Construction of the ELISA assay to quantify Semaphorin 3A in the adult brain

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  • Yamashita Naoya
    Dept. of Applied Biosci., Kanagawa Institute of Technology Dept. of Cell and Mol Pharmacol., Juntendo Univ., Sch. Med. Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ.
  • Mizutani Yui
    Dept. of Cell and Mol Pharmacol., Juntendo Univ., Sch. Med.
  • Jitsuki-Takahashi Aoi
    Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ. Dept. Biochem, Tokyo Women's Medical Univ
  • Lin Ke-Yi
    Chiome Bioscience
  • Nakamura Fumio
    Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ. Dept. Biochem, Tokyo Women's Medical Univ
  • Sakurai Takashi
    Dept. of Cell and Mol Pharmacol., Juntendo Univ., Sch. Med.
  • Goshima Yoshio
    Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ.

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Other Title
  • 成体脳におけるセマフォリン3A発現を定量するELISAシステムの確立

Abstract

<p>Extracellular soluble signals that control several aspects of neuronal development are known to play a critical role in maintaining neuronal function and homeostasis in the mature nervous system. Abnormal expression and/or secretion of these molecules are therefore thought to be associated with the onset of various types of neurological disorders. It has been reported that the expression of Semaphorin 3A (Sema3A), a secreted type of repulsive axon guidance molecule, is impaired in several neurodegenerative disorders. However, due to the lack of a reliable Sema3A antibody, our knowledge about Sema3A expression in the adult brain is still limited. Here we report the identification of a pair of Sema3A monoclonal antibodies for the sandwich ELISA assay using the Autonomously Diversifying Library system. This ELISA assay recognized recombinant human Sema3A in the range of 0-100 pM, but not Sema3F, one of a subfamily of class III semaphorins. The specificity of this assay was also confirmed by using the embryonic brains from sema3A deficient mice as a negative control. Moreover, this assay could measure Sema3A concentration in TBS- and RIPA-soluble lysate obtained from the adult mice brains. These data suggested that our ELISA assay is a reliable tool for the validation of Sema3A as a biomarker in neurodegenerative disorders.</p>

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