Tell me about using a Nanodrop for protein quantitation

[Bonnie]

I've been doing a microplate BCA assay (diluting my samples 1:100 or 1:1000) to quantitate proteins before doing Western blots, but I noticed that another lab in my building has a nanodrop. It would save me a few hours each time if I could use the nanodrop to read undiluted 2 uL samples instead of doing the BCA.

I do an acetone precipitation on all my samples and re-suspend them in plain PBS, so I believe that my proteins should be free of any contaminating substances that would absorb at 280 nm. My protein concentration is generally in the 1 - 8 mg/mL range, which I think is okay for the nanodrop?

Can anyone think of a reason why I shouldn't use the nanodrop? I'm only hesitant because no one I know seems to use it, but it might just be because they're afraid of technology.

[butters]

so have u test it yet? which model of nanodrop are u using? If you only have few samples why not use the nanodrop but is u have a lot then better use BCA assay. The best would be to have both data and u can compare which one u trust more.

nanodrap is great coz no consumable is use except tips .

[Bonnie]

I ran samples on both the nanodrop and the BCA assay and got very poor agreement between them, so I gave up on using the nanodrop for this purpose. The nanodrop and BCA results were often different from each other by as much as a factor of 2, probably due to other stuff still in solution after the acetone precipitation.

[ppo]

You can make a blank following the steps of acetonoe precipitation but without protein. That could help if there is something interfearing with the method. Otherwise Nanodrop has been used because lacking of calibration
Hope you save some time with nanodrop