The invention of trichrome staining.

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Trichrome staining is a process that involves staining combinations that usually contain three dyes of contrasting colours selected to stain connective tissue, muscle, cytoplasm and nuclei in bright colours[1].


The English pathologist Heneage Gibbes invented[2] the trichrome staining process in 1880 after experimenting with various histological stains. Here is the trichrome staining (which he called “treble staining”) method he describes in his 1880 publication:

  • He first stained the histologic sections with picrocarmine and soaked them in acidulated water.
  • He then took a roseine solution and diluted it with alcohol and then immersed the same sections in the solution for 2-3 minutes.
  • Then, he removed the sections and poured denatured alcohol on them to wash off the excess dye.
  • He then placed the sections in a diluted solution of iodine green, after which they were ready for viewing once removed.

On the application of this “treble stain” to histologic sections, he states[2], “This staining process is well shown in a section of the base of a cat or dog’s tongue, cut through one of the circumvallate papillae, the section should be sufficiently large to include some of the mucous glands, of which there are a large number in that region. If the staining is well done it will show all the muscle fibres stained with picro-carmine, the connective tissue, protoplasm of cells, &c., stained with rosein; while all the nuclei in the superficial epithelium, serous glands, non-striped muscle tissue in the vessels, and elsewhere, are stained a brilliant green.” Here is a photomicrograph[3] of a section of a rat’s airway stained with Masson’s trichrome (nuclei are stained dark red/purple, cytoplasm is stained red/pink and connective tissue is stained blue):

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Here is a photograph[4] of Heneage Gibbes:

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  1. stain
  2., pages 39-42.
  4. gibbes-heneage