人血清癌胚抗原的生物素-亲和素ELISA检测方法的建立

目的:建立检测血清癌胚抗原(CEA)的高灵敏度生物素-亲和素酶联免疫检测(BA-ELISA)方法。方法:亲和层析纯化得到CEA,免疫新西兰家兔制备多克隆抗体。将得到的抗体连接生物素和辣根过氧化物酶,在生物素化BSA包被的96微孔板上建立生物素-亲和素ELISA(BA-ELISA)。对这一体系的线性、灵敏度、特异度、稳定性、回收率等参数进行鉴定,并和常规ELISA、放射免疫检测以及化学发光法相比较检测了临床标本中的CEA浓度。结果:线性检测范围为(0.42～50)U/mL,检测结果表明,体系稳定性良好;灵敏度为0.42 U/mL,批内、批间差异均小于10.0%。该研究建立的方法与常规ELISA对比差异有统计学意义,与放射免疫检测对比差别无统计学意义。与化学发光法相比较,回归方程为:y=0.04825+0.99674x,r=0.994,差异无统计学意义。结论:建立的CEA的BA-ELISA检测方法易于操作、灵敏度高、价格低廉,适合应用于临床检测。

Establishment of BA-ELISA method for detecting CEA in human sera

AIM: To establish a sensitive biotin-avidin enzyme linked immunosorbent assay(BA-ELISA) method for detecting carcinoembryonic antigen(CEA) in serum.METHODS: CEA which had been purified by affinity chromatography was used to immunize the rabbits to produce polyclonal antibodies.Then the antibodies were connected with biotin and horseradish peroxidase(HRP).So BA-ELISA method was established on the basis of 96 microwell plates coated with biotinylated BSA.Finally we examined the sensitivity,specificity,stability and recovery rate of this system and compared the BA-ELISA method with the traditional ELISA,radioimmunoassay and chemiluminescence in detecting CEA concentrations.RESULTS: The stability of this system was proved good.The linear range was from 0.42 to 50 U/mL,the sensitivity was 0.42 U/mL,and the intra-differences together with inter-differences were less than 10.0%.There was significant difference between BA-ELISA and traditional ELISA,while there was no significant difference between BA-ELISA and radioimmunoassay.The regression equation of this method was y=0.04825+0.99674x and r=0.994,and there was no significant difference between the BA-ELISA and chemiluminescence.CONCLUSION: The BA-ELISA method we established to detect CEA was easy to operate,highly sensitive,low in price and suitable for application in clinical detection.