Picric acid was the first synthetic dye to be made, in 1771 by Woulfe. It is now most commonly used in conjunction with acid fuchsin in the Van Gieson method which demonstrates collagen (red) in contrast to cytoplasm and muscle (yellow). It is also used, with or without other yellow acid dyes, to colour erythrocytes in trichrome methods for fibrin such as Lendrum's Picro-Mallory. It is a fixing agent and is a constituent of Bouin's fluid. It has been used as a differentiating acid in a variant of Heidenhain's iron hematoxylin, and is a trapping agent for methyl violet in the Gram Weigert stain for bacteria and fibrin.
Important safety note
Picric acid is explosive when dry, although it is safe to handle with standard techniques.
It must be kept moistened with water or ethanol at all times.
Be careful if tissues are immersed in a large container of a picric acid containing fixative by suspending from a string. The string can act as a wick, and large crystals of picric acid can form on it.
Measuring picric acid safely
Do not measure picric acid by drying and weighing. Instead, make a saturated solution and calculate the amount from that.
Saturated aqueous picric acid contains 1 gram in 78 ml.
Saturated ethanolic picric acid contains 1 gram in 12 ml.
Making saturated ethanolic picric acid
Drain water damped picric acid through a standard filter paper.
Drain thoroughly, but do not allow to dry.
Place into a container and just cover with absolute ethanol.
Leave for a few minutes, then drain off the ethanol.
Repeat the last two steps once more.
Cover the picric acid with sufficient absolute ethanol to make a stock solution.
Leave at least 24 hours for a saturated solution to form.
Store away from flames and sparks in an explosian safe container.
Add more absolute ethanol as needed.
Ensure that some undissolved picric acid is always present.
Reference R. D. Lillie. Conn's Biological Stains
Williams & Wilkins, Baltimore, MD., U.S.A.
Edward Gurr, (1971) Synthetic dyes in biology, medicine and chemistry
Academic Press, London, England.
Pope Sine, (2003) Synthetic dyes
Rajat Publications, New Delhi, UK.