摘要
目的纯化人血清中多克隆抗RhD抗体,并对其进行鉴定。方法收集5例RhD阴性者血清,经不规则抗体筛查和确认后,按等比例混合,分别用RhD(-)A型红细胞和RhD(-)B型红细胞对血清中的抗A、抗B抗体于4℃过夜吸收处理;用RhD(+)O型红细胞制备血影细胞,以血影细胞作为抗原,吸附血清中抗RhD抗体,再用低pH值缓冲液洗脱、分离纯化抗RhD抗体,用DEAE-Sephadex A-50层析法提取其中IgG组分;分别用western blot、微柱法鉴定。结果 5名研究对象血清中均含抗RhD抗体;血清中的抗A和抗B血型抗体被完全吸收,纯化后抗RhD抗体IgG组分效价达到1∶32。结论本实验获得了高效价的抗RhD多克隆抗体,为进一步利用噬菌体肽库获得RhD血型抗原表位研究等奠定基础。
Objective To purify and identify polyclonal anti-RhD antibody from human sera.Methods The sera from five RhD-negative cases were screened and affirmed by irregular antibodies and mixed according to equal proportion.The anti-A and anti-B antibodies in the sera were absorbed by using RhD-negative erythrocytes with type A and type B at 4 ℃ overnight.The ghost cells were prepared by RhD-positive erythrocyte with blood type O,and were used to be the antigens to absorb anti-RhD antibody from the mixed sera.These cells were eluted by low pH value buffer and separated by purified anti-RhD antibody.The IgG fractions of anti-RhD antibody were separated by DEAE-Sephadex A-50 column and were identified by western blot and microcolumn technique.Results The sera from all the five investigated subjects contained anti-RhD antibody.The anti-A and anti-B type antibodies in the purified sera were entirely absorbed.The titer of IgG fraction of the purified anti-RhD antibody reached 1∶32.Conclusion The anti-RhD polyclonal antibody was successively obtained in this study and may used to be the material for the research on the epitopes of RhD blood type antigen by phage peptide library.
出处
《临床检验杂志》
CAS
CSCD
北大核心
2012年第3期190-192,共3页
Chinese Journal of Clinical Laboratory Science
基金
国家自然科学基金专项资助项目(30940067)
关键词
RH血型
抗原
抗体
纯化
血影细胞
Rh blood type
antigen
antibody
purification
ghost cells
