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期刊名称:药物分析杂志
主管单位:中国科学技术协会
主办单位:中国药学会
承办:中国食品药品检定研究院
主编:金少鸿
地址:北京天坛西里2号
邮政编码:100050
电话:010-67012819,67058427
电子邮箱:ywfx@nicpbp.org.cn
国际标准刊号:ISSN 0254-1793
国内统一刊号:CN 11-2224/R
邮发代号:2-237
 

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成像毛细管等电聚焦电泳测定单抗等电点的若干影响因素

Several factors on isoelectric point determination of monoclonal antibody by imaging capillary isoelectric focusing electrophoresis(iCIEF)

作者(英文):
分类号:R917
出版年·卷·期(页码):2018,38 (10):1727-1732
DOI: 10.16155/j.0254-1793.2017.01.01
-----摘要:-------------------------------------------------------------------------------------------

目的:评价尿素、聚焦时间、pI标记物3个因素对成像毛细管等电聚焦电泳(iCIEF)测定抗HER2单抗、抗VEGF单抗主峰等电点结果的影响。方法:应用iCIEF技术,通过添加尿素(2 mol·L-1),改变聚焦时间(6、8、10 min)和pI标记物的数量(2个/6个),对2种单抗制品主峰等电点进行测定。结果:抗HER2单抗使用6个pI标记物,添加终浓度2 mol·L-1的尿素,聚焦时间6、8、10 min,主峰等电点分别为9.01±0.00、9.02±0.01、9.06±0.01;使用6个pI标记物,不添加尿素,聚焦时间6、8、10 min,主峰等电点分别为9.07±0.01、9.08±0.01、9.09±0.01;使用2个pI标记物,主峰等电点范围为9.00~9.14(含尿素)、9.04~9.17(不含尿素)。抗VEGF单抗使用6个pI标记物,添加终浓度2 mol·L-1的尿素,聚焦时间6、8、10 min,主峰等电点分别为8.32±0.00、8.35±0.00、8.35±0.07;使用6个pI标记物,不添加尿素,聚焦时间6、8、10 min,主峰等电点分别为8.39±0.01、8.39±0.02、8.41±0.01;使用2个pI标记物,主峰等电点范围为8.26~8.48(含尿素)、8.34~8.49(不含尿素)。结论:使用iCIEF评价所选单抗等电点时,尿素、聚焦时间两因素对等电点的计算结果影响较小,pI标记物的选择对等电点的计算结果影响明显。

-----英文摘要:---------------------------------------------------------------------------------------

Objective:To evaluate the effects of the three factors(urea, focusing time and pI marker) on the determination of the isoelectric points of anti-HER2 and anti-VEGF monoclonal antibody electrophoresis by imaging capillary isoelectric focusing electrophoresis(iCIEF).Methods:By adding urea(2 mol·L-1), changing the focus time(6, 8, 10 min), and the number of pI markers(2/6), the iCIEF technique was applied to determine the isoelectric points of two monoclonal antibody products. Results:The anti-HER2 monoclonal antibody used 6 pI markers, and added urea with a final concentration of 2 mol·L-1. The focusing time was 6, 8 and 10 min, and the isoelectric points of the main peak were 9.01±0.00, 9.02±0.01, 9.06±0.01, respectively. Six pI markers were used without urea, the focusing time was 6, 8, 10 minutes, and the main peak isoelectric points were 9.07±0.01, 9.08±0.01, 9.09±0.01, respectively. Using two pI markers, the pI ranges were 9.00-9.14 (with urea) and 9.04-9.17 (without urea). The anti-VEGF antibody used 6 pI markers, and added urea with a final concentration of 2 mol·L-1. The focusing time was 6, 8 and 10 min, the main peak isoelectric points were 8.32±0.00, 8.35±0.00, 8.35±0.07. Six pI markers were used and no urea was added. The focusing time was 6, 8, and 10 min, and the main peak isoelectric points were 8.39±0.01, 8.39±0.02, 8.41±0.01, respectively. Using two pI markers, the pI ranges were 8.26-8.48 (with urea) and 8.34-8.49 (without urea). Conclusion:When iCIEF is used to evaluate the isoelectric point of the monoclonal antibody, the two factors, urea and focusing time have little influence on the calculation of isoelectric point, and the selection of pI markers has obvious influence on the calculation results of isoelectric point..

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