Immunohistochemistry (IHC) - Detection and Identification of Infectious Disease Processes in Surgical Pathology (Online CE Course)

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Author: Jim Burchette, HT(ASCP)
Reviewer: M. Lamar Jones, BS, HT(ASCP)

This course discusses various entities that cause infectious diseases. These entities are identified in formalin-fixed, paraffin-embedded (FFPE) human tissue by use of specific primary antibodies and immunohistochemical (IHC) techniques. IHC procedures, including chromogenic substrates, enhancement techniques, and alternative methodologies are also discussed.

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Continuing Education Credits

Florida Board of Clinical Laboratory Personnel Credit Hours - Immunohistochemistry/Advanced Histologic Techniques: 2 hour(s)
Approved through 9/1/2022


  • Identify infectious entities on formalin-fixed, paraffin-embedded human tissue by use of specific primary antibodies and immunohistochemistry (IHC) techniques.
  • Describe specific entities that cause various infectious diseases.
  • Discuss both monoclonal and polyclonal antibodies that are available for use in the detection of infectious diseases.
  • Outline IHC techniques, including the use of alternative methodologies, chromogenic substrates, and enhancement techniques.

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Course Outline

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  • Introduction
      • Using Immunohistochemistry (IHC) for the Detection of Infectious Diseases
      • A virus is the ONLY infectious entity that can be detected by IHC techniques.
      • ONLY monoclonal antibodies can be used for IHC techniques for the demonstration of infectious diseases.
      • Disclaimer
      • IHC Detection Systems
      • Tissue Pretreatment: Heat-Induced Epitope Retrieval (HIER)
      • Heat-Induced Epitope Retrieval (HIER) Equipment
      • Tissue Pretreatment: Enzyme-Induced Epitope Retrieval (EIER)
      • Tissue Adhesion
      • Enzyme Labels
      • Chromogenic Substrates
      • Which of the following is an ADVANTAGE to using a polymer or micro polymer detection system?
      • ALL epitopes require some form of heat or enzyme retrieval.
      • The histologist should always ADD tissue section adhesive to the flotation bath when using positive-charged slides.
      • The horseradish peroxidase (HRP) enzyme is derived from the root of the AGAVE plant.
  • Infectious Disease Entities: Antibodies and Reagents used in Immunohistochemistry (IHC)
      • Antibodies
      • Viral Markers
    • Cytomegalovirus (CMV)
      • Cytomegalovirus (CMV)
      • Additional Cytomegalovirus (CMV) Images
      • In CMV infected cells, Feulgen-positive, granular amphophilic inclusions swell inside the nuclei giving the characteristic "owl's eye" nuclear pattern...
      • CMV infection can affect many organs, including the lungs and gastrointestinal (GI) tract.
    • Herpes Simplex Virus (HSV)
      • Herpes Simplex Virus (HSV)
      • Additional Herpes Simplex Virus (HSV) Images
      • In addition to blistering infections of the skin, mucous membranes, and esophagus, both herpes simplex and varicella zoster infections can be associat...
      • ONLY nuclear staining patterns of herpes simplex virus (HSV) can be observed.
    • Adenovirus (ADV)
      • Adenovirus (ADV)
      • Additional Adenovirus (ADV) Image
      • ADV “smudge cells” can be seen in histologic and cytologic preparations.
      • The author's preferred proteolytic enzyme used to pretreat formalin-fixed, paraffin-embedded (FFPE) tissue sections is 0.25% trypsin with pH 7.6-7.8.
    • Epstein-Barr Virus (EBV)
      • Epstein-Barr Virus (EBV)
      • Epstein-Barr Virus (EBV) Latent Membrane Protein 1 (LMP1)
      • Epstein-Barr Virus (EBV) Epstein-Barr Nuclear Antigen 2 (EBNA2)
      • Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) is an IgG1, kappa monoclonal antibody consisting of commercially individual clones CS1, CS2,...
      • Post diaminobenzidine (DAB) metal enhancement, such as 1% copper sulfate and a light hematoxylin counter stain, will improve visualization of a light ...
    • Varicella Zoster Virus (VZV)
      • Varicella Zoster Virus (VZV)
      • Once infected with chickenpox, a person can NEVER again have a varicella zoster virus (VZV) infection.
      • Complications such as pneumonia and encephalitis can develop in adults who were NOT exposed as children, but contract chickenpox later in life.
    • Human Papilloma Virus (HPV)
      • Human Papilloma Virus (HPV)
      • Human papilloma virus (HPV) is an RNA virus that causes proliferation on epidermal and mucosal surfaces.
    • Polyomavirus
      • Polyomavirus
      • JC Polyomavirus (JCV)
      • BK Polyomavirus (BKV)
      • The initials JC and BK were taken from the first patients identified with the specific polyomavirus.
      • Anti-human SV40 monoclonal antibody, clone MRQ-4, can ONLY be used for the demonstration of JC polyomavirus (JCV) in formalin-fixed, paraffin-embedded...
      • Anti-SV40 monoclonal antibody, clone MRQ-4, will usually result in a cytoplasmic localization of the polyomavirus ONLY within the renal tubules.
    • Hepatitis B Virus
      • Hepatitis B Virus (HBV)
      • Hepatitis B Core Antigen (HBcAg)
      • Hepatitis B Surface Antigen (HBsAg)
      • IHC Double Stain for Hepatitis B Surface Antigen (HBsAg) and Hepatitis B Core Antigen (HBcAg)
      • Hematoxylin and eosin (H&E) stain alone is sufficient to asses hepatitis infection.
      • Hepatitis B core antigen (HBcAg) is demonstrated in the cytoplasm of infected hepatocytes.
      • Hepatitis B surface antigen (HBsAg) is a nuclear reaction in type B viral hepatitis infected hepatocytes.
    • Parvovirus
      • Parvovirus (PARVO) B19
      • Parvovirus B19 is a very LARGE virus which causes erythema infectiosum (fifth disease) in children.
      • Parvovirus replicates in bone marrow cells and inhibits red blood cell (RBC) production (erythropoiesis) causing hemolytic complications.
      • Parvovirus B19 has been implicated in hydrops fetalis, which causes spontaneous abortion in humans.
    • Parainfluenza
    • Respiratory scintial virus (RSV)
      • Respiratory Syncytial Virus (RSV)
      • Cell cytoplasm is the target area for demonstrating RSV.
    • Human Herpes Virus 8 (HHV8)
      • Human Herpesvirus 8 (HHV8)
      • Human herpesvirus 8 (HHV8) is an RNA virus that belongs to the Alphaherpes viriniae subfamily.
  • Parasitic or Fungal Entities: Antibodies and Reagents used in Immunohistochemistry (IHC)
      • Pneumocystis Pneumonia (PCP) or Pneumocystosis
      • Toxoplasmosis
      • Additional Images of Toxoplasmosis
      • Toxoplasma will infect cells, where the parasite will remain until the cell dies. When the cell dies, the parasite will infect another cell.
      • What antibody isotype is P. jiroveci mouse monoclonal clone 3F6?
  • Bacterial Entities: Antibodies and Reagents used in IHC
  • References
      • References

