Production and Characterization of Monoclonal Antibodies Which Distinguish Different Surface Structues of Pea (Pisum sativum cv. Alaska) Phytochrome :

  • Nagatani,Akira
    Department of Biology, Faculty of Science, University of Tokyo:Division of Biological Regulation, National Institute for Basic Biology
  • Yamamoto,Kotaro T.
    Division of Biological Regulation, National Institute for Basic Biology
  • Furuya,Masaki
    Department of Biology, Faculty of Science, University of Tokyo:Division of Biological Regulation, National Institute for Basic Biology
  • Fukumoto,Tetsuo
    Department of Anatomy, Hamamatsu University School of Medicine
  • Yamashita,Akira
    Department of Anatomy, Hamamatsu University School of Medicine

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Eight monoclonal antibodies to pea 114,000 dalton phytochrome (mAP-1 to mAP-8) were produced by fusing spleen cells from immunized BALB/C mice with NS-1 myeloma cells. Antibody production by mAP-1 to mAP-7 was detected by cellular radioimmunoassay using sheep red blood cells coated with pea phytochrome. Relative binding positions of these mAPs were determined using a competitive binding assay, and at least 6 different antigenic regions were defined on the 114,000 dalton chromopeptide. Location of the regions in relation to the chromophore was studied by examining the cross-reactions of the relevant mAPs with chromophore-containing proteolytic fragments (62,000, 37,000 and 24,000 daltons) of phytochrome. The following cross-reactions were observed; mAP-1 with 62,000, mAP-6 and mAP-7 with 62,000, 37,000 and 24,000. Only mAP-7 caused "spectral denaturation" of phytochrome. All mAPs reacted with P_R and P_<FR> with similar af Bnity. These mAPs will be useful probes in future studies for detecting each site of the apoproteins of phytochrome. By using another screening method, namely radioimmunoassay with antigen-coupled Sepharose 4B, mAP-8 was also obtained.

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