Analysis of gradients

Once samples have been collected from a gradient, it may be important to determine their density by collecting fractions from a blank gradient run in parallel. The most direct method is to weigh accurately known volumes of liquid using a pycnometer; however, this is very time consuming. 

It is more convenient to determine the density of a fraction by measuring the refractive index (RI), which has the added advantage of requiring as little as 20-50 µl of sample.

For extensive tables relating % (w/v) concentration of iodixanol, density and RI of iodixanol solutions produced by the dilution of OptiPrep with routine buffered saline solutions see Application Sheet C01. 

Because the RI of gradient solutions is increased by the presence of other solutes (e.g. salts, buffer etc), the precise value of the RI will vary with the presence and concentration of these solutes. 

Thus it might be wise to construct a simple graph of RI against iodixanol concentration, from measurements made on solutions of iodixanol prepared by mixing OptiPrep with the specific cell medium used in the study