Development of the high sensitivity measurements for drug-allergy

药物过敏高灵敏度测量方法的开发

• 批准号: 11672310
• 负责人: ITO Masatoshi
• 金额: $ 1.92万
• 依托单位: Toho university
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11672310/
• 关键词: Drug allergy; Anti-drug antibody; DLST; Interleukin-2; RT-PCR; RAD; Affinity content; Competitive ELISA; インタ-ロイキン-2; インターロイキン2; 感作T細胞; 抗薬剤抗体; competitive ELISA; PCR法; サイトカイン

Drug allergy was examined on the supersensitive detection method for the determinant of drug antigen and drag-lymphocyte blastogenesis test. In ELISA, the antibody detection was carried out using 3 kinds of cefaclor/OVA complex of which the cross-link position differed. First, the side-chain(R 1) of cefaclor was recognized by anti-cefaclor IgG antibodies obtained from cefaclor-allergy patient. In contrast, the IgM antibody reacted at 7-aminocephalosporanic acid (7-AMA) of cefaclor. Competitive ELISA shown that anti-cefaclor side-chain antibodies were competitive inhibitied by ampicillin, becampicillin, cefaclor and cephalexin, which were composed from α-phenylglycine side-chain. Anti-cefaclor IgG antibodies reacted to the α-phenylglycine (R1) side- chain in a dose response, but not p-amino phenyl acetic acid and 7-AMA. Furthermore, BIAcore analysis indicated that the affinity of anti-cefaclor antibody was Ka=2. 19 x 10-3M-1and Kd=2.56 x 10-4 M-l , which were almost equal to other the immune antibody. On the other hand, DLST was positive in 3 out of 25 patients, and false positives were 3 patients. In contrast, IL-2 was detected from 4 to 8 hrs after the stimulation by drugs. Moreover,IL-2 mRNA was detected in all patients with the IL-2 production.These results suggested that the detection method of IL-2 protein and mRNA are more super-sensitive than the DLST, and this method is effective diagnosed test for drug-allergy.

采用药物抗原决定簇超灵敏检测法和拖曳淋巴细胞胚细胞试验检测药物过敏情况。在ELISA中，使用交联位置不同的3种头孢克洛/OVA复合物进行抗体检测。首先，头孢克洛的侧链(R 1)被从头孢克洛过敏患者获得的抗头孢克洛IgG抗体识别。相反，IgM 抗体与头孢克洛的 7-氨基头孢烷酸 (7-AMA) 发生反应。竞争性ELISA结果表明，抗头孢克洛侧链抗体可被由α-苯甘氨酸侧链组成的氨苄西林、贝卡西林、头孢克洛和头孢氨苄竞争性抑制。抗头孢克洛 IgG 抗体在剂量反应中与 α-苯基甘氨酸 (R1) 侧链发生反应，但不与对氨基苯乙酸和 7-AMA 发生反应。此外，BIAcore分析表明抗头孢克洛抗体的亲和力Ka=2。 19×10-3M-1，Kd=2.56×10-4M-1，与其他免疫抗体几乎相当。另一方面，25 名患者中有 3 名 DLST 呈阳性，3 名患者呈假阳性。相反，在药物刺激后4至8小时检测到IL-2。而且，所有有IL-2产生的患者中均检测到了IL-2 mRNA。这些结果表明IL-2蛋白和mRNA的检测方法比DLST更灵敏，该方法是药物过敏的有效诊断测试。