Chapter 34: Electron Microscopy

INDICATIONS FOR ELECTRON MICROSCOPY

Ancillary to standard techniques (i.e., light microscopy) to resolve diagnostic difficulties in human histopathology through examination of ultrastructural findings at the cellular and organelle level

• Unclassifiable, undifferentiated neoplasms

• Supporting a diagnosis from a list of differential diagnosis

• Supporting a light microscopic diagnosis

• Determination of the primary site in metastatic neoplasms

• Medical disease of kidney

• Metabolic storage diseases

• Other congenital disorders

• Infectious agents

• Autoimmune diseases

• Certain cutaneous diseases

• Identification of foreign material in tissues

TECHNIQUE AND TISSUE PREPARATION

• Tissue preparation is similar to conventional wax embedding light microscopy: aldehyde fixation, dehydration, embedding, sectioning, and examination of sections exposed to some form of radiation

• Electron microscopy differences include

  • Image is formed by the scattering of electrons by heavy metal atoms (introduced as solutions of uranyl acetate and osmium tetroxide or lead citrate) selectively adherent to tissue sections

  • Tissue is embedded in epoxy, allowing for sectioning to a thickness of 100 nm

  • Tissue should be submitted in appropriate media (i.e., 2.5% glutaraldehyde; Karnovsky's [universal] solution)

• Fixation with glutaraldehyde provides the best structural preservation; unlike formaldehyde, glutaraldehyde is slowly penetrating. Therefore, only very small pieces of tissue are processed (i.e., 0.5–1 mm³ or 2–3 mm² and thickness of about 0.5 mm)

• Although processing and staining are labor intensive, turn around time can be as short as 24 to 48 hours

• Electron microscopy may also be performed on formalin fixed material from deparaffinizing a wax block. Preservation may be sufficient for diagnostic purposes although results may be variable

DIAGNOSTIC CELLS AND ORGANELLES

Langerhans Cell

• Bone marrow–derived (Fig. 34-1)

• Antigen-processing and -presenting cells

• Indented nucleus with ropey nucleolus

• Rod- and racket-shaped (terminal expansion) cytoplasmic granules (Birbeck granules) with central dotted line

• Often seen at the cell surface when membrane-bound antigen is internalized by endocytosis

• Cytoplasm contains dispersed vimentin intermediate filaments

Merkel Cell

• Slowly adapting type I mechanoreceptors located in sites of high tactile sensitivity (Fig. 34-2)

• Present among basal keratinocytes

• Nucleus is lobulated

• Cytoplasm is electron lucent with prominent Golgi

• Margins of cells project cytoplasmic spines toward keratinocytes

• Typical granules (80–200 nm) have a dense core, halo, and a slightly ruffled membrane

• Granules contain neurotransmitter-like substances

• Intermediate filaments are numerous and assume a parallel or whorled arrangement near the nucleus (dot-like pattern)

Lamellar Granules

• In the ...