摘要
目的基于可重复利用的免疫磁珠,建立一种简捷快速的特异性抗体检测方法。方法设计人类巨细胞病毒(humancytomegaloviruses,HCMV)PP150蛋白的抗原表位,并合成8分枝多聚抗原肽PP150-8MAPs。将PP150-8MAPs以共价结合形式包被于DynabeadsM-450Tosylactivated磁珠表面,制备特异性免疫磁珠。用PP150-8MAPs免疫Balb/c小鼠制备抗此抗原表位的标准抗血清。应用免疫磁珠检测标准抗血清中的抗体,优化检测条件。在抗体检测反应结束后,洗脱抗体-二抗复合物,再生后的免疫磁珠重复用于标准抗血清样品的检测分析,并分析免疫磁珠可重复利用的次数。结果制备免疫磁珠时,PP150-8MAPs的最佳包被量为100μg/mL,包被效率为79%。用PP150-8MAPs免疫小鼠后,得到的抗血清滴度可达到112800。用免疫磁珠法检测小鼠标准抗血清,免疫磁珠可重复进行20次以上的检测分析。结论基于酶联免疫检测方法,包被抗原的免疫磁珠可重复应用于血清样品中特异性抗体的检测分析。此实验为建立一种新的可重复检测的高效清样品中特异性抗体的检测分析。此实验为建立一种新的可重复检测的高效免疫磁珠抗体分析方法奠定了基础,在基础研究与临床检测中具有广泛的应用前景。
Objective To develop a new method to detect antibodies quickly and especially based on reusable immunomagnetic beads. Methods Eight branches multiple antigen peptides （PP150-8MAPs） were synthesized according to the epitope sequence from Human cytomegaloviruses pp150 protein. Then the immunomagnetic beads were prepared by coating PP150-8MAPs onto the surfaces of Dynabeads M-450 （Tosylactivated） in covalent binding form. The standard serum sample was obtained by immunizing Balb/c mice with PP150-8MAPs. The antibodies in the standard serum were detected by immunomagnetic beads and the detecting conditions were optimized. The antibody compounds were removed from immunomagnetic beads by citric acid, and then the beads were reused for antibody detection. Results Total 79% PP150-8MAPs could be coated on the surfaces of Dynabeads M-450 （Tosylactivated） when the concentration of PP150-8MAPs was 100 μg/mL. The antibodies titration of the Balb/c mice serum was reached 1 ： 12 800 after immunization. The immunomagnetic beads could be reused to detect antibodies over 20 times. Conclusion The immunomagnetic beads coated antigens can be reused to detect the antibodies in serum samples. This method can be used widely for antibody analysis in immunology research or clinical diagnosis of infection diseases in the future.
出处
《免疫学杂志》
CAS
CSCD
北大核心
2006年第5期577-579,583,共4页
Immunological Journal
关键词
抗原表位
多聚抗原肽
免疫磁珠
Epitope
Multiple antigen peptides
Immunomagnetic beads
