免疫复合物型治疗性乙型肝炎疫苗有效成份佐剂吸附率检测初探  

Explore to Assay the Adsorbing Percentage of Therapeutic Hepatitis B Vaccine Major Effective Component to Alum Adjuvant

作　　者：朱亭玉 徐静[2] 何亚丽 王明莹 齐璇 周根喜 李津[1,3] 

机构地区：[1]北京天坛生物制品股份有限公司乙肝疫苗室,北京100024 [2]北京生物制品研究所免疫研究室,北京100024 [3]北京大学基础医学院病原生物学系,北京100191

出　　处：《生物技术通讯》2010年第1期43-46,共4页Letters in Biotechnology

基　　金：国家重大科技专项(2008ZX10002-002)

摘　　要：目的:探索免疫复合物型治疗性乙型肝炎疫苗"乙克"的主要有效成份HBsAg/抗-HBs复合物(IC)及游离HBsAg的佐剂吸附率检测方法。方法:氢氧化铝佐剂经不同转速离心后,用ICP-AES法检测上清中铝残留量。未吸附IC经不同转速离心后,用电化学发光法检测在各离心转速下上清中HBsAg和抗-HBs含量。用电化学发光法检测未吸附IC与同一批次的佐剂吸附IC的HBsAg含量,计算HBsAg回收率;检测佐剂吸附IC样品离心上清及未离心样品的HBsAg含量,计算吸附百分率及检测的精密度。用凯氏定氮法测定佐剂吸附免疫复合物离心上清与离心前样品蛋白含量。结果:氢氧化铝佐剂经6500r/min离心3min,上清中铝的残留量为0;未吸附IC经2000～9000r/min离心3min,所测上清中HBsAg与抗-HBs均未明显下降。佐剂吸附IC经解离后,HBsAg回收率为91.56%,表明经过解离后铝佐剂不会影响HBsAg含量的检测结果。佐剂吸附IC样品与离心上清比较,蛋白含量减少了3.2mg/mL,表明每毫克铝可吸附约2.54mg蛋白,对总蛋白的吸附率为13.5%。佐剂吸附IC及HBsAg的吸附百分率的重复性检测结果为99.87%±0.15%,CV为0.15%。结论:佐剂吸附的IC在6500r/min离心3min的条件下,上清中未被吸附的HBsAg和抗-HBs不会下沉,而氢氧化铝佐剂连同被吸附IC及HBsAg则被沉淀,且铝佐剂不会对HBsAg的测定造成干扰,故通过检测离心样品上清及未离心样品中的HBsAg含量,可有效测定铝吸附百分率,且精密度较好。Objective：To explore the assay method for adsorbing percentage of therapeutic hepatitis B（HB）vaccine YIC major effective component HBsAg/anti-HBs immune complex（IC）and dissociated HBsAg to alum adjuvant.Methods：Alum adjuvant samples were centrifuged at different speeds,the residual alums in the supernatants were tested with ICP-AES.Pre-adsorbed IC samples were centrifuged at various speed,the residual HBsAg and anti-HBs in the supernatants were detected by ECLIA.ECLIA was employed to accurately measure the HBsAg quantities in the pre-adsorbed IC samples as well as the alum adsorbed IC samples at the same lots,calculate recovery of HBsAg.ECLIA were employed to accurately measure the HBsAg quantities in the alum adsorbed IC samples as well as supernatants of YIC samples centrifuged at 6500 r/min for 3 mins,the saturate protein adsorbed to alum,the adsorbing percentage of IC to alum adjuvant and the precision of latter were induced respectively.Assay of protein in supernatant of centrifuged adjuvant adsorbed IC sample and uncentrifuged sample with Kjeldah method.Results：The trace alum in the supernatants of alum adjuvant centrifuged at 6500 r/min for 3 mins dropped to zero.Un-adsorbed ICs centrifuged at 2000～9000 r/min for 3mins,the HBsAg and anti-HBs inthe supernatants remained roughly at similar residual level,suggesting that the HBsAg would not sediment at the centrifuge condition.After the dissociation of the alum adsorbed IC,the recovery of HBsAg is 91.56%.It reveals that the assay result of HBsAg contents will not be influenced by alum.Compared to YIC samples,the protein concentration in centrifuged supernatants decreased by 3.2 mg/mL（around 13.5%）,implying that per mg alum could adsorb about 2.54 mg protein.Consecutive tests of YIC samples showed that the adsorbing percentage of IC to alum adjuvant was 99.87%±0.15%and the CV was 0.15%.Conclusion：When centrifuged at 6500 r/min for 3 mins,the un-adsorbed HBsAg and anti-HBs would not sediment and interfere the adsorbing percentage assay of

关 键 词：治疗性乙型肝炎疫苗 免疫复合物 氢氧化铝 吸附百分率 

分 类 号：Q502[生物学—生物化学] R392.1[医药卫生—免疫学]