军事医学
軍事醫學
군사의학
BULLETIN OF THE ACADEMY OF MILITARY MEDICAL SCIENCES
2014年
8期
617-620,625
,共5页
侯春梅%郎小玲%肖燕%郑明克%蒋兴伟%肖鹤%陈国江%王仁喜%冯健男%韩根成%沈倍奋%黎燕
侯春梅%郎小玲%肖燕%鄭明剋%蔣興偉%肖鶴%陳國江%王仁喜%馮健男%韓根成%瀋倍奮%黎燕
후춘매%랑소령%초연%정명극%장흥위%초학%진국강%왕인희%풍건남%한근성%침배강%려연
Tim-3%抗体,单克隆%L3D%U937细胞%炎性因子%脓毒症
Tim-3%抗體,單剋隆%L3D%U937細胞%炎性因子%膿毒癥
Tim-3%항체,단극륭%L3D%U937세포%염성인자%농독증
Tim-3%antibodies,monoclonal%L3D%U937 cells%inflammatory factors%sepsis
目的：制备抗人Tim-3单抗并评价其生物学活性，为干预Tim-3表达失调引发的疾病提供实验基础。方法制备重组人Tim-3蛋白，常规方法免疫BALB/c小鼠，筛选出阳性克隆，并对其识别、中和活性进行体内外验证，探讨其作为免疫干预手段的可行性。结果①获得一株能够分泌具有较好结合活性的抗人Tim-3单克隆抗体细胞株（克隆号L3D），其分泌的免疫球蛋白亚型为IgG2a；②流式实验结果表明，该抗体能够结合人U937细胞上的Tim-3分子，且与小鼠RAW 264．7细胞上Tim-3有一定的交叉结合活性；③该抗体能够阻断Gal-9分子诱导的人THP1细胞凋亡，具有良好的体外中和活性；④体内注射L3D，显示该抗体能够显著加重脓毒血症模型小鼠病情，提高炎性因子的表达水平，显示出其良好的体内中和活性。结论成功制备一株分泌抗人Tim-3抗体的细胞株，其良好的结合及中和活性为基于Tim-3表达异常相关疾病的干预提供了实验基础。
目的：製備抗人Tim-3單抗併評價其生物學活性，為榦預Tim-3錶達失調引髮的疾病提供實驗基礎。方法製備重組人Tim-3蛋白，常規方法免疫BALB/c小鼠，篩選齣暘性剋隆，併對其識彆、中和活性進行體內外驗證，探討其作為免疫榦預手段的可行性。結果①穫得一株能夠分泌具有較好結閤活性的抗人Tim-3單剋隆抗體細胞株（剋隆號L3D），其分泌的免疫毬蛋白亞型為IgG2a；②流式實驗結果錶明，該抗體能夠結閤人U937細胞上的Tim-3分子，且與小鼠RAW 264．7細胞上Tim-3有一定的交扠結閤活性；③該抗體能夠阻斷Gal-9分子誘導的人THP1細胞凋亡，具有良好的體外中和活性；④體內註射L3D，顯示該抗體能夠顯著加重膿毒血癥模型小鼠病情，提高炎性因子的錶達水平，顯示齣其良好的體內中和活性。結論成功製備一株分泌抗人Tim-3抗體的細胞株，其良好的結閤及中和活性為基于Tim-3錶達異常相關疾病的榦預提供瞭實驗基礎。
목적：제비항인Tim-3단항병평개기생물학활성，위간예Tim-3표체실조인발적질병제공실험기출。방법제비중조인Tim-3단백，상규방법면역BALB/c소서，사선출양성극륭，병대기식별、중화활성진행체내외험증，탐토기작위면역간예수단적가행성。결과①획득일주능구분비구유교호결합활성적항인Tim-3단극륭항체세포주（극륭호L3D），기분비적면역구단백아형위IgG2a；②류식실험결과표명，해항체능구결합인U937세포상적Tim-3분자，차여소서RAW 264．7세포상Tim-3유일정적교차결합활성；③해항체능구조단Gal-9분자유도적인THP1세포조망，구유량호적체외중화활성；④체내주사L3D，현시해항체능구현저가중농독혈증모형소서병정，제고염성인자적표체수평，현시출기량호적체내중화활성。결론성공제비일주분비항인Tim-3항체적세포주，기량호적결합급중화활성위기우Tim-3표체이상상관질병적간예제공료실험기출。
Objective To develop a human Tim-3 specific monoclonal antibody and evaluate its biological activity and possible use in clinical diseases associated with dysregulated Tim-3 expression .Methods The BALB/c mice were immu-nized by conventional method, and positive clones were used to develop anti-human Tim-3 antibody, the binding and neutralization activities of which in vitro and in vivo were investigated.Results ①A monoclonal antibody (clone L3D) which could specifically bind to human Tim-3 protein in ELISA assay was obtained and the subtype of the monoclonal antibody was IgG2a .②Flow cytometry indicated that the monoclonal antibody could bind to Tim-3 expressed in human U937 cells.This antibody also showed a cross activity to mice′Tim-3.③The monoclonal antibody inhibited the apoptosis of THP1 cells induced by Gal-9, the ligand of Tim-3.④Injection of Tim-3 antibody exacerbated sepsis in mice as marked by the decreased survival rate and increased expression of pro-inflammatory cytokines .Conclusion An anti-human Tim-3 monoclonal antibody is successfully obtained.The excellent binding and neutralization activities of this antibody enable it to be widely used in clinical diseases associated with deregulated Tim-3 expression .
