Standard curve using ImageJ - (Jun/14/2016 )

Hy,

I want to make a standard curve using the intensity/area of DNA Ladder by ImageJ for measuring the amount of dna.

I measured the intensity of ladder's bands (the image from the producer) and I made a standard curve. Now I want to correlate the intensity of dna from the gel.

I don't know if my approach is the best or correct and I would like an opinion from you.

Thank you!

Your general approach is correct, but you must load standards with every gel and use those to generate a standard curve each time. You can not use images from previous gels (or the ones provided by the producer) and different exposures as these will lead to erroneous results.

Bottom line: each time you run a gel, load the standards as well and use those.

Thank you!

I run the samples with 2 microliters dna marker, but the image from the producer was obtained by using 1 microliter. In order to get the standard curve can I double the concentrations of the marker. If 500 bp is coresponding to 73 ng (1 microliter) can I consider for 2 microliter 146 ng?

Yes. In fact you can apply this to any volume you choose. Note that to obtain an accurate standard curve your sample should be about the same length as the standard you are comparing to as length and brightness can be correlated for these sorts of things.

Thank you very much for your help!