LC-MS is a powerful protein characterization tool that can be used to identify post-translational modifications (PTMs) on proteins during the clone selection process. An early read on product quality during clone selection can help avoid CMC pitfalls that can result in costly delays. PTMs can be sequence-dependent and by identifying these PTMs during the sequence optimization stage, enables the selection of constructs with more desirable product quality attributes. Four examples will be shown that highlight the use of mass spectrometry to confirm the presence of expected PTMs such as N-glycosylation of the Fc region in mAb-like molecules, as well as to screen out undesirable PTMs such as proline hydroxylation. These case studies demonstrate that in-depth characterization using mass spectrometry early in the development process can mitigate potential CMC risks and enable the product development and IND timelines to stay on track. 1. Identification of N-terminal pyroglutamate formation – an expected PTM 2. Cleavage site identification to minimize cleavages and increase protein yield 3. Proline hydroxylation PTM – an undesirable PTM 4. O-glycan PTM identification — selecting constructs with low levels of O-glycosylation for reducing manufacturing risk