Oral Presentation 24th Annual Lorne Proteomics Symposium 2019

Laser ablation for tissue sampling for proteomics and metabolomics (#52)

Hartmut Schlüter 1 , Pascal Steffen 2 , Nils-Owe Hansen 3 , R.J.Dwayne Miller 3 , Mark Molloy 2
  1. Mass Spectrometric Proteomics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
  2. Bowel Cancer & Biomarker Lab, Kolling Institute, The University of Sydney, St.Leonards, Sydney , New South Wales, Australia
  3. Atomically Resolved Dynamics Department, Max-Planck Institute for the Structure and Dynamics of Matter (MPSD), Hamburg, Germany

For every omics approach sample preparation is one of the most critical steps.  Especially tissue sampling is very challenging because during the process of homogenization biomolecules are known to be converted by chemical reactions like oxidation as well as enzymatic reactions. The latter are caused by the release of enzymes from their compartments during homogenization. We have recently shown that sampling of tissues for proteomics with a picosecond infrared laser (PIRL) is very soft. Labile molecules like glycoproteins stay intact during laser ablation. Even enzymatic activities of proteins are maintained (1).  Furthermore, the total yield of proteins with respect to their total number and total amount is significantly better compared to conventional homogenization. Proteoforms during tissue sampling with PIRL also remain intact in contrast to conventional homogenization (2). Thus, especially for the future of top-down proteomics PIRL is very promising (3). In addition to the application of PIRL for tissue sampling for proteomics, we performed a series of experiments applying infrared laser systems for tissue sampling for lipidomics and metabolomics. We sampled different pig brain areas with PIRL and investigated the condensates of the tissue aerosols with shot-gun lipidomics. By principle component analysis we were able to differentiate the different brain tissues. We compared the yields of metabolites obtained by conventional tissue sampling and by PIRL. The metabolites of both homogenization procedures were analyzed with p180-kit of Biocrates, by which approximately 180 metabolites are quantified via selected reaction monitoring by LC-MS and flow-injection analysis (FIA) by a triple quadrupole mass spectrometer. The results clearly showed that PIRL is more efficient in tissue sampling for proteomics. Furthermore, PIRL is giving access for volatile labile molecules by irradiation of tissues. In summary tissue sampling with PIRL for lipidomics, metabolomics and proteomics is advantageous compared to classical sampling and homogenization methods, giving higher yields of intact biomolecules and thereby allowing a closer and more realistic view to the original composition of biomolecules in intact tissues.

  1. Ultrafast extraction of proteins from tissues using desorption by impulsive vibrational excitation. Kwiatkowski M, et al., Miller RJ, Schlüter H. Angew Chem Int Ed Engl. 2015 Jan 2;54(1):285-8.
  2. 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. Kwiatkowski M, et al., Steffen P, et al., Miller RJD, Schlüter H. J Proteomics. 2016 Feb 16;134:193-202.
  3. How many human proteoforms are there? Aebersold R, et al. Kelleher NL, et al., Schlüter H, et al., Zhang B. Nature Chem Biol. 2018 Feb 14;14(3):206-214.