High sensitivity LC-MS profiling of antibody-drug conjugates with difluoroacetic acid ion pairing.

Reversed-phase liquid chromatography (RPLC) separations of proteins using optical detection generally use trifluoroacetic acid (TFA) because it is a strong, hydrophobic acid and a very effective ion-pairing agent for minimizing chromatographic secondary interactions. Conversely and in order to avoid ion suppression, analyses entailing mass spectrometry (MS) detection is often performed with a weaker ion-pairing modifier, like formic acid (FA), but resolution quality may be reduced. To gain both the chromatographic advantages of TFA and the enhanced MS sensitivity of FA, we explored the use of an alternative acid, difluoroacetic acid (DFA).