Homogenization is a critical step in the analysis of proteomes of tissues. With many classical homogenization methods not all parts of the tissue are completely dispersed. As a result after centrifugation an insoluble pellet remains, containing proteins, which are lost for the analysis. A problem occurring in each kind of homogenization method is associated with release of enzymes changing the composition of the proteomes. The picosecond infrared laser (PIRL) is promising to improve the quality of tissue homogenates. During irradiation water molecules in the tissue absorb the energy of the laser. Because of the picosecond pulse the absorbed energy of the water molecules is nearly completely transformed into translational energy. Thus during irradiation with PIRL the tissue is ablated by cold vaporization.
In this study by applying diverse proteomic approaches PIRL homogenates were analyzed and compared with homogenates yielded by mechanical homogenization. Comparison of PIRL tissue homogenates with mechanical tissue homogenates by bottom-up proteomics revealed a significant higher yield of the number of identified proteins. Analysis of the different homogenates with two-dimensional electrophoresis showed a larger number of protein species in the case of the PIRL homogenization. The mechanical homogenate was characterized by a larger number of proteolytic degradation products. The lower degree of proteolytic degradation in the PIRL homogenate presumably can be explained by the ultrafast process of the conversion of intact tissue into the frozen condensate of the tissue aerosol.
In conclusion homogenization of tissues with PIRL is advantageous because the dispersion of tissues is more complete, the homogenate is more homogeneous, it is faster and very soft. Furthermore cold vaporization of tissue with PIRL is giving higher yields in the total amount of proteins, in the number of identified proteins and in the number of protein species (proteoforms).