重组人源化兔抗血管内皮生长因子单克隆抗体生物学活性检测方法的建立

Development of a method for determination of biological activity of humanized rabbit anti-vascular endothelial growth factor monoclonal antibody

目的建立重组人源化兔抗血管内皮生长因子(Vascular endothelial growth factor,VEGF)单克隆抗体生物学活性检测方法。方法以人脐静脉血管内皮细胞(Human umbilical vein endothelial cells,HUVECs)作为基质细胞,对23支重组人源化兔抗VEGF单抗参比品、24支供试品原液和10支成品进行生物学活性测定,并计算参比品和供试品的IC50值,计算变异系数;对参比品进行活性赋值后,再计算供试品活性,计算变异系数。结果重组人源化兔抗VEGF单抗参比品、原液和成品的IC50值分别为(49.41±23.60)、(47.42±19.45)和(39.13±16.25)ng/ml,变异系数分别为47.76%、41.02%和41.53%;参比品活性赋值为5.08×104U/ml;供试品原液活性为(0.88±0.19)×106U/ml,变异系数为21.6%,供试品成品活性为(1.49±0.41)×106U/支,变异系数为27.5%。结论成功建立了重组人源化兔抗VEGF单抗生物学活性测定方法,确定了参比品的活性单位,为重组人源化兔抗VEGF单抗生物学活性的检测提供了切实可行的方法。

Objective To develop a method for determination of biological activity of humanized rabbit anti-vascular endothelial growth factor（VEGF） monoclonal antibody（McAb）.Methods The biological activities of 23 containers of reference,24 containers of bulk and 10 containers of final product of recombinant humanized rabbit anti-VEGF McAb were determined using human umbilical vein endothelial cells（HUVECs） as matrix,based on which the IC50 values of reference and test samples as well as their variation coefficients were calculated.After the activity of reference was defined,the activities of test samples and their variation coefficient were calculated.Results The IC50 values of reference,bulk and final product of recombinant humanized rabbit anti-VEGF McAb were（49.41 ± 23.60）,（47.42 ± 19.45） and（39.13 ± 16.25） ng / ml,of which the variation coefficients were 47.76%,41.02% and 41.53%,respectively.The activity of reference was defined as 5.08 × 104 U / ml.The activity of bulk was（0.88 ± 0.19） × 106 U / ml,of which the variation coefficient was 21.6%;however,the activity of final product was（1.49 ± 0.41） × 106 U / dose,of which the variation coefficient was 27.5%.Conclusion A method for determination of biological activity of recombinant humanized rabbit anti-VEGF antibody was successfully developed,and the activity unit of reference was defined,which provided a practical tool for determination of biological activity of recombinant humanized rabbit anti-VEGF McAb.