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Kinase Assay

kinase protocol specific activity dilution

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#1 pa15

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Posted 10 March 2014 - 11:20 AM

Hello,

 

I'm performing a Kinase Assay. The enzyme is made up from a 10 ug to a concentration of 0.1 ug/ml. I need to dilute the enzyme with dilution buffer. I'm given a specific activity plot with the specific activity of the enzyme in nmol/min/mg. How can I determine how much I need to dilute the enzyme based of the specific activity?

 

Thanks.



#2 mdfenko

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Posted 11 March 2014 - 03:52 AM

you need to determine how many units and a convenient volume of enzyme you want to add to the reaction.


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#3 pa15

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Posted 11 March 2014 - 08:30 AM

Thank you for your response.

Could you please be a little more specific regarding the determination of units. I have already found the specific activity to be 125 nmol/min/mg and plan on using 10uL of my diluted enzyme. I'm just not sure how much it should be diluted. 



#4 mdfenko

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Posted 12 March 2014 - 03:57 AM

how many units do you want in each individual reaction? and what is a convenient volume for you to add?

 

which kinase are you using?

 

here is a link to the worthington biochemicals search page, type "kinase" into the search box and you will get a list of kinases with descriptions, references and assays. maybe the information you want is here?


Edited by mdfenko, 12 March 2014 - 04:01 AM.

talent does what it can
genius does what it must
i do what i get paid to do





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