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1 Key words: Varicella-zoster virus, monoclonal antibody, diagnosis, varicella, herpes zoster
2
3 蛍 光標 識 単 抗 体 に よ るVZV感 し た も の に つ い て の み 判 定 し,細 よ び 陰 性 の 標 準 試 料 と し て,VZV感 性お 染 色 は一 般 に 鮮 明 で あ った.陽 性 細 胞 の特 徴 は, お よ び400倍 で 鏡 検 し た. る い はHSV-2と 計43株 鮮 分 離 株 を 選 び,7つ そ れ ぞ れ6株 つ つ に つ い て,内 の交 のHSVお よ の 各 研 究 機 関 で, 容 を 知 らせ ず に ウ イ ル ス 株 の 同 定 を 行 っ た. 標 識 し たVZV,HSV-1お 使 用 し,あ お よ び細 胞 質 に広 汎 に特 異 蛍 光 が み られ る(原 倍 率 66).写 真2も 水 痘 例 のVZV抗 原 の染 色 を 示 し,核 内 と細 胞 質 に 明 瞭 な特 異 蛍 光 が み られ る. 矢 印( )は 多 核 巨細 胞 を 示 す(原 倍 率 132). 暗 赤 色 に染 色 され,黄 緑 色 の 特 異 蛍 光 を 示 す 陽 性 ウ イ ル ス の 分 離 培 養 は,水 疱 内 容 を,VZVで はVero細 細 胞 に 接 種 し て 行 っ た.ウ 水痘例 の水疱 原 を 示 し,核 内 陰 性 細 胞 は エ バ ソ ス ブル ーを 添 加 す る こ と に よ り イ ル ス分 離 培 養 胞 に,HSVで の染 色 の あ る こ とで あ る.写 真1は 蓋 塗 抹 標 本 で 検 出 され たVZV抗 の交差反応性 差 反 応 性 を 検 討 す る た め,合 HEL細 ッ トに よ る染 色 所 見 細 胞 質 全 体 が染 ま る こ と と,核 に特 異 的 な 顆 粒 状 ッ トのHSV-1あ (5)ウ (1)キ 績 胞 を 塗 抹 した ス ラ イ ドグ ラス を 同 時 (4)VZVとHSV間 びVZV新 成 染 お よび 非 感 に 観 察 し た.通 常 は200倍 VZVキ 197 胞 質 と核 に 穎 粒 状 の 特 異 蛍 光 を 認 め た も の を 陽 性 と し た.陽 染 のHEL細 染 症 の診 断 胞 あ る い はHEL イ ル ス の 同 定 はFITC よ びHSV-2のmAbを る い は 中 和 法 な ど で 行 っ た. は 細 胞 か ら鮮 明 に 区 別 で き た.た だ し,膿 疱 よ り得 られ る試 料 の 中 に は 時 に 白血 球 の 細 胞 質 中 に非 特 異 的 な 淡 いdullgreenの が あ った.し 染 色 像 が み られ る こ と か し,核 の 特 異 的 な 顆 粒 状 の 蛍 光 染 色 所 見 を 目安 にす る こ と に よ り,判 定 す る こ とが で ぎた. (2)患 者 検 体 で の抗 原 検 出 写 真1水 痘 検 体 塗抹 標 本 対 象 症 例 数 は431例,そ イル ス分 離 を 行 った.臨 帯 状 疱 疹190例,単 の 内258例 に つ い て は ウ 床 診 断 で は,水 痘105例, 純 ヘ ル ペ ス96例,帯 状 疱 疹,単 純 ヘ ル ペ ス の 区 別 のつ か な か った もの24例,そ 他 の 水 疱7例 で あ った.角 関 連 の症 例 と して 帯 状 疱 疹 が4例,単 Table 1 materials Detection of VZV using antigens 純 ヘル ペス in clinical monoclonal antibody Percentage of positive by immunofluorescence 写真2水 痘 検 体 塗抹 標 本 (): 平成2年2月20日 の 膜 ヘ ル ペ ス お よ び眼 科 cases
4 Table 2 Virological diagnosis by immuno- Table 3 Comparison between virus isolation and immunofluorescence
5 Table 4 The detailed analysis of the 80 cases which were negative both in VZV and HSV isolation
6 5) Goldstein, L. C., Corey, L., McDougall, J. K., Tolentino, E. & Nowinski, R. C.: Monoclonal antibodies to herpes simplex viruses: Use in antigenic typing and rapid diagnosis. J. Infect. Dis., 147: , ) Nowinski, R. C., Tam, M. R., Goldstein, L. C., Stong, L., Kuo, C. C., Corey, L., Stamm, W. E., Handfield, H. H., Knapp, J. S. & Holmes, K. K.: Monoclonal antibodies for diagnosis of infectious diseases in humans. Science, 219: 637 7) Kitamura, K., Namazue, J., Campo-Vera, H., Ogino, T. & Yamanishi, K.: Induction of neutralizing antibody against varicella-zoster virus (VZV) by VZV gp3 and crossreactivity between VZV gp3 and herpes simplex viruses gb. Virology, 149: 74-82, ) Schaeffer, H. J., Beauchamp, L. & de Miranda, P.: 9-(2-Hydroxyethoxy methyl) guanine activity against viruses of the herpes group. Nature, 272: , ) Biron, K. K. & Elion, G. B.: In vitro susceptibility of varicella zoster virus to Acyclovir. Antimicrob. Agents. Chemother., 18: 443 Diagnosis of Varicella-Zoster Virus (VZV) Infection by using FITC-labeled Monoclonal Antibodies Michihito NIIMURA & Mariko HONDA of Dermatology, Jikeikai Univerity School of Medicine Tsuyoshi KURATA & Tetsutaro SATA of Pathology, The National Institute of Health Hiroshi SAKAOKA of Oral Bacteriology, School Dentistry Hokkaido University
