<p>Accurate values of the specific refractive index increment (∂<i>n</i>/∂<i>c</i>) are essential to the accurate determination of molar mass averages, distributions, and related macromolecular parameters by, among others, size-based separations with online static light scattering and differential refractive index (DRI) detection. Examined here is the 100% mass recovery method of calculating the ∂<i>n</i>/∂<i>c</i> of dilute macromolecular solutions, when employing mixed solvents in a size-based separation (e.g., size-exclusion chromatography or SEC) with DRI detection. This method has been used successfully in the past for determining the ∂<i>n</i>/∂<i>c</i> of various polyelectrolytes. It is quicker, generally simpler, and less sample-intensive than its offline, batch-mode DRI counterpart. Whether or not the 100% mass recovery method allows for the necessary solvent equilibration within the immediate vicinity of the polymer chain during a chromatographic run, so as to allow for accurate determination of ∂<i>n</i>/∂<i>c</i> in SEC/DRI experiments, is evaluated here. This is done using a set of three narrow-dispersity linear polystyrene standards covering a 40-fold range in molar mass, dissolved in a 25:75 mix of tetrahydrofuran and <i>N</i>,<i>N-</i>dimethyl formamide. Results are compared to those previously obtained by the batch-mode method and from calculations involving the accurately known molar masses of the polymers and refractive indices of the solvents. The 100% mass recovery method of obtaining ∂<i>n</i>/∂<i>c</i> values, while of great help for determining the ∂<i>n</i>/∂<i>c</i> of, e.g., polyelectrolytes, does not appear able to overcome the obstacle of preferential solvation when analyzing macromolecules in a mix of non-isorefractive solvents with dissimilar second virial coefficients.</p>