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Citrate synthase standard for CS assay


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#1 girlfrommars

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Posted 21 May 2013 - 07:39 AM

Hi,

I recently performed a citrate synthase assay on rat skeletal muscle. I decided to use a commerical citrate synthase as a control for the experiment. The CS control states that 1 unit of it will form 1 umol of citrate from OAA and Acetyl co A. When analysing my data I came across a problem, I am unsure how to determine if the assay worked correctly using this control. I calculated the results to umol/min using 1 unit of citrate synthase. But I got results as low as 0.3 umol/min. If I take into account the mg/ml concentration of the citrate synthase and calculate the data the same as my samples I get ~130 umol/min/mg using 1 unit of citrate synthase.

Has anyone else done this assay? And if so do they know how to do the calculation?
I except you ment to get something around 1, if it is a bit off then you can use this to normalize you data as you know the reaction is slightly off. But I feel like I must be doing something totally wrong for mine to be this off, or I am not taking all factors into account.

Any help would be greatly appreciated!

#2 bob1

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Posted 21 May 2013 - 05:23 PM

Units for enzymes are usually defined by the amount of substrate per time. for many the unit of time is an hour - so perhaps this is your problem?

#3 mdfenko

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Posted 22 May 2013 - 04:49 AM

are you controlling temperature?

mixing thoroughly?

how are you assaying (kit or homebrew)?
talent does what it can
genius does what it must
i do what i get paid to do




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