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

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  • Furukawa Suzuna
    Dept. Applied Biosci., Kanagawa Inst. Tech.
  • Kobayashi Mizuki
    Dept. Applied Biosci., Kanagawa Inst. Tech.
  • 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. Pharmacol., Juntendo Univ. Sch. Med.
  • Goshima Yoshio
    Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ.
  • Yamashita Naoya
    Dept. Applied Biosci., Kanagawa Inst. Tech. Dept. Mol Pharmacol and Neurobiol, Grad. Sch. Med., Yokohama City Univ. Dept. Pharmacol., Juntendo Univ. Sch. Med.

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  • 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. Both Sema3A monoclonal antibodies recognize the Sema domain of human Sema3A and can measure recombinant Sema3A in the range of 0-100 pM by ELISA. The specificity of this assay was confirmed by using the embryonic brains from sema3A deficient mice as a negative control. Moreover, this assay could measure Sema3A concentration in Tris buffered saline-and SDS-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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