Proteomic approach to human kidney glomerulus prepared by laser microdissection from frozen biopsy specimens: Exploration of proteome after removal of blood-derived proteins

Authors

  • Yutaka Yoshida,

    Corresponding author
    • Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Masaaki Nameta,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Masayoshi Kuwano,

    1. Laboratory of Proteome Research, National Institute of Biomedical Innovation, Japan
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  • Ying Zhang,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Xu Bo,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Sameh Magdeldin,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
    2. Department of Physiology, Faculty of Veterinary Medicine, Suez Canal University, Egypt
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  • Zenyui Cui,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Hidehiko Fujinaka,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
    2. Institute of Clinical Research, Niigata National Hospital, Japan
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  • Eishin Yaoita,

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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  • Takeshi Tomonaga,

    1. Laboratory of Proteome Research, National Institute of Biomedical Innovation, Japan
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  • Tadashi Yamamoto

    1. Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, Japan
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Correspondence Dr. Yutaka Yoshida, Department of Structural Pathology, Institute of Nephrology, Graduate School of Medical and Dental Sciences, Niigata University, 1–757 Asahimachi-dori, Chuo-ku, Niigata 951–8510, Japan

E-mail: yyoshi@med.niigata-u.ac.jp

Fax: +81-25-227-0768

Abstract

Purpose

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.

Experimental design

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 and clinical relevance

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.

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