CAP Pro Course - Histology - Immunohistochemistry Part 1

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Author: Carla Shoffeitt, MSM, HT(ASCP)
Reviewer: Alexandru Casapu, MBA, MLS(ASCP)CM, PBTCM

Continuing Education Credits

Objectives

  • Discuss the basic principles of immunohistochemistry (IHC).
  • Review fixation and processing requirements related to IHC.
  • Identify and discuss methods of epitope retrieval.
  • Discuss methods and detection systems used in the visualization of antibody-antigen reactivity.
  • Outline basic IHC staining protocol and staining considerations.
  • Discuss quality control (QC) measures used in IHC, including actions taken when QC results are not acceptable.

Course Outline

  • Discuss the basic principles of immunohistochemistry (IHC).
      • Defining Immunohistochemistry (IHC)
      • Antibodies, Antigens, and Epitopes
      • Antibody-Antigen Binding
      • Monoclonal Versus Polyclonal Antibodies
      • Antibody Dilution and Titration
      • Immunohistochemistry (IHC) is a method of localizing and demonstrating specific antigens in tissues or cells based on antigen-antibody interactions. W...
      • Antibodies used in IHC staining exist as monoclonal and polyclonal antibodies. Which of the following statements MOST accurately describes the differe...
  • Review fixation and processing requirements related to IHC.
      • Fixative Considerations Based on Specimen Type
      • Formalin Fixation
      • Fixation and Processing Considerations for Breast Markers
      • Microtomy and Slide Preparation
      • Prior to performing IHC staining, blood smears, touch-preps, and cytocentrifuged cytology preparations are air-dried to allow cells to fix to the slid...
      • Which of the following statements is TRUE in regard to fixation and processing considerations for breast markers?
      • You have been assigned blocks to section for IHC staining. Which of the following should be considered when sectioning slides for these requests?
  • Identify and discuss methods of epitope retrieval.
      • Unmasking of Epitopes
      • Proteolytic-Induced Epitope Retrieval (PIER)
      • Heat-Induced Epitope Retrieval (HIER)
      • When formalin-based fixatives are utilized, cross-linkages are formed with structural proteins. As a result, antigens can become "masked." Which of th...
      • Heat-induced epitope retrieval (HIER) methods have greatly improved the quality and reproducibility of IHC staining. Which of the following represents...
  • Discuss methods and detection systems used in the visualization of antibody-antigen reactivity.
      • Direct and Indirect Methods of Immunohistochemical (IHC) Staining
      • Peroxidase Anti-Peroxidase (PAP) Method
      • Avidin-Biotin Complex (ABC) and Labeled Streptavidin-Biotin (LSAB) Methods
      • Polymer-Based Immunohistochemistry (IHC)
      • The avidin-biotin complex (ABC) method of IHC staining makes use of the essentially irreversible binding seen between avidin (an egg white protein) an...
      • In polymer-based IHC staining, what component allows for additional binding sites and ultra-sensitivity?
      • Dr. Jones has reviewed an IHC stain on a liver biopsy. He is concerned because there appears to be unexpected positive staining. He has come to you fo...
  • Outline basic IHC staining protocol and staining considerations.
      • Blocking Non-Specific Binding
      • Importance of Washing Steps
      • Incubation of Primary Antibody and Detection System Reagents
      • Chromogen and Substrates
      • Endogenous biotin is bound to enzymes and protein in a variety of tissues and is found in high concentration in brain, kidney, and liver tissue. How i...
      • Which of the following statements with regard to the function of wash buffers is TRUE?
  • Discuss quality control (QC) measures used in IHC, including actions taken when QC results are not acceptable.
      • Controls in Immunohistochemistry (IHC)
      • Validation of New Primary Antibodies
      • Validating New Lots of Antibody and Ancillary Reagents
      • Storage Considerations for Immunohistochemistry (IHC) Reagents
      • Documenting Instrument Function and Maintenance
      • Participation in Quality Assessment (QA) Programs
      • Tate is preparing an early morning staining run on the automated IHC stainer. Upon retrieval of the detection kit and needed antibodies from the refri...
      • You have received an antibody that is new to your laboratory. The antibody requires optimization and validation before being used for patient testing....
      • John receives a new lot of GATA-3 antibody. What important packaging content should John review prior to initiating validation on the new lot?
  • References
      • References

Additional Information

Level of Instruction: Intermediate
Intended Audience: This program is designed as an educational and training tool for HT, HTL, MLS, MT, MLT personnel, medical laboratory science students and interns, pathology residents, and practicing pathologists.
Author 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.
Reviewer Information: Alexandru Casapu, MBA, MLS(ASCP)CM, PBTCM, has over 20 years of experience as a medical laboratory scientist, section supervisor, and laboratory manager. He is the former Director of Clinical Laboratory Technology Program at Georgia Piedmont Technical College. He is currently a Program Director at MediaLab, Inc. Alex holds BS degrees in Biology and Medical Technology from Clark Atlanta University and an MBA from the University of Georgia.

This course is part of the CAP Competency Assessment Hub.

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