Microfabricated modular scale-down device for regenerative medicine process development.
Microfabricated modular scale-down device for regenerative medicine process development.
复制标题
DOI：
10.1371/journal.pone.0052246
复制
发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Szita N
中科院分区：
文献类型：
--
作者：
Reichen M;Macown RJ;Jaccard N;Super A;Ruban L;Griffin LD;Veraitch FS;Szita N
毫升和微升生物反应器加速工艺开发的能力已在传统生物技术中得到成功证明。然而，对于目前的再生医学来说，较小规模的培养方法不能处理需要评估的广泛的处理变量。现有的微制培养设备可以用最少的资源(例如昂贵的培养介质)来测试不同的培养变量，通常在设计时没有考虑到工艺开发。我们提出了一种新的、可高压灭菌的、微细加工的设备，用于再生医学过程的开发。这种微型制造的设备包含一个可重新密封的培养室，便于使用标准培养方案，创建了与传统小规模培养设备的联系，用于验证和放大研究。此外，模块化设计可以很容易地适应不同培养底物/细胞外基质组合的研究。采用标准的静态接种方法，将灭活的小鼠胚胎成纤维细胞(IMEF)和人胚胎干细胞(HESC)克隆接种于明胶包被的组织培养聚苯乙烯(TC-PS)上。该设备中包含的微流控芯片可对设备中的培养基流量和产生的剪应力进行精确和精确的控制。细胞培养2天，灌流浓度为30 0µL·H-−-1的培养液，模拟切应力为1 1×10−4Pa.在灌流后，hESC克隆对不同的多能性标志物呈阳性染色，并保持未分化的形态。开发了一种图像处理算法，可以从相差显微镜图像中定量共培养的集落形成细胞。在不到45秒的高分辨率图像中，根据饲养层细胞(IMEF)的背景对hESC的集落大小进行了量化，这将允许在未来的实验中实时监控培养过程。该装置是利用微流控技术的优势进行再生医学过程开发的第一步。
The capacity of milli and micro litre bioreactors to accelerate process development has been successfully demonstrated in traditional biotechnology. However, for regenerative medicine present smaller scale culture methods cannot cope with the wide range of processing variables that need to be evaluated. Existing microfabricated culture devices, which could test different culture variables with a minimum amount of resources (e.g. expensive culture medium), are typically not designed with process development in mind. We present a novel, autoclavable, and microfabricated scale-down device designed for regenerative medicine process development. The microfabricated device contains a re-sealable culture chamber that facilitates use of standard culture protocols, creating a link with traditional small-scale culture devices for validation and scale-up studies. Further, the modular design can easily accommodate investigation of different culture substrate/extra-cellular matrix combinations. Inactivated mouse embryonic fibroblasts (iMEF) and human embryonic stem cell (hESC) colonies were successfully seeded on gelatine-coated tissue culture polystyrene (TC-PS) using standard static seeding protocols. The microfluidic chip included in the device offers precise and accurate control over the culture medium flow rate and resulting shear stresses in the device. Cells were cultured for two days with media perfused at 300 µl.h−1 resulting in a modelled shear stress of 1.1×10−4 Pa. Following perfusion, hESC colonies stained positively for different pluripotency markers and retained an undifferentiated morphology. An image processing algorithm was developed which permits quantification of co-cultured colony-forming cells from phase contrast microscope images. hESC colony sizes were quantified against the background of the feeder cells (iMEF) in less than 45 seconds for high-resolution images, which will permit real-time monitoring of culture progress in future experiments. The presented device is a first step to harness the advantages of microfluidics for regenerative medicine process development.
登录
查看更多内容
影响因子：
2.9
作者：
Aunins, JG;Bader, B;Zhou, WC
通讯作者：
Zhou, WC
影响因子：
2.8
作者：
Korin, Natanel;Bransky, Avishay;Levenberg, Shulamit
通讯作者：
Levenberg, Shulamit
DOI：
10.1126/science.1171643
发表时间：
2009-06-26
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Discher DE;Mooney DJ;Zandstra PW
通讯作者：
Zandstra PW
影响因子：
3.8
作者：
Korin, Natanel;Bransky, Avishay;Levenberg, Shulamit
通讯作者：
Levenberg, Shulamit
影响因子：
6.1
作者：
Kamei, Ken-ichiro;Guo, Shuling;Tseng, Hsian-Rong
通讯作者：
Tseng, Hsian-Rong