Alkaline Phosphatase-86

Reagent Preparation

Reagents are provided ready for use. Fixative: Take 25 mL Citrate solution and add 65 mL Acetone, and 8 mL formaldehyde. Seal tightly and refrigerate for up to 4 weeks.


  1. Add 1 mL Sodium Nitrite solution to 1 mL FRV-Alkaline solution OR add 1 mL Sodium Nitrite solution to 1 mL FBBAlkaline solution. Mix by gentle inversion and allow to stand for 2 minutes.
  2. Add solution from step 1 to 45 mL of room temperature deionized water.
  3. Add 1 mL Naphthol AS-BI Alkaline solution to diazonium salt solution (step 2). Mix thoroughly and pour into a Coplin jar.
  4. Fix slides Citrate/Acetone fixative at room temperature for 30 seconds.
  5. Rinse gently in deionized water.
  6. Place slides in Naphthol/diazonium salt solution and incubate for 15 minutes protected from direct light.
  7. Rinse slides for 2 minutes in deionized water. Do not allow slides to air dry.
  8. Counterstain for 2 minutes. If using FRV-Alkaline solution counterstain with Hematoxylin solution. If using FBBAlkaline solution counterstain with Neutral Red solution.
  9. Rinse slides thoroughly in tap water and air dry.


Sites of alkaline phosphatase activity will appear as blue or red granular staining.

See product insert sheet for method of scoring stain intensity.

Alkaline Phosphatase-86


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