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The page below is a sample from the LabCE course Histology Special Stains: Connective Tissue. Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

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Phosphotungstic Acid-Hematoxylin (PTAH) Stain - Chemistry

The PTAH staining method relies on acid-base chemistry to stain collagen and muscle fibers. These fibers are demonstrated using a tungsten mordant provided by the phosphotungstic acid. This mordant binds hematin and stains selective tissue components blue, while the phosphotungstic acid is believed to stain other tissue components a red-brown color.
Traditionally, tissue sections were preferably fixed in Zenker's fixative. However, due to regulatory initiatives to remove all mercury from laboratories, Zenker's is no longer readily used. Several commercial stain vendors now offer Zinc chloride as an alternative to Zenker's.
A section of skeletal muscle tissue can be used for quality control.
The amount of phosphotungstic acid in the staining solution is far greater than the amount of hematein and it is believed that tungsten binds all available components blue, while the phosphotungstic acid is thought to stain the red-brown components. This stain has been referred to as a polychrome stain because one solution gives two major colors. The components colored red-brown will lose this color with water or prolonged alcohol washes. Therefore, dehydration of the tissue section following staining MUST be rapid.