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Dried blood spots internal standard marker protein

dried blood spot protein internal standard

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#1 biochem06

biochem06

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Posted 30 October 2013 - 06:04 AM

Hi,

 

I'm using dried blood spots (whole blood, not separated) to measure some markers of inflammation eg CRP. I want to compare the results from the dried blood spots to frozen plasma.

 

The problem is this: I add 10ul of blood to filter paper and let it dry etc. Then puch out a 3mm circle of the dried blood for analysis. This 3mm circle is extracted in 50ul of extraction buffer and then assayed. However, I don't know how much blood I have actually extracted from the dried blood spot and it has also been diluted in extraction buffer.

 

I want to compare the results to a known volume of frozen plasma. So how do I do this?

 

I was thinking of adding an internal standard to the sample before adding the blood to filter paper, that way I will know how much I have extracted as I can measure the amount of internal standard.

 

What could I use for the internal standard? It must be cheap, stable at room temperature and easy/cheap to measure. It must also not react with the blood.

 

I was thinking maybe a protein from another species e.g. horse myoblobin (could measure by ELISA although this is expensive) or a stable dye that can be measured using a spectrophotometer (we don't have access to mass spec).

 

any thoughts or suggestions would be greatly appreciated!







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