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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. 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. 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. 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. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Citation

Yutaka Yoshida, Masaaki Nameta, Masayoshi Kuwano, Ying Zhang, Xu Bo, Sameh Magdeldin, Zenyui Cui, Hidehiko Fujinaka, Eishin Yaoita, Takeshi Tomonaga, Tadashi Yamamoto. Proteomic approach to human kidney glomerulus prepared by laser microdissection from frozen biopsy specimens: exploration of proteome after removal of blood-derived proteins. Proteomics. Clinical applications. 2012 Aug;6(7-8):412-7

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PMID: 22927354

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