Laser cell patterning is a distinctly effective technique for creating cell arrangements in culture that replicate in vivo tissue structure for studying contact-mediated cell-cell interactions. Conventional laser-based single-cell-manipulation techniques are limited by their inability to pattern irregularly shaped cells, such as rod-shaped cardiomyocytes. We report use of a spatial light modulator loaded with a computer-generated phase map to shape a single laser source into multiple laser-guidance beams distributed around the outer contour of an irregularly shaped cell to achieve accurate cell patterning. In addition to describing the principle and practice of the system design, we present what is to our knowledge the first achievement of patterning large, irregularly (rod) shaped adult rat cardiomyocytes in an end-to-end connected alignment to replicate the in vivo heart muscle cell connection without use of substrate surface modifications, which can interfere with in vivo-like cell-cell and cell-extracellular matrix interactions. Our research demonstrates that two-stage multiple beam laser guidance is effective: 1) A dual-beam configuration horizontally translates a cell from the outlet of the microfluidic cell delivery channel to a position above the cell deposition site and 2) A quad-beam configuration rapidly propels the cell axially through the suspension medium to the culture substrate with vertical movement of the cell patterning chamber. Our study reveals that 90% of the patterned cells maintained end-to-end connection 30 min after patterning, and mechanical junctions could be reinstalled between laser connected cells after overnight incubation. This demonstrates that multiple beam laser patterning is an outstanding tool for in vitro studying contact-mediated cell-cell interactions among irregularly shaped cells.

激光细胞图案化是在培养中创建可复制的细胞排列的独特有效技术 体内用于研究接触介导的细胞间相互作用的组织结构。常规的基于激光的单细胞操作技术由于无法对不规则形状的细胞（例如棒状心肌细胞）进行构图而受到限制。我们报告使用加载有计算机生成的相位图的空间光调制器，以将单个激光源整形为分布在不规则形状的单元的外部轮廓周围的多个激光引导光束，以实现精确的单元模式。除了描述系统设计的原理和实践之外，我们还提供了我们所知的首个以端对端连接排列方式复制大型，不规则（杆状）成年大鼠心肌细胞图案以复制模型的成就。体内心肌细胞连接而无需使用底物表面修饰，这可以干扰体内样细胞-细胞和细胞-细胞外基质的相互作用。我们的研究表明，两阶段多光束激光导引是有效的：1）双光束配置将细胞从微流细胞递送通道的出口水平转移到细胞沉积部位上方的位置，以及2）四光束配置随着细胞构图室的垂直运动，细胞迅速地通过悬浮介质轴向地推动细胞到达培养底物。我们的研究表明，图案化后30分钟，有90％的图案化细胞保持端对端连接，过夜孵育后，可以在激光连接的细胞之间重新建立机械连接。体外 研究形状不规则的细胞之间的接触介导的细胞间相互作用。