Fluroscent Glutamate Release Assay using Synaptosomes

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Saurabh 5555
Saurabh 5555's picture
Fluroscent Glutamate Release Assay using Synaptosomes

I am trying to establish a fluorescent glutamate release.  This assay uses NADP, Calcium chloride and Glutamate dehydrogenase enzyme. Glutamate dehydrogenase acts on released glutamate( which is released on depolarizing synaptosomes by KCl, 4-AP) and NADPH is produced which gives a florescence signal at 340 nm (emission wavelength) and 460 nm (emission wavelength) I am preparing Synaptosomes by Percoll gradient method. I am not able to deploarize synaptosomes using potassium chloride. I have tried increasing NADP, Glutamate dehydrogenase but nothing seems to give a signal with KCl. I am not able to find a reason for no effect by KCl.

Filsy
Filsy's picture
 Hi Saurabh,

 Hi Saurabh,

I saw your post from 2010 on glutamate release. Have you been successful in getting the assay to work with synaptosomes. I am now studying glutamate release from synaptosome. If you have a protocol that works will be good. 

Thank you,
Filsy