Department of Bio and Brain Engineering, Korea Advanced Institute of Science and Technology (KAIST), 291 Daehak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.
Synthetic Biology and Bioengineering Research Center, Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 34141, Republic of Korea.
Lab Chip. 2021 Oct 26;21(21):4155-4165. doi: 10.1039/d1lc00581b.
Hanging drop plates and low-attachment well plates are suitable for a high throughput screening model of a spheroid, because each drop (or well) contains a single spheroid and the spheroid environment are separated from each other. However, uniform spheroid culture on these devices is difficult as the liquid around the spheroid is replaced by direct pipetting, which can cause spheroid damage or loss, and well-to-well variation. If spheroids need to be cultured for a long time or analyzed through chemical treatment of immunostaining, it becomes a more considerable problem as the number of pipetting action increases. To address these problems, we have developed a poly(dimethylsiloxane) (PDMS)-based drop array chip (DAC) and a pillar array chip (PAC) that can apply a droplet contact-based spheroid transfer (DCST) technique to multiple reagent change or washing steps of spheroid assays. Unlike previous DCST devices, 3D-printed mold-based DCST devices showed stable spheroid manipulation during repetitive drop contact and facile transfer of spheroid arrays to the next reagent-loaded DAC while minimizing cross-contamination of the reagents. Compared to the conventional manual or machine pipetting method, the DCST method showed lower user-to-user variation and a higher spheroid retention rate in the manipulation of the spheroid array. Live/dead staining, hypoxia staining, and immunofluorescence staining of the spheroid array were performed on a breast cancer cell line, BT-474. Furthermore, four clearing methods were applied to the spheroid array as a proof of concept, and we have identified the applicability of the DCST platform as a pretreatment platform for whole spheroid analysis.
悬滴板和低附着孔板适用于球体的高通量筛选模型，因为每个滴（或孔）包含一个单独的球体，并且球体环境彼此分离。然而，由于通过直接移液来替代球体周围的液体，这些设备上的球体均匀培养较为困难，这可能导致球体损坏或丢失，以及孔与孔之间的差异。如果需要长时间培养球体或通过化学处理免疫染色进行分析，随着移液操作次数的增加，这会成为一个更严重的问题。为了解决这些问题，我们开发了一种基于聚二甲基硅氧烷（PDMS）的液滴阵列芯片（DAC）和柱阵列芯片（PAC），可将基于液滴接触的球体转移（DCST）技术应用于球体分析的多个试剂更换或洗涤步骤。与以前的 DCST 设备不同，3D 打印模具基 DCST 设备在重复液滴接触过程中表现出稳定的球体操作，并且能够轻松地将球体阵列转移到下一个装有试剂的 DAC，同时最大限度地减少试剂的交叉污染。与传统的手动或机器移液方法相比，DCST 方法在球体阵列的操作中显示出更低的用户间差异和更高的球体保留率。对乳腺癌细胞系 BT-474 的球体阵列进行了死活染色、缺氧染色和免疫荧光染色。此外，还应用了四种透明化方法对球体阵列进行了概念验证，并且我们已经确定了 DCST 平台作为整体球体分析预处理平台的适用性。
