NADH NBT Staining
From Bridges Lab Protocols
Revision as of 18:20, 27 August 2018 by Lgunder (Talk | contribs) (Created page with "== Tris Buffer .2M pH 7.4 == #Warm Tris buffer to 37°C in water bath. #Use hot plate to keep the buffer at 37°C, pH the buffer with HCl. (pH changes with temperature) ==Met...")
Tris Buffer .2M pH 7.4
- Warm Tris buffer to 37°C in water bath.
- Use hot plate to keep the buffer at 37°C, pH the buffer with HCl. (pH changes with temperature)
Methods
- 20mg NBT aka Nitrotetrazolium Blue Chloride or Nitro Blue Tetrazolium
- 80mg of NADH aka β-Nicotinamide adenine dinucleotide, reduced disodium salt hydrate
- 100mL of Tris Buffer
- 100mL fits in a staining dumpster
- Incubate for 30 minutes at 37°C
Dehydrate and coverslip
- Rinse 3-4 times under running DI water
- Immerse in 50% ethanol for 2x 30 seconds
- Immerse in 70% ethanol for 2x 30 seconds
- Immerse in in 95% ethanol for 2x 30 seconds
- Immerse in in 100% ethanol for 2x 30 seconds
- Immerse in 1:1 xylene ethanol for 2 minutes for 2x 30 seconds
- Repeat twice more using fresh xylene. Check staining under the microscope
- Coverslip using Permount
