Rational cell culture optimization enhances experimental reproducibility in cancer cells

Marina Wright Muelas*, Fernando Ortega, Rainer Breitling, Claus Bendtsen, Hans V. Westerhoff

*Corresponding author for this work

Research output: Contribution to JournalArticleAcademicpeer-review


Optimization of experimental conditions is critical in ensuring robust experimental reproducibility. Through detailed metabolomic analysis we found that cell culture conditions significantly impacted on glutaminase (GLS1) sensitivity resulting in variable sensitivity and irreproducibility in data. Baseline metabolite profiling highlighted that untreated cells underwent significant changes in metabolic status. Both the extracellular levels of glutamine and lactate and the intracellular levels of multiple metabolites changed drastically during the assay. We show that these changes compromise the robustness of the assay and make it difficult to reproduce. We discuss the implications of the cells' metabolic environment when studying the effects of perturbations to cell function by any type of inhibitor. We then devised 'metabolically rationalized standard' assay conditions, in which glutaminase-1 inhibition reduced glutamine metabolism differently in both cell lines assayed, and decreased the proliferation of one of them. The adoption of optimized conditions such as the ones described here should lead to an improvement in reproducibility and help eliminate false negatives as well as false positives in these assays.

Original languageEnglish
Article number3029
Pages (from-to)1-16
Number of pages16
JournalScientific Reports
Issue number1
Publication statusPublished - 14 Feb 2018


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