细胞骨架蛋白4荧光免疫层析检测方法的建立

Establishment of fluorescence immunochromatography detection for cytoskeleton-associated protein 4

摘要:

采用基于免疫层析技术与荧光微球标记技术相结合的方式，建立一种快速、简单的定量检测肝癌肿瘤标记物的方法。依据双抗体夹心原理，将细胞骨架蛋白4 (CKAP4) 配对抗体分别作为标记与包被抗体，羊抗兔多抗作为质控线包被抗体，制备CKAP4荧光免疫层析试纸条，并对试纸条的线性、精密性、稳定性等各项性能指标进行评价。结果表明，采用时间分辨荧光微球所制备的CKAP4免疫层析试纸条灵敏度高，特异性好，精密性在15%以内，回收率在85%–115%之间，线性范围为25–1 000 pg/mL，可在37℃稳定保存20 d，与商业化的ELISA试剂盒的相关性良好。结论：初步建立了CKAP4荧光免疫层析方法，能够定量检测血清中CKAP4的含量，且具有快速、灵敏、简便、经济、可单人份操作等优点，有望成为肝癌辅助诊疗的新方法。

Abstract:

A rapid and simple method to detect tumor markers in liver cancer was established by combining immunochromatography technique with fluorescent microsphere labeling. According to the principle of double antibody sandwich, the cytoskeleton-associated protein 4 (CKAP4) paired antibody was used as the labeled and coated antibody, and the goat anti-rabbit polyclonal antibody was used as the quality control line coated antibody in the preparation of the CKAP4 fluorescent immunochromatographic test strips. After the preparation, the test strips were evaluated on various performance indicators, such as linearity, precision and stability. The CKAP4 immunochromatographic strip prepared by time-resolved fluorescent microspheres had high sensitivity, and good specificity. Its precision was within 15%, recovery between 85% and 115%, and linear range between 25 and 1 000 pg/mL. The test strip could be kept stable at 37 °C for 20 days, and it correlated well with commercial ELISA kits. The CKAP4 fluorescence immunochromatography method can quantitatively detect the content of CKAP4 in serum. Furthermore, it is rapid, sensitive, simple, economical and single-person operation. This method has the potential of becoming a new method for the diagnosis and treatment of liver cancer.