胎盘生长因子串联scFv质粒构建表达及亲和力测定  

Construction,expression and affinity determination of placental growth factor tandem scFv plasmid

作　　者：闫若辰 王文皓 魏琳纳 袁玉华[3] YAN Ruo-chen;WANG Wen-hao;WEI Lin-na;YUAN Yu-hua(Department of Clinical Laboratory,General Hospital,Tianjin Medical University,Tianjin 300052,China;Department of Clinical Laboratory,Blood Diseases Hospital,Chinese Academy of Medical Sciences,Tianjin 300020,China;Department of Clinical Laboratory,General Hospital Airport Site,Tianjin Medical University,Tianjin 300308,China)

机构地区：[1]天津医科大学总医院检验科,天津300052 [2]中国医学科学院血液病医院临床检测中心,天津300020 [3]天津医科大学总医院空港医院检验科,天津300308

出　　处：《天津医科大学学报》2023年第1期78-82,共5页Journal of Tianjin Medical University

基　　金：国家重点研发计划(2020YFC0843100)。

摘　　要：目的:构建并表达胎盘生长因子串联scFv,提升对胎盘生长因子的亲和力。方法:设计不同长度的柔性连接子连接的串联scFv,构建串联scFv质粒,并分别将串联scFv与亲本scFv于原核表达系统进行表达。通过间接ELISA分别测定串联scFv及对应亲本scFv亲和力,分别对数据进行单因素ANOVA方差分析,并使用Dunnett-t检验进行组间比较。结果:柔性连接子长度的不同,会对串联scFv亲和力造成差别。单因素ANOVA方差分析结果表明,各组亲和常数间存在明显的统计学差异,Dunnettt检验组间比较结果可知,当串联scFv间柔性连接子(GGGGS)n长度n=4和n=5时,亲和力较亲本提升不明显(P>0.05);但亲和力在n=6(t_(335)=33.90,t_(7A10)=32.00,均P<0.000 1)及n=7(t_(335)=91.68,t7A10=90.71,均P<0.000 1)时较两亲本均有明显提升。结论:串联scFv可以明显提高单体scFv亲和力。Objective:To construct and express placental growth factor(PLGF) tandem scFv,and determine the affinity for PLGF.Methods:Tandem scFvs connected by flexible linkers of different lengths were designed to construct the tandem scFv plasmid,and the tandem scFv and their parental scFv were expressed in the prokaryotic expression system respectively.The affinity of tandem scFvs and parental scFvs were measured by indirect ELISA.One-way ANOVA analysis was performed on the data and the Dunnett-t test was used to compare between groups.Results:The different lengths of the flexible linkers would cause differences in the affinity of the tandem scFv.The results of the one-way ANOVA analysis showed that there were statistically significant differences between the affinity constants of the groups.The Dunnett-t test for comparison between groups showed that when the length of the flexible linker(GGGGS)n between the tandem scFv was n=4 and n=5,there was no significant increase in affinity over the parents(P>0.05).However,the affinity was significantly higher than that of both parents at n=6 (t_(335)=33.90,t_(7A10)=32.00,both P<0.000 1)and n=7 (t_(335)=91.68,t_(7A10)=90.71,both P<0.000 1).Conclusion:Tandem scFv can considerably enhance monomeric scFv affinity.

关 键 词：胎盘生长因子 串联scFv SCFV 亲和力 

分 类 号：R392-33[医药卫生—免疫学]