title: Quantitative proteomic analysis of formalin-fixed, paraffin-embedded clear cell renal cell carcinoma tissue using stable isotopic dimethylation of primary amines.
abstract: BACKGROUND
  Formalin-fixed, paraffin-embedded (FFPE) tissues represent the most abundant resource of archived human specimens in pathology. Such tissue specimens are emerging as a highly valuable resource for translational proteomic studies. In quantitative proteomic analysis, reductive di-methylation of primary amines using stable isotopic formaldehyde variants is increasingly used due to its robustness and cost-effectiveness.
  
  
  RESULTS
  In the present study we show for the first time that isotopic amine dimethylation can be used in a straightforward manner for the quantitative proteomic analysis of FFPE specimens without interference from formalin employed in the FFPE process. Isotopic amine dimethylation of FFPE specimens showed equal labeling efficiency as for cryopreserved specimens. For both FFPE and cryopreserved specimens, differential labeling of identical samples yielded highly similar ratio distributions within the expected range for dimethyl labeling. In an initial application, we profiled proteome changes in clear cell renal cell carcinoma (ccRCC) FFPE tissue specimens compared to adjacent non-malignant renal tissue. Our findings highlight increased levels of glyocolytic enzymes, annexins as well as ribosomal and proteasomal proteins.
  
  
  CONCLUSIONS
  Our study establishes isotopic amine dimethylation as a versatile tool for quantitative proteomic analysis of FFPE specimens and underlines proteome alterations in ccRCC.
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, Freiburg, Germany. JWeisser@cemm.oeaw.ac.at.|Present address: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, A-1090, Vienna, Austria. JWeisser@cemm.oeaw.ac.at.
authors.forename: J
authors.initials: J
authors.lastname: Weißer
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, Freiburg, Germany. zon.lai@mol-med.uni-freiburg.de.
authors.forename: Z W
authors.initials: ZW
authors.lastname: Lai
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. peter.bronsert@uniklinik-freiburg.de.|Comprehensive Cancer Center Freiburg, Freiburg, Germany. peter.bronsert@uniklinik-freiburg.de.|German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany. peter.bronsert@uniklinik-freiburg.de.
authors.forename: P
authors.initials: P
authors.lastname: Bronsert
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. markus.kuehs@uniklinik-freiburg.de.
authors.forename: M
authors.initials: M
authors.lastname: Kuehs
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. vanessa.drendel@uniklinik-freiburg.de.
authors.forename: V
authors.initials: V
authors.lastname: Drendel
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. sylvia.timme@uniklinik-freiburg.de.
authors.forename: S
authors.initials: S
authors.lastname: Timme
authors.affiliation: Clinic for General and Visceral Surgery, University Medical Center Freiburg, Freiburg, Germany. simon.kuesters@uniklinik-freiburg.de.
authors.forename: S
authors.initials: S
authors.lastname: Kuesters
authors.affiliation: Urologische Klinik und Zentrale Klinische Forschung, Klinikum der Universität Freiburg, Freiburg, 79106, Germany. cordula.jilg@uniklinik-freiburg.de.
authors.forename: C A
authors.initials: CA
authors.lastname: Jilg
authors.affiliation: Clinic for General and Visceral Surgery, University Medical Center Freiburg, Freiburg, Germany. dr.ulrich.wellner@gmail.com.|Present address: Clinic for Surgery, University Clinic of Schleswig-Holstein Campus Lübeck, Lübeck, Germany. dr.ulrich.wellner@gmail.com.
authors.forename: U F
authors.initials: UF
authors.lastname: Wellner
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. silke.lassmann@uniklinik-freiburg.de.|BIOSS Centre for Biological Signaling Studies, University of Freiburg, D-79104, Freiburg, Germany. silke.lassmann@uniklinik-freiburg.de.|Comprehensive Cancer Center Freiburg, Freiburg, Germany. silke.lassmann@uniklinik-freiburg.de.|German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany. silke.lassmann@uniklinik-freiburg.de.
authors.forename: S
authors.initials: S
authors.lastname: Lassmann
authors.affiliation: Department of Pathology, University Medical Center Freiburg, Freiburg, Germany. martin.werner@uniklinik-freiburg.de.|Comprehensive Cancer Center Freiburg, Freiburg, Germany. martin.werner@uniklinik-freiburg.de.|German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany. martin.werner@uniklinik-freiburg.de.
authors.forename: M
authors.initials: M
authors.lastname: Werner
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, Freiburg, Germany. martin.biniossek@mol-med.uni-freiburg.de.
authors.forename: M L
authors.initials: ML
authors.lastname: Biniossek
authors.affiliation: Institute of Molecular Medicine and Cell Research, University of Freiburg, Freiburg, Germany. oliver.schilling@mol-med.uni-freiburg.de.|BIOSS Centre for Biological Signaling Studies, University of Freiburg, D-79104, Freiburg, Germany. oliver.schilling@mol-med.uni-freiburg.de.|German Cancer Consortium (DKTK) and German Cancer Research Center (DKFZ), Heidelberg, Germany. oliver.schilling@mol-med.uni-freiburg.de.
authors.forename: O
authors.initials: O
authors.lastname: Schilling
mesh_terms: D000588:Amines / Q000737:chemistry*; D002247:Carbon Isotopes / Q000737:chemistry; D002292:Carcinoma, Renal Cell / Q000378:metabolism / Q000473:pathology*; D002851:Chromatography, High Pressure Liquid; D005557:Formaldehyde / Q000737:chemistry; D006801:Humans; D007553:Isotope Labeling; D007680:Kidney Neoplasms / Q000378:metabolism / Q000473:pathology*; D016612:Paraffin Embedding; D020543:Proteome / Q000032:analysis*; D040901:Proteomics; D053719:Tandem Mass Spectrometry
chemical_list: D000588:Amines; D002247:Carbon Isotopes; D020543:Proteome; D005557:Formaldehyde