The Degree of Labeling (DOL), sometimes called Degree of Substitution (DOS), is a particularly useful parameter for characterizing and optimizing bioconjugates, such as fluorophore-labeled proteins. It is expressed as a molar ratio in the form of label/protein. In general, conjugates with lower DOL values tend to have weaker fluorescence intensities. However, bioconjugates with very high DOL values (e.g. DOL > 6) may also experience reduced fluorescence. This often occurs as a result of self-quenching from the fluorophore. An accurate DOL value will allow a researcher to find the most optimal bioconjugation ratio for a given labeling as well as ensure reproducibility in conjugation results.

For antibodies, the optimal DOL usually falls between 2 to 10. A more precise value, however, will largely depend on the properties of the label and protein. This means that for many bioconjugations, the optimal DOL must be experimentally determined, often through several small-batch labelings. For AAT Bioquest's antibody labeling kits, the optimal DOL and reagent quantities have already been tested. For other bioconjugates, the tool below can be used to calculate the degree of labeling.