液相ELISA法测定人血浆抗英夫利西单抗抗体及在克罗恩病患者中应用
Measurements of antibodies to infliximab in plasma by solution ELISA and its application in patients of Crohn's disease
朱扣柱 1丁肖梁 2王燕 3陈卫昌 4缪丽燕2
作者信息
- 1. 苏州大学附属第一医院药学部,江苏苏州 215006;江南大学附属儿童医院(无锡市儿童医院)药学部,江苏无锡 214023
- 2. 苏州大学附属第一医院药学部,江苏苏州 215006
- 3. 江南大学附属儿童医院(无锡市儿童医院)药学部,江苏无锡 214023
- 4. 苏州大学附属第一医院消化内科,江苏苏州 215006
- 折叠
摘要
目的:建立人血浆抗英夫利西单抗抗体(antibodies to infliximab,ATI)免疫分析方法并进行方法学验证.方法:用甘氨酸酸解英夫利西单抗(infliximab,IFX)与ATI结合形成的复合物,然后用生物素标记IFX和辣根过氧化物酶(HRP)标记IFX结合ATI形成桥联复合物,再将溶液转移到链霉亲和素包被的孔板上进行检测.方法建立后采用51例未曾使用过IFX的克罗恩病患者血浆进行验证.结果:检测ATI的ELISA方法经验证筛选临界值为1.16,确证临界值为17.21％,滴度临界值为1.34,批内和批间精密度均小于20％,灵敏度达50ng·mL-1,当阳性对照抗体质量浓度为100ng·mL-1时,药物耐受水平为12.5μg·mL-1.结论:成功建立并验证了检测人血浆ATI的免疫分析方法,适合临床开展治疗药物检测.
Abstract
OBJECTIVE To develop and validate a solution enzyme-linked immunosorbent assay(ELISA)method for detecting antibodies to infliximab(ATI).METHODS The infliximab(IFX)/ATI complexes were dissociated by glycine acid,then biotin-IFX and HRP-IFX conjugated with ATI and then resulting bridging complexes were transferred to a streptavidin matrix coated 96-well plate.After an establishment of ATI assay,plasma samples from 51 IFX-naïve Crohn's disease(CD)patients were utilized for method validation.RESULTS Based upon a panel of plasma samples from 51 IFX-naïve CD patients,the screening cut-point was 1.16(S/N).The confirmatory cut-point was determined as a decrease in signal of＞17.21％.The titer cut-point was 1.34.Both intra-and inter-assay precisions were＜20％of CV.The relative sensitivity of this assay was 50 ng·mL-1.Drug-tolerance at the concentration of 100 ng·mL-1 positive control was ≤12.50 μg·mL-1 IFX.The relative sensitiv-ity of this assay was 50 ng·mL-1.CONCLUSION The immunoassay for measuring ATI has been successfully developed and validated,this method can be used for therapeutic drug monitoring.
关键词
英夫利西单抗/抗英夫利西单抗抗体/克罗恩病/酶联免疫吸附分析法Key words
infliximab/antibodies to infliximab/Crohn's disease/enzyme-linked immunosorbent assay引用本文复制引用
基金项目
苏州市姑苏人才项目(GSWS2019001)
国家自然科学基金青年项目(82003857)
出版年
2024
NETL
NSTL
万方数据