长型肌球蛋白轻链激酶4Ig结构域的表达及单克隆抗体的制备

长型肌球蛋白轻链激酶(long myosin light chain kinase,L-MLCK)及其分子内结构域的功能还不清楚.在研究其功能时,特异性抗体是一基本工具,但目前国际上还没有制备成功特异性较好的抗体.为此,我们先在大肠杆菌E.coliBL21中重组表达了鸡L-MLCK的4Ig蛋白片段(MLCK4Ig),蛋白经纯化后作为抗原免疫小鼠.在小鼠血清中检测到特异性抗体的基础上,建立了2株杂交瘤细胞株,获得抗鸡MLCK4Ig的单克隆抗体.该抗体可以特异与鸡L-MLCK和鸡MLCK4Ig反应.本文并对该抗体的特性及应用进行了讨论.结果表明,我们部分解决了L-MLCK的抗体缺乏问题,为研究L-MLCK及MLCK4Ig片段的功能提供了重要工具.

Expression and Monoclonal Antibody Preparation of 4Ig Fragment of Long Myosin Light Chain Kinase

The functions of long myosin light chain kinase(MLCK)and the multiple domains within the molecule were poorly understood.To our knowledge,there is no specific antibody against L-MLCK so far,which is essential for the study of L-MLCK.In this report,the 4Ig fragment of chicken L-MLCK(MLCK4Ig)was expressed by using of E.coli BL21 system,the purified protein was used as antigen to induce antibody.Specific antibody was detected in the sera of mouse injected with chicken MLCK4Ig.We further established two hybridoma cell lines producing monoclonal antibody,which specifically reacted with chicken MLCK4Ig or chicken L-MLCK.The characteristic and usefulness of the antibody were also discussed.The results showed that our work partially solved the problem of L-MLCK antibody deficiency,which provides an important tool for the functional investigation of L-MLCK and MLCK4Ig fragment.