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Metabolomics represents the global assessment of metabolites in a biological sample and reports the closest information to the phenotype of the biological system under study. Mammalian cell metabolomics has emerged as a promising tool with potential applications in many biotechnology and research areas. Metabolomics workflow includes experimental design, sampling, sample processing, metabolite analysis, and data processing. Given their influence on metabolite content and biological interpretation of data, a good experimental design and the appropriate choice of a sample processing method are prerequisites for success in any metabolomic study. The use of mammalian cells in the metabolomics field involves harder sample processing methods, including metabolism quenching and metabolite extraction, as compared to the use of body fluids, although such critical issues are frequently overlooked. This review aims to overview the common experimental procedures used in mammalian cell metabolomics based on mass spectrometry, by placing special emphasis on discussing sample preparation approaches, although other aspects, such as cell metabolomics applications, culture systems, cellular models, analytical platforms, and data analysis, are also briefly covered. This review intends to be a helpful tool to assist researchers in addressing decisions when planning a metabolomics study involving the use of mammalian cells.