Enzymatic Dissociation Induces Transcriptional and Proteotype Bias in Brain Cell Populations
Daniele Mattei, Andranik Ivanov, Marc van Oostrum, Stanislav Pantelyushin, Juliet Richetto, Flavia S. Mueller, Michał Beffinger, Linda Schellhammer, Johannes vom Berg, Bernd Wollscheid, Dieter Beule, Rosa Chiara Paolicelli, Urs Meyer
Abstract
Different cell isolation techniques exist for transcriptomic and proteotype profiling of brain cells. Here, we provide a systematic investigation of the influence of different cell isolation protocols on transcriptional and proteotype profiles in mouse brain tissue by taking into account single-cell transcriptomics of brain cells, proteotypes of microglia and astrocytes, and flow cytometric analysis of microglia. We show that standard enzymatic digestion of brain tissue at 37 °C induces profound and consistent alterations in the transcriptome and proteotype of neuronal and glial cells, as compared to an optimized mechanical dissociation protocol at 4 °C. These findings emphasize the risk of introducing technical biases and biological artifacts when implementing enzymatic digestion-based isolation methods for brain cell analyses.