Additional Information

Level of instruction: Intermediate to advanced
Intended audience:  This program is designed for the clinical immunohistochemist that has a working knowledge of immunohistochemistry (IHC) technique and can also be used as a reference for proper staining patterns and morphology. This course is also appropriate as an educational and training tool for histology laboratory personnel. 
Author information: Jim Burchette, HT(ASCP) recently retired after 34 years at Duke University Medical Center, Durham, North Carolina. As an Immunopathology Analytical Specialist in Molecular Pathology, Jim's primary job functions included daily IHC quality control and managing the daily technical aspect of the Clinical Immunopathology operation. He has over 38 years of experience in surgical pathology histotechnology with 33 years of immunopathology specialty. Jim has an extensive experience validating new antibodies for research and diagnostic immunohistochemical use. To date, Jim has 80 publications (70 of which are peer-reviewed) and two book chapters, as well as many abstracts, posters, and technical acknowledgments in peer reviewed scientific articles. He has also served as a consultant for Triangle Biomedical Sciences (TBS), Dako North America, Leica Microsystems, and Polysciences.
Reviewer information:  Lamar Jones, BS, HT(ASCP) holds a BS degree from the University of Kentucky, Lexington, Kentucky. He received histotechnology training from the School of Histotechnology at the University of Tennessee Medical Units, Memphis, Tennessee. Lamar is currently the Technical Coordinator in the Department of Pathology at Emory University Hospital in Atlanta, Georgia. He also serves as the HT Program Director for the School of Histotechnology at Davidson County Community College, Mocksville, North Carolina and serves as lecturer, author, workshop director, and presenter for local, state, and national histotechnology meetings. 
Course description:  This course discusses various entities that cause infectious diseases. These entities are identified in formalin-fixed, paraffin-embedded (FFPE) human tissue by use of specific primary antibodies and immunohistochemical (IHC) techniques. IHC procedures, including chromogenic substrates, enhancement techniques, and alternative methodologies are also discussed.

AD09-173 Bone Marrow Toxoplasma g.
Case of disseminated Toxoplasmosis infection in bone marrow from a heart transplant patient demonstrated with Toxoplasma gondii mouse monoclonal antibody, clone TP3. DAB chromogen.

EBV positive high grade malignant lymphoma immunoblastic plasmacytoid type. Nuclear reaction shown with EBV EBNA2 antibody. DAB chromogen.

Helicobacter pylori bacteria demonstrated in gastrointestinal tissue using rabbit polyclonal antibody HRP labeled detection system with DAB chromogen.

Toxoplasmosis infection in lung demonstrated with Toxoplasma gondii mouse monoclonal antibody, clone TP3. Fast red chromogen. Notice the clusters of T gondii protozoa and individual protoza. Tissue cysts will rupture, releasing individual protoza to form new cysts.

Virus-like particles in the nuclei of abnormal oligodendrocytes are labeling with mouse monoclonal SV40 antibody, clone MRQ-4. DAB chromogen.

CMV image A

Treponema pallidum (spirochetes) demonstrated in a cutaneous biopsy from an HIV patient with epidermal ulcerations and dermal lymphohistiocytic inflammation. Fast red chromogen.

Pretreatment (PT) module