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OD as units for amount of DNA, or concentration?

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OD as units for amount of DNA, or concentration?

Postby microphobe » Mon Aug 08, 2005 10:33 am

I have always thought that OD (optical density) was a measure of concentration, but now I am confused.

I have seen more and more often, OD being used to refer to absolute amounts of DNA or RNA. For instance, tubes of lyophilised PCR primers that give the "OD units" but not the volume - so AFAIK you cannot figure out the yield, can you?

Also, Maniatis says to load 0.5-0.7 OD260 units of RNA per lane for a Northern blot. How do you convert to nanograms without knowing the volume of solution??

Thanks
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Postby iri_black » Mon Aug 08, 2005 12:40 pm

Optical density is the absorbance of an optical element for a given wavelength λ per unit distance.
It is used in spectrophotometry, equal to 2 minus the log of the percentage transmittance of light.

At liest I think so...... :oops: :?
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Postby chemistry_freako » Mon Aug 08, 2005 1:51 pm

yea..it measures the absorbance of a sample - quantitative analysis. Most spectrophotometers are accurate for values lower than 1 - cos the standard curve starts to deviate from the Beer- Lambert Law at high concentrations (e.g. due to stray light).
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Postby canalon » Mon Aug 08, 2005 2:38 pm

I have always thought that OD (optical density) was a measure of concentration, but now I am confused.

I have seen more and more often, OD being used to refer to absolute amounts of DNA or RNA. For instance, tubes of lyophilised PCR primers that give the "OD units" but not the volume - so AFAIK you cannot figure out the yield, can you?


Yep, PCR primers are often sold as unit (You order from at least 0.5U to 10U depending on your needs). Of course those are concentrationd of DNA after synthesis, and do noy correspond to anything once lyophilised... But you will usually find the absolute quantity in ng witten on the tube, which is the only useful information.

As for the Maniatis, it just give the OD as a shortcut: Since you measur you RNA concentration by OD, it gives you an OD value (if you have 0.1OD 260 Units per ul, load 5 to 7 ul... and so on). At least you won't make any miscalculations in your conversions :lol:

HTH

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