Staining

Question 1

Stain Profile

PTAH
Phosphotungstic acid hematoxylin

Purpose, principle, optimal fixative, & procedure

From Self Instruction textbook, notes & etc

Answer 1

Purose: Method of Mallory used to demonstrate the cross-striations arrangements of actin & myosin contractile proteins, and also fibrin in skeletal muscle. The cross-striations are a diagnostics feat of rhabdomyosarcomas (et other skeletal muscle diseases). Can be used to demo glial fibers & myelin. Outdated because of IHC.
Principle: PTA to hematein 20:1, tungsten bins to all available hematein to give blue lake. Metal-hematein lake stains blue (cross-strations, fibrin & nuclei) and PTA stains red-brown (collagen). A polychrom stain.
Fixative: Zenker>NBF

Question 2

Stain Procedure

Phosphotungstic acid hematoxylin (PTAH)

From Self Instruction textbook, notes & etc

Answer 2

1. Deparaffinize sections & hydrate to distilled water.
   2.Formalin fixed, then mordant in Zenker with 5% acetic overnight
   3.Rinse in tap
   4.Place in Gram iodine for 15min
   5.Rinse in tap
   6.Place in 5% aqueous sodium thiosulface 3 min (mercury removal)
   7.Wash in tap for 10min
   8.Place sections in 0.25% potassum permanganete for 5min
   9.Rinse in tap
   10.PLace in 5% oxcalic acid for 1min
   11.Wash in running tap 10 min
   12.Stain in PTAH solution overnight
   13.Dehydrate quickly in 2x 95% & 100% ROH, clear in xylene & then mount with synthetic resin
   Remember Zenker contains mercuric chloride so the steps with gram iodine & sodium thiosulfate are critical to avoid fixation pigment