I don't know if you can trust any staining procedure to leave the cells
totally unharmed, but methylene blue has been used for decades in crustacean
systems. The nerves remain functional with staining.
I don't have my lab notes with me here at home. The histology texts include
elaborate procedures for preparing reduced methylene blue. However, the
physiological preparation usually just involves dissolving the oxidized
(blue) form in solution (Van Harreveld's?). Make sure you filter it before
using -- it leaves lots of needle-like crystals deposited over the tissue.
The concentration is not too important -- try saturated. Drop a few
droplets of the stain directly on the tissue and let is sit a few minutes.
Then rinse everything off in fresh saline and wait for the dye to "develop".
I believe it has to be reduced to colorless form and taken into the cells,
then re-oxidized back to blue. Keep adding more dye and washing it away as
needed. Be patient, it may take ten or twenty minutes for things to start.
Also note that torn muscle and connective tissue take up the stain, so
careful and clean dissection is important.
Of course you should practice it first on a test system. The walking legs
are good preparations, especially the meropodites. On a good preparation
you can see the individual fibers innervating the flexor and extensors, as
well as the accessory flexors. I have used it on the cardiac ganglion and
the stomatogastric system, tracing the pathway of nerves to the stomach
muscles. It is also nice for showing the complex innervation of the
abdominal stretch receptors. And, of course, you can always try it on the
superficial flexors!
Soon Hyouk Lee wrote in message <7u5gei$9en$1 at merrimack.Dartmouth.EDU>...
>Would methylene blue end up destroying the cell? The safety sheet for
>methylene blue states it being somewhat toxic, and I'm afraid that the
nerve
>will be unusable for recording and/or stimulation after treatment with such
>a stain.
>>Befuddled,
>>Soon Hyouk
>>Soon.Hyouk.Lee at dartmouth.edu>>----------
>In article <cobN3.10370$Ud2.372896 at typhoon1.rdc-detw.rr.com>, "Richard
>Norman" <rsnorman at mediaone.net> wrote:
>>>> Would a vital stain like methylene blue work?
>> It should stain the nerve well enough so that you can dissect it out of
the
>> capsule.
>>>>
