Re: fluorescence in situ zymography

From:Thu-Thao Pham

I have been dissolving my PMSF in 100% Ethanol and have no problem.


>From: "Abigail.Mackey" 
>To: "''" 
>Subject: fluorescence in situ zymography
>Date: Thu, 06 Dec 2001 17:58:12 +0000
>Hello All,
>One other issue with this method to resolve, hopefully the last one!
>I need some advice on the use of PMSF(phenylmethylsulphonylfluoride) in
>solution. The solution contains:
>50mmol Tris-HCl, pH 7.4,
>10mmol CaCl2*2H2O,
>0.05% Brij 35,
>5mmol PMSF
>Fluorescence-conjugated gelatin is then added to this solution and 
>with tissue sections to detect gelatinase activity. As I understand it, 
>is added to inhibit any serine proteases which may be present in the 
>To get the PMSF to dissolve, the solution needs to be heated to almost
>boiling. My first question is will the heating destroy the PMSF action? If
>so how do I get it to dissolve another way, can it be dissolved in ethanol
>for example? Will the addition of alcohol have any other effects on my
>thanks in advance,
>Abigail Mackey
>Department Sport and Exercise Sciences
>University of Limerick
>tel +353 (0)61 20 28 84
>fax +353 (0)61 33 04 31

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