抗人β-HCG单克隆抗体的制备及化学发光检测方法的建立

Preparation of Monoclonal Antibodies Against Human β-HCG and Establishment of a Sandwich CLIA System

目的:制备人绒毛膜促性腺激素(β-HCG)单克隆抗体,建立人β-HCG双抗体夹心CLIA检测方法。方法:用人β-HCG抗原免疫小鼠,通过细胞融合、筛选后得到杂交瘤细胞株,然后将细胞株扩大培养并纯化上清液获得抗体,测定抗体亲和力、特异性及表位,最后建立双抗体夹心CLIA方法。结果:获得4株抗人β-HCG的杂交瘤细胞株(β-1-1、β-2-1、β-3-1、β-4-1)。用β-1-1和β-2-1建立的双抗体夹心法的检测范围为0.5 mIU/mL-800 mIU/mL,灵敏度0.23 mIU/mL,检测结果的相对偏差均在±10%内,回收率在90%以上。结论:本研究最终成功制备了抗人β-HCG mAb,建立了定量检测人β-HCG的双抗体夹心CLIA方法,为β-HCG检测及疾病的诊断奠定基础。

Objective： To prepare human chorionie gonadotrophin （β-HCG） monoclonal antibody, and establish double antibody sandwich CLIA detecting system for human β-HCG. Methods： Mice were immunized with human β-HCG antigen, and then hybridoma cell lines were obtained by cell fusion and screened for the clones which expressed the antibody. The positive cell clones were expanded further for a large volume of supematant, from which the antibodies were purified and tested for affinity, specificity and epitope. Finally the obtained antibodies were used to establish the double antibody sandwich CLIA system. Results： 4 monoclones of anti-human β-HCG （β-1-1,β-2-1, β-3-1, β-4-1） antibody were obtained. β-1-1 and β-2-1 were used to establish the double antibody sandwich system with detection range of 0.5 mIU/mL-800 mlU/mL, sensitivity of 0.23mlU/mL, relative detection deviation of±10% and recovery rate of over 90%. Conclusion： The double antibody sandwich CLIA system for quantitative detection of human β-HCG, using the prepared anti-human β-HCG mAbs, was successfully established, which lay the foundation for the diagnostic β-HCG detection in the diseases.