Analysis of shear stress due to shaking culture vessels or agitation of medium in plant cell culture

这项研究的目的是分析液体培养中细胞或细胞团受到的物理压力，这些压力是由于为细胞提供氧气或混合培养基而进行的搅拌或曝气造成的。采用的分析方法是将媒介的运动形象化并加以分析。所选择的培养容器为摇瓶和生物反应器，可视化方法采用示踪法，以尼龙颗粒为示踪剂。通过拍摄尼龙颗粒在短时间内的运动轨迹，直观地显示了液体的运动。当还需要流线方向的数据时，闪光镜发出光来标记流线的开始和结束。用计算机对得到的流线图像进行分析，计算出流线、旋转和剪切应力在流场中的分布。这些流动特性被选为与细胞的静水压力有关的流动特性。在摇瓶培养的情况下，使用激光光片(激光在薄平面上传播)来显示液体在液体中间的水平面上的运动。结果表明，该流型由高速区、中速区和低速区组成。图案本身根据烧瓶旋转而旋转。这些图案取决于旋转速度。随着转速的增加，旋转应力和剪应力绝对值的平均值增大。不同于摇床培养的情况，生物反应器中的液体运动是完全三维的，我们需要从两个角度同时拍摄两幅图像，在这种情况下，要求在两幅图像中识别相同的颗粒。我们正在努力克服这一困难。

The objectives of the study were to analyze the physical stress imposed on cells or cell aggregates in liquid culture caused by agitaion or aeration for supplying oxygen to cells or for mixing the medium. The adopted methods of analysis were to visualize the medium movement and to analyze it. The selected culture vessels for analysis were flasks for shake culture and bioreactors.The tracer method was adopted for visualization method, in which we utilized nylon particles for tracers. By photographing the trajectories of the nylon particles during a short period, the liquid movement was visualized. When data on streamline direction were also needed, light was emitted by a stroboscope to mark the beginning and end of the streamlines. The obtained images of streamlines were analyzed by a computer and distribution of streamlines, rotation and shear stress in the flow field were calculated. These flow characteristics were selected as ones relevant to hydrostatic stresses to the cells.In the case of the flask shake culture, a laser light sheet (laser light spread in thin plane) was used to visualize the liquid movement in a horizontal plane at the middle of the liquid. The results showed a flow pattern which consisted of high velocity region, moderate velocity region, and low velocity region. The pattern itself rotated in accordance with the flask rotation. The patterns were dependent on the rotation speed. With the increase of the rotation speed, the averages of the absolute values of rotation and shear stress increased. Unlike the case of shake culture, liquid movement in a bioreactor was fully three-dimensional, and we need two images taken simultaneously from two angles, In this case, identifying the same particle in the two images were requested. We are making efforts to overcome this difficulty.