Lapis ISSN 2398-2969

Staining technique: Papanicolaou

Synonym(s): Pap

Contributor(s): Molly Varga, Sarah Pellett

Introduction

  • Papanicolaou was originally associated with cervical screening, staining cervical cells in three stages with competing dyes.
  • The Papanicolaou stain is also used for clinical material and is normally used to stain bodily secretions, fine needle aspirates and touch preparations.
  • Two dyes are used that differ in their affinity for various sites within a cell, ie nucleus and cytoplasm.
  • Different to Romanowsky stains in that there is no chemical interaction between nuclear and cytoplasmic solutions. They bind electrostatically, resulting in salt formation.
  • Similar to hematoxylin and eosin, except that eosin is replaced with two cytoplasmic counterstains. This highlights keratinization in squamous epithelium.
  • The important elements of Papanicolaou staining methods are nuclear staining followed by orange and polychromic cytoplasmic staining (hematoxylin stain + orange stain + polychromic stain = Papanicolaou stain).
  • Using this method, cells are highly transparent allowing diagnosis, even when cells are overlapping and when mucus or inflammatory cells are present.
  • The staining method can be applied either manually with the aid of cuvettes, or mechanically in automated staining systems.

Uses

  • Used in special diagnostic situations, especially where neoplasia is suspected.
  • Provides architectural information for cytopathologists and allows nuclear detail to be visualized even in thick smears and tissue fragments.

Advantages

  • Provides excellent detail of nuclear and nucleolar changes.
  • Unlike other stains, it highlights cytoplasmic keratin; stain appears orange to orange-red.
  • Wet fixation causes red cell lysis, improving visual examination of clumps of cells; useful for smears heavily contaminated in blood.

Disadvantages

  • Compared with Romanowsky stains, cytoplasmic detail and organisms, including bacteria, are not well defined.
  • Cells must be immediately wet fixed in alcohol or by using a spray fixative, unless use modified Papanicolaou where smears are rehydrated.
  • Slides must not be allowed to air-dry between steps or between staining and coverslip mounting.
  • Adhesive-coated slides (poly-l-lysine, gelatin) may aid cell retention.
  • Time consuming; not very practical for a veterinary practice laboratory.

Requirements

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Procedure

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Further Reading

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