Application of antibody engineering in forensic toxicological analysis

• Project/Area Number: 24590859
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Legal medicine
• Research Institution: Kumamoto University
• Principal Investigator: SASAO Ako 熊本大学, 生命科学研究部, 助教 (80284751)
• Co-Investigator(Renkei-kenkyūsha): MORIOKA Hiroshi 熊本大学, 大学院生命科学研究部, 教授 (20230097) ; NISHITANI Yoko 熊本大学, 大学院生命科学研究部, 教授 (30359997) ; YONEMITSU Kosei 熊本大学, 大学院生命科学研究部, 准教授 (10128332)
• Project Period (FY): 2012-04-01 – 2015-03-31
• Project Status: Completed (Fiscal Year 2014)
• Budget Amount: ¥5,330,000 (Direct Cost: ¥4,100,000、Indirect Cost: ¥1,230,000); Fiscal Year 2014: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2013: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2012: ¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
• Keywords: 薬毒物スクリーニング / 法医中毒学 / 組換え抗体 / 一本鎖抗体 / フルボキサミン / 向精神薬 / 法中毒

Outline of Final Research Achievements

The aim of this study is to facilitate preparation of anti-drug antibodies　using antibody engineering such as phage display technology. In this study, we acquired cDNAs that encode immunoglobulin variable regions (VL and VH) from hybridoma cells producing anti-drug monoclonal antibodies (antidepressant, fluvoxamine; FLV, antipsychotic, phenothiazines; PTZ), and determined their DNA sequences. Then, we constructed expression vectors and prepared single chain variable fragment (scFv) recognizing FLV or PTZ. Their physicochemical properties were examined by ELISA or SPR analysis. Anti- FLV or anti-PTZ scFvs were found to have specific and sufficient binding to their each antigen. Also, we prepared an FLV Quenchbody protein that was a novel antibody-based fluorescent biosensor and examined its physicochemical properties.

Research Products

• [Journal Article] Single-chain variable fragment technology in forensic toxicological analysis: production of an antibody to fluvoxamine (2013)
  • Author(s): Ako Sasao・ Yoshiaki Suwa・ Taemi Aso・ Hiroe Kohmatsu・ Yuki Ohtsu・ Satoko Mishima・ Kosei Yonemitsu・ Hiroshi Morioka・ Yoko Nishitani
  • Journal Title: Forensic Toxicology Volume: 31 Issue: 1 Pages: 62-66
  • DOI: 10.1007/s11419-012-0163-4
  • Peer Reviewed
• [Presentation] 抗うつ薬フルボキサミンに対する一本鎖抗体の調製とその特性 (2015)
  • Author(s): 笹尾亜子、高木美智代、大津由紀、三島聡子、米満孝聖、森岡弘志、西谷陽子
  • Organizer: 第99次日本法医学会学術全国集会
  • Place of Presentation: 高知市、高知県
  • Year and Date: 2015-06-10 – 2015-06-12
• [Presentation] Application of antibody engineering to drug screening: Preparation of the anti-fluvoxamine Quenchbody, a novel fluorescent biosensor (2014)
  • Author(s): Sasao A, Takaki M, Jeong HJ, Ohtsu Y, Mishima S, Yonemitsu K, Morioka H, Ueda H, Nishitani Y
  • Organizer: 9th International Symposium on Advances in Legal Medicine and 98th Congress of the Japanese Society of Legal Medicine
  • Place of Presentation: Fukuoka, Japan
  • Year and Date: 2014-06-16 – 2016-06-20
• [Presentation] Application of antibody engineering to drug screening: Preparation of the anti-fluvoxamine Quenchbody, a novel fluorescent biosensor (2014)
  • Author(s): Ako Sasao, Michiyo Takaki, Hee-Jin Jeong, Yuki Ohtsu, Satoko Mishima, Kosei Yonemitsu, Hiroshi Morioka, Hiroshi Ueda, Yoko Nishitani
  • Organizer: 第9回国際法医学シンポジウム・第98次日本法医学会学術全国集会
  • Place of Presentation: 福岡市・福岡国際会議場
• [Presentation] Application of recombinant antibody engineering to drug screening. Report 1: preparation of anti-fluvoxamine single chain variable fragment (scFv) antibody. (2012)
  • Author(s): Sasao A, Suwa Y, Ohtsu Y, Mishima S, Yonemitsu K, Morita H, Nishitani Y.
  • Organizer: The 50th Annual Meeting of the International Association of Forensic Toxicologists（TIAFT2012）
  • Place of Presentation: アクトシティ浜松（静岡県）
• [Presentation] 抗うつ薬フルボキサミンに対する一本鎖抗体の調製 (2012)
  • Author(s): 笹尾亜子，諏訪喜昭，大津由紀，三島聡子，米満孝聖，森岡弘志，西谷陽子
  • Organizer: 第96次日本法医学会学術全国集会
  • Place of Presentation: アクトシティ浜松（静岡県）