Department of Biomedical Engineering, University of Rochester, Rochester, NY, USA.
Biomed Microdevices. 2013 Jun;15(3):453-63. doi: 10.1007/s10544-013-9745-0.
Development of micro-well array systems for use in high-throughput screening of rare cells requires a detailed understanding of the factors that impact the specific capture of cells in wells and the distribution statistics of the number of cells deposited into wells. In this study we investigate the development of microbubble (MB) well array technology for sorting antigen-specific B-cells. Using Poisson statistics we delineate the important role that the fractional area of MB well opening and the cell seeding density have on determining cell seeding distribution in wells. The unique architecture of the MB well hinders captured cells from escaping the well and provides a unique microenvironmental niche that enables media changes as needed for extended cell culture. Using cell lines and primary B and T cells isolated from human peripheral blood we demonstrate the use of affinity capture agents coated in the MB wells to enrich for the selective capture of B cells. Important differences were noted in the efficacy of bovine serum albumin to block the nonspecific adsorption of primary cells relative to cell lines as well as the efficacy of the capture coatings using mixed primary B and T cells samples. These results emphasize the importance of using primary cells in technology development and suggest the need to utilize B cell capture agents that are insensitive to cell activation.
开发用于高通量筛选稀有细胞的微孔阵列系统需要详细了解影响细胞在孔中特异性捕获的因素，以及细胞在孔中沉积数量的分布统计。在这项研究中，我们研究了用于分选抗原特异性 B 细胞的微泡（MB）孔阵列技术的开发。我们使用泊松统计方法阐明了 MB 孔开口的分数面积和细胞接种密度对确定细胞在孔中接种分布的重要作用。MB 孔的独特结构阻止了被捕获的细胞从孔中逃逸，并提供了一个独特的微环境小生境，使细胞培养过程中能够根据需要进行介质更换。我们使用细胞系和从人外周血中分离的原代 B 和 T 细胞，证明了使用涂覆在 MB 孔中的亲和捕获剂来富集选择性捕获 B 细胞的用途。与细胞系相比，牛血清白蛋白在阻止原代细胞非特异性吸附方面的功效以及使用混合原代 B 和 T 细胞样本的捕获涂层的功效存在重要差异。这些结果强调了在技术开发中使用原代细胞的重要性，并表明需要使用对细胞激活不敏感的 B 细胞捕获剂。
