文档介绍：新生大鼠心肌细胞原代培养方法的改良
【摘要】目的探讨更为简单、有效的新生大鼠心肌细胞分离、培养方法。方法取出生24 h内大鼠左心室,剪碎,%胰蛋白酶、%胶原酶Ⅰ分离,离心收集心肌细胞,差速贴壁法和化学试剂抑制非心肌细胞生长,纯化后培养于DMEM培养基。免疫荧光鉴定纯度,%台盼蓝染色检查心肌细胞成活率。结果心肌细胞纯度为95%,平均成活率96%,并出现同簇细胞的同步跳动。结论本研究应用改良的新生大鼠心肌细胞培养方法,心肌细胞存活率高,纯度高,且操作简便,重复性好,是一种较为理想的心肌细胞原代培养方法,可满足实验要求。
【关键词】心肌细胞原代培养胶原酶Ⅰ
Abstract: Objective To find an easy way to separate and culture the myocardial cells of rat neonate. Methods The left ventricular myocardium was removed from neonate rats within 24 hours after birth. The myocardial cells were separated by digestion with % trypsin and % collagenase I and collected by centrifugation. The myocardial cells were then cultured in DMEM and harvested by the time of adherence. The myocardial cells cultured were observed under light microscope, identified with cTnI and stained with % trypan blue to count the living cells. Results 95 percent of the cultured cells were myocardial cells and 96 percent of them were alive. All the myocardial cells were beating synchronously. Conclusion This is an effective way to obtain myocardial cells for scientific experimentation.
Key words: myocardial cell;primary culture;collagenaseⅠ
体外培养的心肌细胞具有自发节律性和收缩性特征,能够保持其在体内原有的许多结构和功能,同时排除了神经、体液等因素的干扰,可以从细胞、分子水平阐明一些基本理论,在心肌细胞生长发育、生理、代谢、病理等研究中具有重要作用。因此,心肌细胞原代培养技术已广泛应用于心血管疾病机理及心血管药物和心脏组织工程学的研究。自1960年Harary等[1]首次对Wister乳鼠的心肌细胞进行培养并成功地维持其自发性节律搏动长达40天以来,国内外许多学者对心肌细胞原代培养方法不断地进行研