Relevance to Theme Selected “Big data” approaches, such as multi-omics-based characterisation of cellular function, show great potential in aiding academic and industrial endeavours to improve therapeutics production. Impact/Novelty The utilisation of state-of-the-art host cell-omics to supplement conventional, medium-based measurements as proxies for cellular function will further the understanding of producer CHO cell lines. Introduction To meet the growing demand for recombinant biotherapeutics employed today, producer cell lines are under ever-constant development to improve their productivity, stability and product quality. Accurate analysis of protein expression and post-translational modification of proteins by reversible phosphorylation can effectively decipher vital cellular processes and their regulation. Methods/Approach The development of a novel, fast, and representative multi-omics sampling approach for suspension cultures allowing analysis of the host-cell proteome, phosphoproteome, metabolome, and secretome was carried out. To demonstrate the application of this approach, a fed-batch culture of the openly available producer cell line NISTCHO was consequently investigated for the impact of feeding on cellular function. Sampling took place in both the exponential and stationary growth phases to reflect the most critical stages of the bioprocess. Results were integrated into the parallel monitoring of cell proliferation and viability, as well as nutrient consumption and metabolic excrements. Results Analysis of bioprocess parameters and host cell-omic datasets of multiple time points before and after scheduled feeding of NISTCHO revealed distinct responses to altered nutrient availability. This work provides time-resolved and highly comprehensive profiling of the acute impact of feeding on the cellular landscape, including the proteome and phosphoproteome. Conclusion A novel mass spectrometry-based multi-omics method can offer an in-depth analysis of cellular function in various product and/or process-relevant biological settings. The advanced characterisation of an academically available producer cell line will drive research in recombinant protein production and bioprocess engineering.