Worm Breeder's Gazette 7(1): 72

These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.

Histochemical Staining of C. elegans Embryos for Acetylcholinesterase Activity

S. Siddiqui

Figure 1

I have used the following procedure to stain C.  elegans embryos 
histochemically for AChE activity.
1.  On a microscope slide subbed with a 1% BSA solution, place a few 
young adults into a 20 l drop of 2% glutaraldehyde solution in 100 mM 
maleate buffer, pH 6.0.  The animals are cut with a scalpel blade at 
the vulva to release the embryos.
2.  Very gently, a 16 X 16 mm coverslip is placed on top of the 
buffer drop.  The slide is then immersed into a liquid nitrogen 
container for about 15 seconds, and the coverslip is quickly pried off 
with a scalpel blade.
3.  Another 16 X 16 mm coverslip with its two opposite edges coated 
with grease is placed over the embryos.  The slide is incubated in a 
humid chamber at O C for two hours.
4.  About 10 l of the staining solution ( containing 10 mg 
acetylthiocholinechloride, 8.8 ml 100 mM maleate buffer, pH 6.0.  1ml 
50 mM sodium citrate, 100 l 300 mM copper sulfate, and 100 l potassium 
ferrocyanide, added in this order) is applied at the open edge of the 
coverslip, and is absorbed at the other open edge with an absorbant 
paper ( see fig.).  This step is repeated 3-4 times to ensure that all 
embryos have been immersed in the staining solution.  The slide is 
then incubated in a humid chamber at room temperature for about 6 
hours before light microscopy.  A reddish-brown stain appears in the 
regions showing AChE activity.
[See Figure 1]
The above procedure can also be used for larvae and the adult 
animals.  Significantly, this method reveals the same areas of AChE 
activity in C.  elegans as in the method reported by Culotti et al.  (
Genetics: 97, 281-305,1981), in which the animals are treated with 95% 
acetone for 3 minutes.  The prominent areas showing the stain for AChE 
activity are, the nerve ring, lateral ganglion, ventral nerve cord, 
dorsal nerve cord, pre-anal ganglion, and the pharryngeo-intestinal 
valve cells.

Figure 1