Immunopathology

Immunopathology

The Immunopathology Unit, located on the 5th floor of the Warren building, is composed of the following diagnostic laboratories:

The diagnostic testing includes immunohistochemical, serological studies, flow cytometric analysis and electron microscopy.

Faculty

• Frederic I. Preffer, PhD
• A. Bernard Collins
• Atul K. Bhan, MD
• G. Petur Nielsen, MD
• John L. Niles, MD

In addition, the following faculty members closely interact with the Unit:

• Robert B. Colvin, MD
• R. Neal Smith, MD
• James R. Stone, MD, PhD

Immunohistochemistry and Serological Laboratories

The Immunopathology Unit performs all the immunohistochemical studies (immunofluorescence and immunoperoxidase) for surgical pathology, cytopathology and autopsy services. This includes studies performed on renal, skin, heart, and liver biopsies, allografts (kidney, liver and heart), tumor markers and hormone receptors (estrogen and progesterone receptors) and prognostic markers (HER-1/c-erbB-2). In addition, serological assays for the detection of antineutrophil cytoplasmic antibodies (ANCA) and anti-glomerular basement membrane (anti-GBM) antibodies, which were developed in the laboratory, are performed and available for reference laboratory testing. Immunopathology is also a reference laboratory for C4d staining by both indirect immunofluorescence and by immunohistochemistry for renal and cardiac transplants.

The immunoperoxidase laboratory has taken a leading role in establishing strategies for the use of differentiation antigens in tumor diagnosis and classification and has provided a stimulating influence on the staff members involved in diagnostic pathology. This has led to a series of investigative studies and publications in surgical pathology, including hematopoietic, soft tissue and epithelial tumors. There has been a substantial increase in the number of immunoperoxidase tests performed in the last several years (approximately 40,000 diagnostic slides in 2009). This reflects an increase in surgical pathology cases as well as addition of many new immunoperoxidase stains for the detection of tumor markers. The number of tests performed for the detection of hormone receptors (estrogen and progesterone) and for HER-2/c-erbB-2 (HER-2) for breast cancer and gynecological tumors (about 110 cases/month) has also significantly increased and FISH for HER-2 gene amplification is performed in the Diagnostic Molecular Pathology Laboratory on more than 1200 cases per year.

Immunopathology plays an important role in renal pathology. Drs. Colvin and Smith are the staff members of the renal service. Mr. Collins assists in the interpretation of immunofluorescence and anti-GBM studies. Diagnostic immunofluorescence is performed routinely on all renal biopsies for medical renal disease, mostly glomerulonephritis. These studies are followed up by electron microscopy to verify and substructure the complexes. An additional immunofluorescence panel has been developed to sub-type amyloid deposits for the renal and cardiovascular surgical pathology services. In 1999 Mr. Collins developed a sensitive three-step immunofluorescence technique for the detection of C4d in tissue. He and Dr. Eveline Schneeberger and Dr. Colvin were first to report that capillary C4d deposition in renal allograft biopsies is a marker of both acute and chronic humoral rejection and correlated with presence of circulating antibodies to donor-specific HLA class I and class II antigens. The C4d assay is now used in transplant centers worldwide in the diagnosis of antibody-mediated rejection of allografts.

The Immunopathology Unit is a major reference laboratory for the diagnosis of ANCA and anti-GBM autoimmune disease. Dr. Niles and Mr. Collins direct the ANCA lab. ANCA are detected by indirect immunofluorescence and verified by specific ELISA. Autoantibodies against the glomerular basement membrane (anti-GBM or Goodpasture's disease) are detected by Western blot and quantitated by an antigen specific ELISA.

Dr. Stone is involved in the identification and characterization of amyloid in tissues. Amyloid deposits in tissue are subtyped by immunofluorescence using a panel of antibodies encompassing antibodies specific to common forms of amyloid, including lambda light chains, kappa light chain, serum amyloid A, Apolipoprotein-A1, transthyretin and fibrinogen. The technique requires fresh frozen tissue and is most commonly performed on heart and kidney, but is applicable to any tissue.

Flow Cytometry Laboratory

The Flow Cytometry Laboratory, directed by Dr. Frederick I. Preffer, utilizes a three-laser BD FACSCanto II capable of correlated 10 parameter analysis and one two-laser FACSCalibur capable of correlated 6 parameter analysis. Specimen prep instruments include a BD Lyse/Wash Assistant, A BD Sample Prep Assistant and a BD Medi-Machine.

The laboratory serves the entire Mass General clinical community, as well as outside cases from MEEI, the North Shore Cancer Center and other institutions. The majority of specimens include those obtained from peripheral blood, lymph nodes and bone marrow. However, fluids from various body cavities, [e.g., spinal, thoracic fluids] account for a significant number of additional cases. The laboratory works closely with the department's cytopathology service in immunophenotyping FNA's of lymph nodes and other hematopoietic tissue as well as lymph node and other tissue biopsy specimens obtained in the frozen section laboratory.

Between 1996 and 2009 the number of cases has increased significantly. Some of the laboratory growth has been appreciated in the form of additional new tests offered [e.g. stem cell & T subset analysis]. For example the "stat" stem cell/CD34+ cell assay was instituted in 1999. The introduction of this innovative assay in the Flow Cytometry lab greatly relieved the Pheresis and Bone Marrow Transplantation Unit of unnecessary stem cell collections, saving valuable hospital resources for more necessary uses.

In 2009, the Flow Cytometry laboratory processed leukocyte immunophenotyping for lymphoma and leukemia (3,110 cases), organ transplant T cell monitoring [e.g. ATG/OKT3; 265 cases), HLA purity evaluation (213 cases), Rituximab monitoring (555 cases), stem/CD34+ cell monitoring (456 cases) and T-cell subset (CD4+/CD8+) monitoring of HIV+ and other immunodeficiency patients (4,790 cases).

The laboratory worked very closely with Becton Dickinson to develop a staining robot (SPA) that would automatically process whole blood in a 'hands-off' way, and prepare sample tubes for direct analysis on the carousel of the FACSCalibur. The additional robotics has proved effective and useful in specimen preparation for flow cytometric analysis.

Teaching, Educational Activities and Research

Find more information on the Flow Cytometry Laboratory page.

Electron Microscopy Laboratory

The Electron Microscopy Laboratory, directed by Dr. G. Petur Nielsen, provides both diagnostic and research transmission electron microscopy services. The types of tissues received by the Electron Microscopy Laboratory include tumor specimens, renal specimens (both native and allograft kidneys), neuropathology specimens (skin, muscle and nerve), cytology specimens (fine needle aspirates, bronchial and peritoneal lavages, voided urine, and pleural effusions) autopsy specimens, teaching specimens (cases done for academic interest), "other" specimens (infectious, genetic, and metabolic diseases) and research specimens.