两种供试品不同浸提条件及作用剂量下的体外人淋巴细胞增殖实验  

In vitro human lymphocyte proliferation assay under different extraction conditions and doses of two types of test samples

作　　者：许建霞[1] 付海洋[1] 曲守方[1] Xu Jianxia;Fu Haiyang;Qu Shoufang(National Institutes for Food and Drug Control,Beijing 102629,China)

机构地区：[1]中国食品药品检定研究院,北京市102629

出　　处：《中国组织工程研究》2024年第31期5017-5021,共5页Chinese Journal of Tissue Engineering Research

摘　　要：背景:体外淋巴细胞增殖实验常用于检测医疗器械潜在的免疫原性,但在相关标准中均未给出详尽的浸提条件及作用剂量。目的:考察供试品不同浸提条件及作用剂量对体外人淋巴细胞增殖的影响,思考在选择体外淋巴细胞增殖实验条件时需考虑的因素。方法:实验检测同种骨修复材料与肝素修饰人工晶状体两种供试品,均分为以下12组:①实验组1:供试品24 h完全培养基(含体积分数10%胎牛血清的RPMI改良培养基)浸提液200μL+淋巴细胞悬液50μL;②阴性对照组1:24 h完全培养基200μL+淋巴细胞悬液50μL;③实验组2:供试品24 h完全培养基浸提液100μL+淋巴细胞悬液100μL;④阴性对照组2:24 h完全培养基100μL+淋巴细胞悬液100μL;⑤实验组3:供试品72 h RPMI改良培养基浸提液(实验前加体积分数10%胎牛血清)200μL+淋巴细胞悬液50μL;⑥阴性对照组3:72 h RPMI改良培养基(实验前加体积分数10%胎牛血清)200μL+淋巴细胞悬液50μL;⑦实验组4:供试品72 h RPMI改良培养基浸提液(实验前加体积分数10%胎牛血清)100μL+淋巴细胞悬液100μL;⑧阴性对照组4:72 h RPMI改良培养基(实验前加体积分数10%胎牛血清)100μL+淋巴细胞悬液100μL;⑨阳性对照组1:含10μg/mL植物血凝素M的完全培养基200μL+淋巴细胞悬液50μL;⑩阳性对照组2:含10μg/mL植物血凝素M的完全培养基100μL+淋巴细胞悬液100μL;⑪空白对照组1:250μL完全培养基;⑫空白对照组2:200μL完全培养基。培养3 d后,采用CCK-8法检测淋巴细胞增殖。结果与结论:①不同实验条件下,同种骨修复材料浸提液均可增强人淋巴细胞的活性,以RPMI改良培养基浸提72 h、浸提液与淋巴细胞悬液的体积比为4∶1的实验条件最为显著;肝素修饰人工晶状体在该条件下对淋巴细胞活性有明显的抑制作用,可能与浸提液中的肝素有关,但在完全培养基浸提24 h、浸提液与淋巴细胞悬液的体积比为4∶1BACKGROUND:In vitro lymphocyte proliferation test is often used to detect the potential immunogenicity of medical devices,but no detailed extraction conditions and dose are given in the relevant standards.OBJECTIVE:To investigate the effects of different extraction conditions of the test product and different doses of the extract on in vitro human lymphocyte proliferation,and to consider the factors that need to be considered when selecting test conditions for in vitro lymphocyte proliferation test.METHODS:In the experiment,the homogenous bone repair material and heparin-modified intraocular lens were divided into the following 12 groups:(1)Experimental group 1:24-hour complete medium(RPMI modified medium containing 10%fetal bovine serum)extract of 200μL+lymphocyte suspension of 50μL;(2)negative control group 1:24-hour complete medium 200μL+lymphocyte suspension 50μL;(3)experimental group 2:24-hour complete medium extract 100μL+lymphocyte suspension 100μL;(4)negative control group 2:24-hour complete medium 100μL+lymphocyte suspension 100μL;(5)experimental group 3:72-hour RPMI modified medium extract(addition of 10%fetal bovine serum before experiment)200μL+lymphocyte suspension 50μL;(6)negative control group 3:72-hour RPMI modified medium(addition of 10%fetal bovine serum before experiment)200μL+lymphocyte suspension 50μL;(7)experimental group 4:72-hour RPMI modified medium extract(addition of 10%fetal bovine serum before experiment)100μL+lymphocyte suspension 100μL;(8)negative control group 4:72-hour RPMI modified medium(addition of 10%fetal bovine serum before experiment)100μL+lymphocyte suspension 100μL;(9)positive control group 1:complete medium containing 10μg/mL plant hemagglutinin-M 200μL+lymphocyte suspension 50μL;(10)positive control group 2:complete medium containing 10μg/mL plant hemagglutinin-M 100μL+lymphocyte suspension 100μL;(11)blank control group 1:250μL complete medium;(12)control group 2:200μL complete medium.After 3 days of culture,the proliferation of lymphocytes was d

关 键 词：体外人淋巴细胞增殖 不同实验条件 浸提液 同种骨修复材料 肝素修饰人工晶状体 

分 类 号：R459.9[医药卫生—治疗学] R392.12[医药卫生—临床医学] R392.8