7 Rinji KAWANA of Bacteriology, Iwate Medical University Takashi KAWANA of Obstetrics and Gynecolog, School Medicine, The University of Tokyo Akira HIDANO of Dermatology, Tokyo Women Medicine College Kunihiko TAMAKI of Dermatology, School Medicine, The University of Tokyo Ryo HONDO of Pathology, The Institute of Medical Science, The University of Tokyo Ryoichi MURAKI Dermatology, The Salvation Army Booth Memorial Hospital Yoshizo ASANO Pediatric, Fujita Gakuen Health University Hitoshi KAMIYA & Masahiro ITO of Pediatrics, School Medicine, Mie University Michiaki TAKAHASHI & Koichi YAMANISHI of Virology, Reearch Institute for Microbial Diseaes, Osaka University Masami YOSHIDA Dermatology, School Medicine, Kinki University Yoshikazu SHIMOMURA of Ophthalmology, School Medicine, Osaka University Shiro NII of Virology, School Medicine, Okayama University Ryoichi MORI & Yoko MINAGAWA of Virology, School Medicine, Kyushu University Yoichi MINAMISHIMA of Microbiology, Miyazaki Medical University Anti-varicella zoster virus (VZV) mouse monoclonal antibodies conjugated with fluorescein isothiocyanate were evaluated for their usefulness as a practical diagnostic tool in the clinical field by examining cells infected with isolated herpes viruses and 431 clinical samples. The kit stained clearly the cells infected with 14 isolated VZV strains without cross reaction to 15 isolated herpes simplex virus type-1 strains (HSV-1) and 14 type-2 (HSV-2) strains. In clinical specimens, viral antigens of VZV were detected in 92/105 (87.6%) cases of varicella and in 176/190 (92.6%) cases of herpes zoster. Specific fluorescence of VZV was also observed in 5 out of 96 cases diagnosed as HSV infections, although these samples had no specific reaction to HSV when tested by the commercially available diagnostic kit. In 24 cases which could not be clinically diagnosed as herpes zoster or herpes simplex, the VZV antigen was demonstrated in 9 cases. All 109 VZV-positive cases in virus isolation by culture were also judged VZV-antigen positive by the kit, while all 69 HSV-positive cases in virus isolation were VZV-antigen negative. Furthermore, the VZV antigen was detected by the kit in 53/60 clinical diagnoses of varicella or herpes zoster without successful virus isolation. These results clearly indicate the usefulness of the kit as a practical VZV diagnostic reagent, especially in terms of specific sensitivity and easy techical manipulation.
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