Abstract：
Antibody drugs play a vital role in diagnostics and therapy. However, producing antibodies from mammalian cells is challenging owing to cellular heterogeneity, which can be addressed by applying droplet-based microfluidic platforms for high-throughput screening (HTS). Here, we designed an integrated system based on disulfide-bonded redox-responsive hydrogel beads (redox-HBs), which were prepared through enzymatic hydrogelation, to compartmentalize, screen, select, retrieve, and recover selected Chinese hamster ovary (CHO) cells secreting high levels of antibodies. Moreover, redox-HBs were functionalized with protein G as an antibody-binding module to capture antibodies secreted from encapsulated cells. As proof-of-concept, cells co-producing immunoglobulin G (IgG) as the antibody and green fluorescent protein (GFP) as the reporter molecule, denoted as CHO(IgG/GFP), were encapsulated into functionalized redox-HBs. Additionally, antibody-secreting cells were labeled with protein L-conjugated horseradish peroxidase using a tyramide amplification system, enabling fluorescence staining of the antibody captured inside the beads. Redox-HBs were then applied to fluorescence-activated droplet sorting, and selected redox-HBs were degraded by reducing the disulfide bonds to recover the target cells. The results indicated the potential of the developed HTS platform for selecting a single cell viable for biopharmaceutical production.
摘要：
抗体药物在诊断和治疗中起着至关重要的作用。然而,由于细胞异质性，从哺乳动物细胞中产生抗体具有挑战性，这可以通过应用基于液滴的微流体平台进行高通量筛选(HTS)来解决。这里,我们设计了一个基于二硫键氧化还原响应水凝胶珠（氧化还原-HBs）的集成系统，它们是通过酶水解制备的，划分，屏幕,选择,检索,并恢复分泌高水平抗体的选定中国仓鼠卵巢（CHO）细胞。此外，氧化还原-HBs用蛋白G功能化作为抗体结合模块以捕获从包封细胞分泌的抗体。作为概念证明，细胞共同产生免疫球蛋白G（IgG）作为抗体和绿色荧光蛋白（GFP）作为报告分子，表示为CHO（IgG/GFP），被封装到功能化的氧化还原-HBs中。此外,抗体分泌细胞用蛋白质L-结合辣根过氧化物酶使用酪胺扩增系统标记，能够对捕获在珠子内的抗体进行荧光染色。然后将氧化还原-HBs应用于荧光激活的液滴分选，选择的氧化还原-HBs通过还原二硫键来降解以回收靶细胞。结果表明所开发的HTS平台用于选择生物药物生产中可行的单细胞的潜力。
