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Occam's Maser's Content

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#145446 RNA isolation from small numbers of cells

Posted by Occam's Maser on 17 November 2012 - 03:46 PM in Molecular Biology

If I might offer one further problem for discussion; I just yesterday discovered how terribly 260/280 absorbance quantitates DNA at low concentrations. I am extracting DNA (and hopefully RNA) from small numbers of FACS sorted human cells. Turns out NanoDrop/Vue grossly overestimate DNA concentrations. I was recommended the Qubit fluorometry system which worked well at my first attempt and gave results far more in keeping with the number of input cells (500) rather than 2-3 logs higher concentrations claimed by absorbance. I am now very suspicious of my "good" RNA readings from NanoDrop... sigh... Qubit has an RNA option too, which I hope to try. Good luck and please update us if your protocols all work out.




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