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Staining techniques: Wright's stain

ISSN 2398-2977


Synonym(s): Romanowsky stain

Podcast: Staining techniques: Wright's stain

Introduction

Rationale

Uses

  • Hematology and cytology.

Pros

  • If Hemofast is used, it contains neutral methyl alcohol, ie does not contain acetone, therefore the solution can fix itself and there is no need for prior fixing of the smeared slides.

Cons

  • Must use distilled water for Hemofast.
  • Cannot use tap water since the latter contains interfering substances, particularly calcium which can cause precipitation of eosin to an insoluble calcium salt.

Alternatives

Work time

  • 5-15 min depending on whether the stain has been prepared or is ready mixed.
  • Refer to methodology for details.

Requirements

People competence
  • Veterinary: medium.
  • Nursing: medium.

Instrumentation

Equipment

  • Minimum: Coplin jars, light microscrope.

Consumables

  • Minimum: distilled water, clean microscope slides, Wright's stain and buffer or Hemofast.

Method

  • Prepare Wright's stain and Wright's buffer or use commercially prepared solutions, eg Hemofast by Mascia Brunelli (see below).
  • Prepare thin blood films or cytology preparations, air dry quickly.
  • Cover slide with Wright's stain.
  • Leave immersed for 3 min.
  • Add an equal volume of Wright's buffer.
  • Gently rock the slide to mix the solutions until a green metallic sheen forms on the surface of the mixture.
  • Leave stain - buffer solution on for 6 min.
  • Rinse under running tap water.
  • Air dry.
  • Examine with light microscope as recommended blood film.

Manual staining with Hemofast

  • Prepare Wright's stain and Wright's buffer or use commercially prepared solutions, eg Hemofast by Mascia Brunelli.
  • Prepare thin blood smears or cytology preparations, air dry quickly.
  • Fill a Coplin Jar with Hemofast stain.
  • Immerse the slides for 5 min at room temperature.
  • Gently rinse slides under running distilled water.
  • Place slides in another Coplin jar filled with distilled water for 10 min (diluted stain).
  • Remove slides and allow to drain and dry.
  • Examine with light microscope.

Result data

Normal
  • Colors of cells/components vary slightly with the variation of pH of the distilled water (Hemofast stain).

Storage

  • Must be stored in dark (non-metal) container in a dry area, since light and humidity alter Hemofast.
  • Poisonous and flammable, therefore store away from heat sources.

Artefacts

Errors

Further Reading

Publications

Refereed papers

Other sources of information

  • Lewis H B & Rebar A H (1979) Bone Marrow Evaluation in Veterinary Practice. Ralston Purina Co. pp 59.