Establishment of the anti-klotho monoclonal antibodies and detection of klotho protein in kidneys

Yukinari Kato, Emi Arakawa, Satoko Kinoshita, Akio Shirai, Akiko Furuya, Kazuya Yamano, Kazuyasu Nakamura, Akihiro Iida, Hideharu Anazawa, Noritoshi Koh, Akiko Iwano, Akihiro Imura, Toshihiko Fujimori, Makoto Kuro-O, Nobuo Hanai, Kazuhiko Takeshige, Yo Ichi Nabeshima

Research output: Contribution to journalArticle

112 Scopus citations

Abstract

A novel gene, klotho (kl), which is involved in the development of a syndrome resembling human aging in mice, was recently identified. The kl gene encodes a single-pass membrane protein whose extracellular domain carries homology to b-glucosidases. There also exists a splice variant of kl mRNA which encodes a putative secreted protein in both human and mouse. In this study, to characterize the physiological roles of Klotho protein, we established three monoclonal antibodies (mAbs) against the recombinant human Klotho protein. The mAbs are named KM2076 (rat IgG2a), KM2119 (rat IgG2b), and KM2365 (mouse IgG1). In Western blots, KM2076 and KM2119 specifically recognized a 130 kDa Klotho protein in the mouse and human kidney membrane fractions. To detect the human Klotho protein, the sandwich-type ELISA system with KM2076 and KM2365 was established. Using the ELISA system, we detected the human Klotho protein as low as 20 ng/ml in the supernatant of Chinese hamster ovary cells (CHO cells), introduced the human klotho gene. KM2076 and KM2119 specifically gave a positive staining by immunohistochemical staining in paraffin or frozen sections of the kidneys from wild-type mice but not in those from kl mice. Strong staining was observed especially in cortical renal tubules of the mouse kidney, where expression of klotho transcripts overlaps. KM2076 also showed a similar reaction pattern in the paraffin sections of rat and human kidneys. The mAbs established in this paper will serve as useful analytical, pathological, and diagnostic tools to disclose the role of Klotho protein in the suppression of a syndrome resembling human aging. (C) 2000 Academic Press.

Original languageEnglish (US)
Pages (from-to)597-602
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume267
Issue number2
DOIs
StatePublished - Jan 19 2000

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Kato, Y., Arakawa, E., Kinoshita, S., Shirai, A., Furuya, A., Yamano, K., Nakamura, K., Iida, A., Anazawa, H., Koh, N., Iwano, A., Imura, A., Fujimori, T., Kuro-O, M., Hanai, N., Takeshige, K., & Nabeshima, Y. I. (2000). Establishment of the anti-klotho monoclonal antibodies and detection of klotho protein in kidneys. Biochemical and Biophysical Research Communications, 267(2), 597-602. https://doi.org/10.1006/bbrc.1999.2009