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revised on 19.07.2018, 08:23 and posted on 19.07.2018, 12:14by Andrey Krutilin, Stephanie Maier, Raphael Schuster, Sebastian Kruber, Marcel Kwiatkowski, Wesley D. Robertson, Dwayne R. J. Miller, Hartmut Schluter
The analysis of proteomes directly from tissues requires the proteins to be released from the cells and their compartments and solubilized, which usually is achieved by mechanical homogenization. It was recently shown, that sampling of tissues with the novel picosecond infrared laser (PIRL) offers higher yields of proteins with respect to the total amount and total number of individual proteins in comparison to mechanical homogenization. Furthermore, proteins obtained from tissues by homogenization with PIRL are significantly less enzymatically degraded, giving improved access to the original composition of proteoforms. The effective cold vaporization of tissue with PIRL is very soft, which is responsible for the phenomenon, that even enzymatic activities of proteins in the tissue aerosol are maintained. In contrast, the energy following irradiation of tissue with microsecond infrared laser (MIRL) pulses is not thermally and acoustically confined to the ablated volume. In this study, PIRL (1 J·cm-2) and MIRL (40-60 J·cm-2) were compared for sampling different tissue types for bottom-up proteomics. We showed that PIRL at low fluence is optimal for soft tissue and desired in scenarios were enzymatic activities of proteins must be maintained as well as were no residual tissue damage is a requirement. MIRL could be well suited for scenarios were enzymatic activities must be suppressed within the intact tissue and thermal and acoustic damage is not a concern.