Abstract:
Adiponectin (ADPN), which serum/plasma adiponectin levels are closely associated with insulin resistance and type 2 diabetes, and lower adiponectin levels predict an increased risk of diabetes, is a strong indicator of diabetes risk in people at high risk of diabetes in different races. Using the unique principle and performance advantages of chemiluminescence immunoassay (CLIA), an ADPN-CLIA method with high sensitivity, high specificity and wide detection range was established based on the principle of two-steps method of sandwich-type, with the magnetic particles (MPs) as the solid phase carrier and acridinium ester (AE) as the chemiluminescence reaction system. The selection of the main raw materials required, the preparation conditions of MPs-coated antibodies, the methods of AE-labeled antibodies, sample requirements and reaction modes were optimized and evaluated. AE labeling experiment was successfully performed with the labeling efficiency of 8.366 and the antibody utilization rate of 96.8%. The chemiluminescent immunoassay for ADPN had a good linear relationship from 0 ng/mL to 250 ng/mL (R2 =0.9993), with the detection limit of 0.05 ng/mL. The coefficient of variation (CV) of intra-assay and inter-assay precision were both less than 5% respectively. The recovery rates for accuracy were from 91.26% to 107.46%. The comparison experiment of 80 clinical serum samples between the developed ADPN-CLIA with the immunoturbidimetry showed that the correlation coefficient was 0.956, and the Bland-Altman analysis showed that the limits of agreement were - 0.364 and 0.433.
摘要:
脂联素(ADPN),血清/血浆脂联素水平与胰岛素抵抗和2型糖尿病密切相关,较低的脂联素水平预测糖尿病风险增加,是不同种族糖尿病高危人群糖尿病风险的有力指标。利用化学发光免疫分析(CLIA)的独特原理和性能优势,一种高灵敏度的ADPN-CLIA方法,基于夹心式两步法的原理,建立了高特异性和宽检测范围,以磁性颗粒(MPs)为固相载体,吖啶酯(AE)为化学发光反应体系。所需主要原材料的选择,MPs包被抗体的制备条件,AE标记抗体的方法,样品要求和反应模式进行了优化和评估。成功进行了AE标记实验,标记效率为8.366,抗体利用率为96.8%。ADPN的化学发光免疫分析在0ng/mL到250ng/mL之间具有良好的线性关系(R2=0.9993),检出限为0.05ng/mL。测定内和测定间精密度的变异系数(CV)均小于5%。准确度的回收率为91.26%至107.46%。开发的ADPN-CLIA与免疫比浊法之间的80份临床血清样品的比较实验表明,相关系数为0.956,Bland-Altman分析表明,一致的极限为-0.364和0.433。