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NeuroScience Associates

Campbell-Switzer Alzheimer Silver Stain

 

Plaques and Tangles: A Controlled and Enhanced Method

Staining Procedure:

(125×65 dishes for up to 30 sxns max & 100×50 dishes for 13 or less sxns.)

  1. Prepare Physical Developer Solutions A, B & C
  2. Nitric acid wash all glassware.
  3. Remove sections from storage containers and place in dH2O 3 x 10 min each. Use baskets to transfer sections through all steps except the SPC and Physical Developer ABC steps.
  4. Prepare SPC Solutions. Stir until needed. Prepare other solutions.
  5. Place sections in 2% NH4OH. Stir on stirring table for 5 min.
  6. Place in dH2O 2 x 1 min. each.
  7. Place in the SPC Soln. and cover. Stir gently for 40 min.
  8. Place in 1% Citric Acid for 3 min.
  9. Place in 4.99 pH Acetate Buffer Working Solution until ready for Physical Developer.
  10. Prepare fresh Physical Developer ABC Sol’n. for each batch. Develop 10-15 sections at a time. Use fewer sections if they appear crowded.
  11. Place in Physical Developer ABC Sol’n. over a light source. The development time is visually assessed.
  12. Stop development by placing in 4.99 pH Acetate Buffer Working Solution briefly.
  13. Place in fresh 4.99 pH Acetate Buffer Working Solution. Timing is not critical.
  14. Place in fresh 4.99 pH Acetate Buffer Working Solution. Timing is not critical.
  15. Place into fresh dH2O 30 sec.
  16. Place into 0.5% Sodium Thiosulfate solution for 45 sec.
  17. Place into fresh dH2O 3 x 2 min. each
  18. Mount sections.
  19. Coverslip.

 

Other links of possible interest:

FAQ regarding silver stains:

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For further reference, please see: Stains

Solutions:

Silver Stain Solution #1: 1% Silver Nitrate (AgNO3)
100 ml dH2OPrepare and mix well.
1.0 g AgNO3
100 ml 1% AgNO3
Silver Stain Solution #2: 1% Potassium Carbonate (K2CO3)
100 ml dH2OPrepare and mix well.
1.0 g K2CO3
100 ml 1% K2CO3
Silver Stain Solution #3: Silver-Pyridine-Carbonate (SPC)

Batch Sizes
1% AgNO360 ml120 ml180 ml240 ml300 ml360 ml
Pyridine17 ml34 ml51 ml68 ml85 ml102 ml
1% K2CO345 ml90 ml135 ml180 ml225 ml270 ml
Total Volume:122 ml244 ml366 ml488 ml610 ml732 ml
Silver Stain Solution #4: 2% Ammonium Hydroxide
98 ml dH2OPrepare and mix well.
2 ml NH4OH (27-29% concentrate)
100 ml 2% NH4OH
Silver Stain Solution #5: 1% Citric Acid
100 ml dH20Prepare and mix well.
1.0 g Citric Acid
100 ml 1% Citric Acid
Silver Stain Solution #6a: 4.99pH Acetate Buffer Stock
60 ml 1M Acetic AcidPrepare and mix well.
140 ml 1M Sodium Acetate
200 ml 4.99 pH Acetate Buffer Stock
Silver Stain Solution #6b: 4.99pH Acetate Buffer Working Solution
238 ml dH2OPrepare and mix well.
12 ml 4.99 pH Buffer
250 ml 4.99 phH Acetate Buffer Working Solution
Silver Stain Solution #7: 0.5% Sodium Thiosulfate
400 ml dH2OPrepare and mix well.
2 g Sodium Thiosulfate
400 ml 0.5% Sodium Thiosulfate
Silver Stain Solution #8a: Physical Developer
Solution APrepare and mix well.
500 ml dH2O
25 g Na2CO3
500 ml Solution A
Solution BPrepare and mix.
500 ml dH2O
1.0 g NH4NO3
1.0 g AgNO3
5.0 g Tungstosilicic Acid
500 ml Solution B
Solution CPrepare and mix well.
250 ml dH2O
0.5 g NH4NO3
0.5 g AgNO3
2.5 g Tungstosilicic Acid
1.75 ml 37% Formaldehyde
250 ml Solution C
Silver Stain Solution #8b: Physical Developer ABC
Free-Floating SectionsParaffin Sections
Solution A100 ml100 ml
Solution B90 ml80 ml
Solution C10 ml20 ml