tDespite many advances in the generation of high producing recombinant mammalian cell lines overthe last few decades, cell line selection and development is often slowed by the inability to predict acell line’s phenotypic characteristics (e.g. growth or recombinant protein productivity) at larger scale(large volume bioreactors) using data from early cell line construction at small culture scale. Here wedescribe the development of an intact cell MALDI-ToF mass spectrometry fingerprinting method formammalian cells early in the cell line construction process whereby the resulting mass spectrometrydata are used to predict the phenotype of mammalian cell lines at larger culture scale using a PartialLeast Squares Discriminant Analysis (PLS-DA) model. Using MALDI-ToF mass spectrometry, a library ofmass spectrometry fingerprints was generated for individual cell lines at the 96 deep well plate stage ofcell line development. The growth and productivity of these cell lines were evaluated in a 10 L bioreactormodel of Lonza’s large-scale (up to 20,000 L) fed-batch cell culture processes. Using the mass spectrometryinformation at the 96 deep well plate stage and phenotype information at the 10 L bioreactor scale a PLS-DA model was developed to predict the productivity of unknown cell lines at the 10 L scale based upontheir MALDI-ToF fingerprint at the 96 deep well plate scale. This approach provides the basis for the veryearly prediction of cell lines’ performance in cGMP manufacturing-scale bioreactors and the foundationfor methods and models for predicting other mammalian cell phenotypes from rapid, intact-cell massspectrometry based measurements.