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MTT assay, protein assay analysis


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6 replies to this topic

#1 ayman

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Posted 15 April 2011 - 05:24 AM

For an experiment i recently conducted as part of a university practical, not much assistance was given in how analysis of both these techniques is meant to be done and after days of searching the internet for examples I've come up empty handed. So i was wondering if anyone could offer some assistance in aiding me with analysis the data. If so could you drop me an email at ae86@uni.brighton.ac.uk

#2 ayman

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Posted 15 April 2011 - 05:25 AM

or offer ur assistant on here as well, thanks

#3 laurequillo

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Posted 15 April 2011 - 06:35 AM

With the MTT is easy, you just have to asign 100% to your positive control, and check what is the percentage of your treated samples. That is the easiest way, where you have the % of living cells respect your control. You can rest as well the value of the blanks if you want.

If you have a curve with a known number of cells, you just have to create your equation with those values (value/Number of cells), and one you get the ecuation (your tendency line), you just introduce the values of your samples in that ecuation and you will have the number of surviving cells per sample
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#4 ayman

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Posted 19 April 2011 - 06:22 AM

With the MTT is easy, you just have to asign 100% to your positive control, and check what is the percentage of your treated samples. That is the easiest way, where you have the % of living cells respect your control. You can rest as well the value of the blanks if you want.

If you have a curve with a known number of cells, you just have to create your equation with those values (value/Number of cells), and one you get the ecuation (your tendency line), you just introduce the values of your samples in that ecuation and you will have the number of surviving cells per sample


Thanks

#5 ayman

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Posted 19 April 2011 - 06:42 AM

what is the purpose of preparing standard curves for cell numbers v MTT activity prior to live/dead, hydrogen peroxide killing experiments?

#6 laurequillo

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Posted 21 April 2011 - 02:04 AM

you use to have number of living cells, instead of % of living cells respect a control (I am not sure if I understood your question)
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#7 ayman

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Posted 22 April 2011 - 12:40 AM

you use to have number of living cells, instead of % of living cells respect a control (I am not sure if I understood your question)


I meant the first experiment we did was the MTT assay to provide a standard curve vs MTT activity, prior to doing the live/dead assay and the protein assay. A question that arose was why would we do the MTT assay first before the other two but I guess, like you said we would have needed a control for the experiment and if I am right in assuming, the MTT assay is the most accurate and precise of the three assays?, therefore the most plausible as the control assay?




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