著者

Izumi Morita Hiroyuki Oyama Mayumi Yasuo Kazuhisa Matsuda Kengo Katagi Aya Ito Hiroka Tatsuda Hiroyuki Tanaka Satoshi Morimoto Norihiro Kobayashi

出版者

The Pharmaceutical Society of Japan

雑誌

Biological and Pharmaceutical Bulletin (ISSN:09186158)

巻号頁・発行日

vol.40, no.2, pp.174-181, 2017-02-01 (Released:2017-02-01)

参考文献数

40

被引用文献数

10 17

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Law enforcement against illicit use of cannabis and related substances requires rapid, feasible, and reliable tools for on-site testing of cannabinoids. Notably, methods based on cannabinoid-specific antibodies enable efficient screening of multiple specimens. Antibody engineering may accelerate development of modern and robust testing systems. Here, we used in vitro affinity maturation to generate a single-chain Fv fragment (scFv) that recognizes with high affinity the psychoactive cannabinoid, Δ9-tetrahydrocannabinol (THC). A mouse monoclonal antibody against THC, Ab-THC#33, with Ka 6.2×107 M−1 (as Fab fragment) was established by the hybridoma technique. Then, a “wild-type” scFv (wt-scFv) with Ka, 1.1×107 M−1 was prepared by bacterial expression of a fusion gene combining the VH and VL genes for Ab-THC#33. Subsequently, random point mutations in VH and VL were generated separately, and the resulting products were assembled into mutant scFv genes, which were then phage-displayed. Repeated panning identified a mutant scFv (scFv#m1-36) with 10-fold enhanced affinity (Ka 1.1×108 M−1) for THC, in which only a single conservative substitution (Ser50Thr) was present at the N-terminus of the VH-complementarity-determining region 2 (CDR2) sequence. In competitive enzyme-linked immunosorbent assay (ELISA), the mutant scFv generated dose–response curves with midpoint 0.27 ng/assay THC, which was 3-fold lower than that of wt-scFv. Even higher reactivity with a major THC metabolite, 11-nor-9-carboxy-Δ9-tetrahydrocannabinol, indicated that the mutant scFv will be useful for testing not only THC in confiscated materials, but also the metabolite in urine. Indeed, the antibody fragment is potentially suitable for use in advanced on-site testing platforms for cannabinoids.

著者

Gerard B. Remijn Emi Hasuo Haruna Fujihira Satoshi Morimoto

出版者

一般社団法人 日本音響学会

雑誌

Acoustical Science and Technology (ISSN:13463969)

巻号頁・発行日

vol.35, no.5, pp.229-242, 2014-05-01 (Released:2014-09-01)

参考文献数

109

被引用文献数

11

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In 1939, Pauline Davis reported the first study on event-related potentials (ERPs) performed on awake humans. ERPs are time-locked brain potentials that occur in response to cognitive, motor or perceptual events. The events used by Davis were sounds, and in the decades that followed her landmark study ERP research significantly contributed to the knowledge of auditory perception and neurophysiology we have today. ERPs are very well suited to study neural responses to sound stimuli, since the researcher can monitor the brain's registration of sound edges and spectral changes in sound on a millisecond-by-millisecond basis. In this overview we will introduce basic concepts of auditory ERP research. The overview includes descriptions of typical ERP components, experimental paradigms, sound stimuli, research methodology, and ways to analyze data.