Ultra-fast transient cell adhesion and its application for high-throughput microfluidic cell sorting

生物细胞利用其表面的各种黏附分子相互作用，并与环境相互作用。这些黏附分子的类型、数量和组合携带着关于特定细胞条件的重要信息，可以用来描述细胞的特征和诊断疾病。根据细胞黏附分子的表达来分选和分离细胞是许多生物医学分析中的关键步骤。目前的细胞分选方法使用与表达特定黏附分子的细胞结合的标签。这些附加的标签可以实现高纯度的分选，但标签可能会改变细胞状态，并可能导致不想要的细胞激活，阻止进一步使用标记的细胞。该项目将开发一种微流控技术，能够在不使用任何标签或标签的情况下，根据黏附分子的表达对生物细胞进行快速有效的分选。这项技术利用了一种带有周期性狭窄的微流体通道，上面覆盖着分子，这些分子在细胞表面短暂地与黏附分子相互作用。相互作用导致感兴趣细胞的轨迹改变，而不会引起不想要的激活。该项目将研究微通道内细胞相互作用的机制，并将探索使用这种微流控分离技术来分离具有高度特异性黏附分子的淋巴细胞，这些淋巴细胞可用于癌症治疗。这项研究将涉及本科生和研究生，该团队将向各级学生开展几项外展活动，包括开发科学和工程竞赛项目。该项目将开发一种基于分子表面标记对目标配体的亲和力的高通量、无标记细胞分选和分离的微流控方法。鉴定和分离表达所需分子表面标记的细胞在生物科学、细胞治疗和医学诊断中的各种应用中都是必需的。该项目将结合微流体实验和计算机模拟，在尚未探索的超快时间尺度上检查生物细胞的瞬时黏附。细胞将被推进通过装饰有对角脊的微流体通道，该通道略微压缩细胞，以促进黏附分子与覆盖微通道表面的配体之间的结合。结合事件改变了微通道中表征细胞黏附的细胞轨迹。细胞和微通道表面之间的短接触时间将防止不必要的细胞激活。该项目将在广泛的时间范围内系统地探索限制对瞬时细胞黏附的影响。此外，该项目将使用微流控细胞分选技术来检测T细胞对靶抗原的选择性，这在临床应用中是重要的，而不会激活细胞。将研究对T细胞新抗原的阳性和阴性选择，以鉴定疾病选择性T细胞。这一研究项目的结果将在医学诊断、治疗、细胞工程和细胞制造方面具有广泛的意义，是一种高通量无标记细胞分离的新方法。此外，微流控方法可以直接测量重要的生理配体和黏附细胞分子之间的瞬时相互作用，这将有助于开发新的诊断方法。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Biological cells use various adhesion molecules on their surfaces to interact with each other and their environment. The type, amount, and combination of these adhesion molecules, which carry important information about specific cell conditions, can be used to characterize cells and diagnose disease. Sorting and separating cells based on their expressions of adhesion molecules are critical steps in many biomedical assays. Current cell sorting approaches use tags that bind to cells expressing specific adhesion molecules. These attached tags enable high purity sorting, but the tags can alter cell state and can lead to unwanted cell activation preventing further use of the labeled cells. This project will develop a microfluidic technology that enables rapid and efficient sorting of biological cells based on the expression of adhesion molecules without the use of any tags or labels. The technology utilizes a microfluidic channel with periodic constrictions coated with molecules that briefly interact adhesion molecules on the cells' surfaces. The interaction causes a change in the trajectories of the cells of interest without inducing unwanted activation. The project will investigate the mechanics of cell interactions within the microchannel and will probe the use of this microfluidic separation technique to isolate lymphocytes with highly specific adhesion molecules that can be used in cancer therapies. The research will involve undergraduate and graduate students, and the team will conduct several outreach activities to students at all levels, including developing projects for science and engineering competitions.This project will develop a microfluidic approach for high-throughput, label-free cell sorting and separation based on the affinity of molecular surface markers for target ligands. Identifying and isolating cells that express desired molecular surface markers are required in a variety of applications in the biological sciences, cell therapy, and medical diagnostics. The project will integrate microfluidic experiments and computer simulations to examine transient adhesion of biological cells at ultrafast time scales that have not yet been explored. Cells will be propelled through a microfluidic channel decorated with diagonal ridges that slightly compress the cells to promote binding between adhesion molecules and ligands covering the microchannel surfaces. The binding events alter cell trajectories in the microchannel characterizing cell adhesion. The short contact time between cells and microchannel surfaces will prevent unwanted cell activation. The project will systematically probe effects of confinement on transient cell adhesion for a wide range of time scales. Furthermore, the project will employ the microfluidic cell sorting technique to examine T cell selectivity to target antigens, which is important in clinical applications, without activating the cells. Positive and negative selection to T cell neoantigens will be investigated to identify disease-selective T cells. The results of this research project, a new microfluidic method for high-throughput label-free cell separation, will have broad implications in medical diagnostics, therapeutics, cell engineering, and cell manufacturing. Furthermore, the microfluidic method can enable direct measurements of transient interactions between important physiological ligands and adhesive cell molecules that will benefit the development of novel diagnostic methods.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.