Nissl Staining for Neurons
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Description
From the lab of Lucien T. "Tres" Thompson, Ph.D. The University of Texas at Dallas
Nissl Staining for Neurons
This is a general protocol adapted from a protocol on ihcworld.com.
Purpose: Nissl staining uses a cresyl violet solution to stain RNA , which is most abundant in the rough endoplasmic reticulum of nuclei. This method is used to detect the nuclei of neurons in fixed, embedded, and frozen tissue.
Materials
30-50 μm fixed frozen/vibratome sections, mounted on slides
Beakers to mix and hold solutions
Incubation chamber
Solutions
Cresyl fast violet
Glacial acetic acid
100% ethyl alcohol
Chloroform
95% ethyl alcohol / 5% deionized H2 O mixture
Xylene
Resin medium
1) Prepare 0.1% cresyl violet solution by mixing 0.1 g cresyl fast violet mixed in 100 mL deionized H2 O. Add 10 drops of glacial acetic acid just before use and filter.
2) De-fat the tissues by soaking in a 1:1 alcohol/chloroform mixture overnight.
3) Rehydrate the slices in 100% alcohol followed by a 95% alcohol / 5% deionized H2 O mixture.
Note: Putting the slides in pure water would cause the frozen tissue to come off the slides.
4) Stain in cressyl violet for 3-5 minutes.
5) Rinse in distilled water.
6) Soak in 95% ethyl alcohol for 5-30 minutes. Check microscopically for staining .
7) Dehydrate in 100% alcohol for five minutes. Replace alcohol and repeat.
8) Clear in xylene for five minutes. Replace xylene and repeat.
9) Mount with resin medium.
Check for staining . Nissl bodies (neurons) will be stained red-violet.