Special staining

Special staining

1. 1.Golden & sweets

   • for reticulin fiber
   • Principal
     • Oxidation
       • oxidation of the hexose sugars in reticulin fiber to expose aldehyde
     • Sensitization
       • metallic complexes is deposited at the aldehyde groups
     • Silver impregnation
       • 1.silver salt and ferric complex exchange ions 2.ammoniacal silver solution is reduced by the exposed aldehyde group to metallic silver
     • Developing
       • further reduction of diamine silver by transferring the section to formaldehyde
   • Procedure
     • 1.Toning:metallic silver is converted to metallic gold(more stable and better contrast)
     • 2.Fixing:unreduced silver and excess gold chloride is remove By sodium thiosulphate
     • 3.Counterstain:use nuclear fast red or light green

2. 2.Jones Methenamine silver stain

   • used for demonstration of glomerular and tubular basement membrane
   • Principal
     • oxidation
       • by periodic acid/chromic acid to form aldehyde groups
     • silver impregnation
       • aldehyde groups reduce silver ions from: methenamine silver to metallic silver
   • Procedure
     • 1.argentaffin:use of sensitizing and reducing solution are not necessary 2.toning:gold chloride convert metallic silver to metallic gold 3.fixing:use sodium thiosulfate to remove unreduced silver 4.counterstained:using light green

3. 3.Verhoeff-van gieson stain(VVG)

   • 1.regressive staining method-overstained with a solution of Hx-ferric chloride-iodine  for the demonstration of elastic fibers
   • Role of ferric chloride and iodine -act as a mordant -act as oxidizer to convert hematoxylin to hematein
   • 2.differention
     • use diluted ferric chloride:break the tissue-mordant-dye complex
     • stained elastic fiber
       • it has a higher affinity for iron-Hx complex, dye retained
   • 1.overstained
     • Hx-ferric-chloride-iodine
   • 3.removal
     • use sodium thiosulfate to remove excess iodine and ferric chloride
   • 4.counterstain use van-gieson
   • Results
     • 1.Collagen:red 2.elastic fibers and nuclei:black 3.Others:yellow
   • Techniques
     • 1.flash verhoeff solution is used 2.overdifferentiation:loss finer fibers 3.prolonged staining in van gieson counterstain:continuous to differentiate(contain picric acid)

4. 4.Trichrome

5. 5.Phosphotungstic acid-Hx

   • to demonstrate cross-striations of skeletal muscle
   • results
     • collagen and muscle fibers are demonstrated -using a tungsten mordant :it binds hematein and stains selective tissue component blue
     • phosphotunstic acid: stain other tissue components red-brown color
   • enhancement
     • post-fixed in zenker's solution: intensify staining reactions
   • technical considerations

6. 8.Unna orcein Technique

   • aldehyde residues are responsible:there are responsible for cross-linking during elastic fiber maturation.
   • counterstain
     • van gieson,H&E and methylene blue
   • oxidation
     • elastic fibers naturally contain aldehydes: oxidation by potassium permanganate:increase the number of free aldehyde groups
     • increase the amount of reactive groups facilitate binding of dye molecules and intense staining of elastic and pre-elastic fibers

7. 6.Iron Hx**

   • stain nuclei and other structure blue to black

8. 7. lipid demonstration

   • stain with Sudan and oil red O
   • principal
     • 1.demonstrate physical processes rather than chemical interactions
   • features of dye
     • The dyes are dissolve in lipid solvent: The dye molecule are found to be more soluble in cellular  lipid than in the dye solvent
     • importing positive stain color to the tissue lipid
   • technical corncern
     • 1.frozen section must be used. 2.tissue must not be exposed to fat solvent: such as xylene, alcohols 3.aqueous mounting media must be used