Hi Weikan, if you mentioned nucleic acids-SIP, the easiest way to know heavy fraction after ultracentrifugation is running a standard with a mixture of 13C-labeled and unlabeled nucleic acids (DNA or RNA). Then you can easily see the density shift between heavy and light fractions.

You need to consider if you used the fully labeled substrates in your incubations. Density shift will be happened when 13C-labeling level increases about 20%. Also, density of nucleic acids depend on GC content, especially for DNA-SIP. Generally, the densities of fully labeled nuleic acids are about 1.757 and 1.815 g/ml for DNA- and RNA-SIP respectively (Lueders et. al 2004) Article Enhanced sensitivity of DNA- and rRNA-based stable isotope p...

Dear Weikan,

I fully agree with Xiuran Yin's suggestions. Regarding the practical approach, how to differentiate isotopically "heavy" and "light" fractions in a nucleic acid SIP experiment, you need a refractometer for measuring the density of fractions as described in Lueders et al. 2004, or Neufeld et al. 2007 Nature Protocols. An important point for running isopycnic centrifugation is the calibration of your gradient buffer, which is done gravimetrically (e.g., weighing defined volumes of the gradient buffer and of fractions after isopycnic centrifugation on a sensitive analytical balance).

I also fully agree with the comments. I recommend you a JoVE video protocol which was published by Dunford and Neufeld, 2010. The video will also answer your question.