP)

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This course will explore the history and application of hematoxylin and eosin (H&E) stains used in the clinical histology laboratory. Basic H&E staining mechanisms and the classification of biological stains will be discussed. The material presented details the chemistry, diagnostic application, and problem-solving strategies for H&E stains.

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Continuing Education Credits

P.A.C.E.® Contact Hours (acceptable for AMT, ASCP, and state recertification): 1.5 hour(s)
Approved through 8/31/2024
Approved through 8/31/2024


  • Differentiate between natural and synthetic dyes.
  • Analyze progressive staining versus regressive staining.
  • Distinguish between the oxidants and mordants in common hematoxylin and eosin (H&E) stains.
  • Identify cell constituents demonstrated with the H&E stain.
  • Describe the purpose of and reagents used for deparaffinization, hydration, and dehydration.
  • Identify potential problems encountered with routine staining and solutions to resolve them.

Customer Ratings

(based on 93 customer ratings)

Course Outline

Click on the links below to preview selected pages from this course.
  • Introduction
      • Clinical Significance and Correlation of Histology
      • How is Histology Useful to the Pathologist?
      • Application of the Hematoxylin and Eosin (H&E) Stain
      • Why is the H&E Stain the Most Common?
      • Which of the following statements about histology are TRUE? (Choose all that apply.)
  • Categories of Biological Stains
      • Three Broad Categories of Biological Stains
      • Which one of the following categories of stains is used to differentiate between a cell's nucleus and cytoplasm?
      • Which category of stain would you employ if asked to demonstrate muscle in a tissue sample?
  • Classification of Biological Stains
      • Classification of Biological Stains
      • An acidic dye has a pH less than 7 and has an affinity for __________ tissue components.
      • Origin of Dyes: Natural
      • Origin of Dyes: Synthetic
      • The first natural dyes were made from aniline and are therefore sometimes referred to as aniline dyes.
      • Examples of Synthetic Dyes
      • Chemistry of Dyes
      • Mechanisms of Action
      • What is the staining mechanism that overstains tissue and then employs a differentiation step?
      • Progressive or Regressive Hematoxylin Staining
      • Which type of staining employs the process of differentiation to delineate desired tissue elements?
  • Routine Staining in the Histology Laboratory
      • Hematoxylin Oxidation
      • Mordants
      • Commonly used Hematoxylins: Their Oxidizers and Mordants
      • Differentiating
      • The Bluing Step
      • Eosin as a Counterstain
      • Types of Eosin
      • Eosin Differentiation
      • Match the following solutions with its' appropriate use.
      • One advantage of using alcohol-based eosins over aqueous eosins is that they will stain slower and produce a wider range of shades.
      • Other than the cytoplasm of cells, what other tissue constituents are stained with eosin? (Choose all that apply.)
  • Applications of the H&E Stain
      • Healthy Versus Diseased Tissue with H&E Stain
      • Uterus
      • Liver
      • Breast
      • Skin
      • Appendix
      • Fallopian Tube
  • Additional Steps Employed in Routine H&E Staining
      • Paraffin Sections
      • Paraffin Sections, continued:
      • Staining Time Comparisons
      • General H&E Staining Procedures
      • When performing regressive H&E staining, the slides must be deparaffinized and ___________ before they will be able to take up the aqueous-based h...
  • Some Common Problems Encountered in H&E Staining
      • Nuclear Staining Errors
      • Poor Eosin Differentiation
      • Poor Contrast Between the Hematoxylin and the Eosin
      • Reddish- Brown Nuclei
      • Dark Precipitate On The Slide
      • Uneven Staining
      • White Patches on Slides After Deparaffinization Step
      • Clear Patches on Tissue After Hydrating
      • Contaminated Clearing Agent
      • Which of the following observations would indicate that tissue sections have not been sufficiently deparaffinized? (Choose all that apply.)
      • What is the function of the hydration alcohols? (Choose all that apply.)
      • What is the indication that the tissue did not go through the bluing step?
  • References
      • References

Additional Information

Level of instruction: Intermediate
Intended audience: Clinical laboratory histotechnologists and technicians,and other medical laboratory personnel who have an interest in this subject matter. This course is also appropriate for histology and clinical laboratory science students,  pathology residents, and practicing pathologists. 
Author information: Anita Buchiane, HT(ASCP) QHIC, is the Lead Histology Technician at Brattleboro Memorial Hospital in Brattleboro, Vermont. Prior to this position, she worked as a Neuropathology/Special Procedures Technician at Baystate Medical Center in Springfield, Massachusetts.  Ms. Buchiane is a graduate of Hartford Hospital School of Allied Health in Connecticut, where she earned her Certificate in Histotechnology, and Greenfield Community College, Greenfield, Massachusetts, where she earned an associates degree in Liberal Arts with a math and science concentration. She is a member of the National Society for Histotechnology and the VT/NH Society for Histotechnology.  She was the recipient of the 2007 Lee G. Luna Foreign Travel Scholarship awarded by the National Society for Histotechnology and the 2010 Region 1 Scholarship awarded by the VT/NH  Society for Histotechnology.
Reviewer information: Carla J. Shoffeitt, MSM, HT(ASCP) is the System Director of Anatomic Pathology for Emory Healthcare in Atlanta, Georgia. She holds a Master of Science Degree in Healthcare Management as well as a Bachelor of Science degree in Biology from Troy University, Troy, Alabama. She is certified as a Histotechnician and has 29 years of experience in the field of Histology.
Course Description: This course will explore the history and application of hematoxylin and eosin (H&E) stains used in the clinical histology laboratory. Basic H&E staining mechanisms and the classification of biological stains will be discussed. The material presented details the chemistry, diagnostic application, and problem solving strategies for H&E stains. 

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breast 40x_edit

Breast H&E S11-792a with arrows


Fallopian Tube H&E with arrows


liver bx 40x H&E with arrows

not blued_edit