Dear Maria,
When assaying PLA2's in the past I used the assay of Radvanyi, F., Jordan,
L., Russo-Marie, F. & Bon, C. (1989) Anal. Biochem. 177, 103-109. It's a
fluorescence based assay so you need access to a spectrofluorimeter. I
found there was a real art to achieving good reproducibility with this
assay, but it works quite well when you have a feel for it.
Alternatively, there is the assay of Seilhamer, J. J., Plant, S.,
Pruzanski, W., Schilling, J., Stefanski, E., Vadas, P. & Johnson, L. K.
(1989) J. Biochem. 106, 38-42 is based on radiolabelled phospholipids. I
didn't use this assay personally, but it was used in my old lab. To me this
assay seemed more robust.
As far as getting activity goes, that is going to depend on your particular
PLA2 enzyme that you are measuring. You will need to vary the type of
phospholipid substrate according to the enzyme. As you may know, some PLA2
will only work on some substrates, so you need to ensure you have the right
type of substrate for your enzyme.
Please get back to me directly should you have more questions.
Regards,
Peter.
At 19:11 27/10/00 +0200, mlorenzi@ascu.unian.it wrote:
>Hi, I would like to have an assay of this enzyme. I have the commercial one
>but it seems not to be active or the assay which I use is not the right one.
>Please again, I would like to know a sure assay to test this enzyme and if
>the lipid which I have to use like substrate have to be previously treatetd
>to break the micelles.
>I hope to receive an answer soon
>Thanks
>Maria Lorenzi
I'm afraid I don't have a clever saying to put here.
Peter Hains (PhD) Ph. +61 2 9850 6216
Australian Proteome Analysis Facility Fax. +61 2 9850 6200
Level 4 Building F7B E-mail. phains@proteome.org.au
Macquarie University, Sydney 2109 Web. www.proteome.org.au
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