Therapeutic monoclonal antibodies (mAbs) are rapidly taking over the treatment of many malignancies, and an astonishing number of mAbs is in development. This causes a high demand for quantification of mAbs in biomatrices both for measuring therapeutic mAb concentrations and to support pharmacokinetics and pharmacodynamics studies. Conventionally, ligand-binding assays are used for these purposes, but LC-MS is gaining popularity. Although intact (top-down) and subunit (middle-down) mAb quantification is reported, signature peptide (bottom-up) quantification is currently most advantageous. This review provides an overview of the reported bottom-up mAb quantification methods in biomatrices as well as general recommendations regarding signature peptide and internal standard selection, reagent use and optimization of digestion in bottom-up quantification methods.