title: Proteomic approach to human kidney glomerulus prepared by laser microdissection from frozen biopsy specimens: exploration of proteome after removal of blood-derived proteins.
abstract: OBJECTIVE
  Abundance of blood-derived proteins in glomeruli prepared by laser microdissection from human kidney biopsy specimens has hampered in-depth proteomic analysis of glomeruli. We attempted to establish experimental platform for in-depth proteomic analysis of glomeruli by removal of blood-derived proteins from frozen biopsy samples.
  
  
  METHODS
  Frozen sections of biopsy samples were exposed to repeated PBS washes prior to laser microdissection to remove blood-derived proteins, and glomerular dissectants were analyzed by MS. The depth of proteomic analysis was evaluated by dynamic range of identified proteins and detection of low-abundance proteins.
  
  
  RESULTS
  Two times PBS washes of frozen sections effectively eliminated blood-derived proteins in laser-microdissected glomeruli and gave an increased number of identified proteins. Analysis of glomeruli from single specimens by a linear ion trap-Orbitrap mass analyzer generated nonredundant, high-confidence datasets of more than 400 identified proteins with high reproducibility, which attained to a considerable depth of the glomerulus proteome as revealed by a wide dynamic range and identification of low-abundance proteins.
  
  
  CONCLUSIONS
  Implementation of washing of frozen section with PBS successfully removed blood-derived proteins and resulted in an in-depth proteomic analysis of laser-microdissected glomeruli, suggesting applicability to clinical study.
authors.affiliation: Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan. yyoshi@med.niigata-u.ac.jp
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Yoshida
authors.forename: Masaaki
authors.initials: M
authors.lastname: Nameta
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Kuwano
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Xu
authors.initials: X
authors.lastname: Bo
authors.forename: Sameh
authors.initials: S
authors.lastname: Magdeldin
authors.forename: Zenyui
authors.initials: Z
authors.lastname: Cui
authors.forename: Hidehiko
authors.initials: H
authors.lastname: Fujinaka
authors.forename: Eishin
authors.initials: E
authors.lastname: Yaoita
authors.forename: Takeshi
authors.initials: T
authors.lastname: Tomonaga
authors.forename: Tadashi
authors.initials: T
authors.lastname: Yamamoto
mesh_terms: D000328:Adult; D000368:Aged; D001706:Biopsy; D001798:Blood Proteins / Q000302:isolation & purification*; D005260:Female; D005629:Frozen Sections / Q000379:methods*; D006801:Humans; D007678:Kidney Glomerulus / Q000378:metabolism* / Q000473:pathology*; D061346:Laser Capture Microdissection / Q000379:methods*; D008297:Male; D008875:Middle Aged; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*
chemical_list: D001798:Blood Proteins; D020543:Proteome