CAJ | 학술논문

目的建立定量检测人血清中金属蛋白酶抑制剂-Ⅰ (TIMPⅠ)的微孔板化学发光酶免疫分析方法.方法以辣根过氧化物酶标记TIMP Ⅰ单抗,以过氧化氢(H202)-鲁米诺(luminol)化学发光体系作为检测体系,采用双抗体夹心反应模式,优化筛选温育条件、抗体包被条件、酶标记物稀释度和发光反应时间等指标.同时进行了回收实验,热稳定性实验,并随机选取60例肝纤维化患者血清与进口试剂进行比对实验.结果所建立方法的线性范围为0.2 ～ 12 ng/ml,相关系数0.996,检出限为0.12 ng/ml,批内、批间变异均小于10％.检测TIMP Ⅰ的临床高、中、低值血清回收率为分别为100.6％、96.5％和106.5％;在4℃和37℃条件下分别进行了3、5、7d的稳定性考察,线性相关系数均＞0.98,标准偏差＜6％；比对实验分析证明2种方法相关性具有统计学意义.结论成功建立定量TIMP Ⅰ微孔板化学发光酶免疫分析方法,该方法具有较高的准确性、灵敏度及良好的重复性,并与进口试剂检测结果一致.
목적 건립정량검측인혈청중금속단백매억제제-Ⅰ (TIMPⅠ)적미공판화학발광매면역분석방법.방법 이랄근과양화물매표기TIMP Ⅰ단항,이과양화경(H202)-로미낙(luminol)화학발광체계작위검측체계,채용쌍항체협심반응모식,우화사선온육조건、항체포피조건、매표기물희석도화발광반응시간등지표.동시진행료회수실험,열은정성실험,병수궤선취60례간섬유화환자혈청여진구시제진행비대실험.결과 소건립방법적선성범위위0.2 ～ 12 ng/ml,상관계수0.996,검출한위0.12 ng/ml,비내、비간변이균소우10％.검측TIMP Ⅰ적림상고、중、저치혈청회수솔위분별위100.6％、96.5％화106.5％;재4℃화37℃조건하분별진행료3、5、7d적은정성고찰,선성상관계수균＞0.98,표준편차＜6％；비대실험분석증명2충방법상관성구유통계학의의.결론 성공건립정량TIMP Ⅰ미공판화학발광매면역분석방법,해방법구유교고적준학성、령민도급량호적중복성,병여진구시제검측결과일치.
Objective To establish microplate chemiluminescence enzyme immunoassay (CLEIA) for quantitative analysis of tissue inhibitor of metalloproteinases Ⅰ (TIMP Ⅰ) in human serum.Methods A sandwich reaction was preformed with horseradish peroxidase(HRP) labeled monoclonal antibody of TIMP Ⅰ as the catalytic enzyme and the H2O2-luminol as the luminescence reagent.Several physical and chemical parameters were studied and optimized such as immunoreaction conditions,the dilution ratio of TIMP IHRP,luminescence reaction time and so on.In order to evaluate the method,recovery test,heat stabilization test and comparison test were carried out.Results The linear range was 0.2-12 ng/ml with r =0.996.The detection limit was 0.12 ng/ml.Inter-assay and intra-assay RSD were both less than 10％.The recoveries of three different spiked concentration samples were 100.6％,96.5％ and 106.5％.After stored at 4℃ and 37℃ for 3,5,7 days,the analysis showed correlation coefficient higher than 0.998 and RSD lower than 6％.The detected results with CLEIA closely corresponded to those with imported ELISA in 60 patients sera with liver fibrosis.Conclusion Established CLEIA for quantity determination of serum TIMP I has high accuracy,sensitivity and repeatability.