Homogenization of tissues via picosecond-infrared laser (PIRL) ablation: Giving a closer view on the in-vivo composition of protein species as compared to mechanical homogenization

Posttranslational modifications and proteolytic processing regulate almost all physiological processes. Dysregu-lation can potentially result in pathologic protein species causing diseases. Thus, tissue species proteomes of dis-eased individuals provide diagnostic information. Since the composition of tissue proteomes can rapidly changeduring tissue homogenization by the action of enzymes released from their compartments, disease specific protein species patterns can vanish.Recently, we described a novel, ultrafastand soft method for cold vaporization of tissue via desorption by impulsive vibrational excitation (DIVE) using a picosecond-infrared-laser (PIRL). Given that DIVE extraction may provide improved access to the original composition of protein species in tissues, we compared the proteome composition of tissue protein homogenates after DIVE homogenization with conventional homogenizations. A higher number of intact protein species was observed in DIVE homogenates. Due tothe ultrafast transfer of proteins from tissues via gas phase into frozen condensates of the aerosols, intact proteinspecies were exposed to a lesser extent to enzymatic degradation reactions compared with conventional proteinextraction. In addition, total yield of the number of proteins is higher in DIVE homogenates, because they are veryhomogenous and contain almost no insoluble particles, allowing direct analysis with subsequent analytical methods without the necessity of centrifugation. Biological significance: Enzymatic protein modifications during tissue homogenization are responsible for changes of the in-vivo protein speciescomposition. Cold vaporization of tissues byPIRL-DIVE is comparablewith taking a snapshot at the time of the laser irradiation of the dynamic changes that occur continuously under in-vivo conditions. At that time point all biomolecules are transferred into an aerosol, which is immediately frozen