摘要
目的建立重组人肠道病毒71型(enterovirus 71,EV71)病毒样颗粒(virus-like particles,VLPs)等电点(isoelectric point,pI)全柱成像毛细管等电聚焦电泳(whole column imaging detection-capillary isoelectric focusing,WCID-CIEF)测定方法,并进行验证。方法采用WCID-CIEF测定重组人EV71 VLPs的pI,对WCID-CIEF的聚焦时间和尿素浓度进行优化,并对方法的专属性、准确度、重复性和中间精密度进行验证。结果选择3000 V聚焦5 min作为WCIDCIEF的聚焦条件,样品处理时不添加尿素。空白缓冲液对重组人EV71 VLPs pI的测定无干扰;WCID-CIEF分别重复测定pI marker 5.85和pI marker 7.05样品6次的相对标准偏差(relative standard deviation,RSD)分别为0.18%和0.06%;重复测定6次重组人EV71 VLPs pI的RSD为0.08%;不同实验人员在不同日期重复测定12次重组人EV71 VLPs pI的RSD为0.14%。结论WCID-CIEF测定重组人EV71 VLPs pI的专属性、准确度、重复性和中间精密度均良好,该方法快速、准确,可用于重组人EV71 VLPs的质量评价。
Objective To develop and verify a method for determination of isoelectric point(pI)of virus-like particles(VLPs)of recombinant enterovirus(EV)71(EV71 VLPs)by whole column imaging detection-capillary isoelectric focusing(WCID-CIEF)electrophoresis.Methods The pI of recombinant human EV71 VLPs was determined by WCID-CIEF electrophoresis.The developed method was verified for specificity,accuracy,reproducibility and intermediate precision.Results In WCID-CIEF electrophoresis,the focusing was performed at 3000 V for 5 min,and no urea was added when the samples were treated.Blank buffer showed no interference to the determination of pI of EV71 VLPs.The relative standard deviations(RSDs)of determination results of pI marker 5.85 and pI marker 7.05 in six repeat tests by WCIUDCIEF electrophoresis were 0.18%and 0.06%respectively,while that of EV71 VLPs was 0.08%.However,the RSD of determination results of pI of recombinant human EV71 VLPs in 12 repeat tests by different personnel on different dates was 0.14%.Conclusion The WCID-CIEF electrophoresis for determination of pI of recombinant human EV71 VLPs showed high specificity,accuracy,reproducibility and intermediate precision,which might be used for the quality evaluation of recombinant human EV71 VLPs.
作者
王文伟
何成
王蓓
楼觉人
WANG Wen-wei;HE Cheng;WANG Bei;LOU Jue-ren(Shanghai Wellvax Co.,Ltd.,Shanghai 200051,China;不详)
出处
《中国生物制品学杂志》
CAS
CSCD
北大核心
2021年第11期1358-1361,1369,共5页
Chinese Journal of Biologicals
基金
上海市科委生物医药领域科技支撑项目(13431900600)。
关键词
肠道病毒71型
病毒样颗粒
全柱成像毛细管等电聚焦电泳
等电点
Enterovirus 71(EV71)
Virus-like particles(VLPs)
Whole column imaging detection-capillary isoelectric focusing(WCID-CIEF)electrophoresis
Isoelectric point(pI)
