Guldevall Karolin, Brandt Ludwig, Forslund Elin, Olofsson Karl, Frisk Thomas W, Olofsson Per E, Gustafsson Karin, Manneberg Otto, Vanherberghen Bruno, Brismar Hjalmar, Kärre Klas, Uhlin Michael, Önfelt Björn
Science for Life Laboratory, Department of Applied Physics, KTH - Royal Institute of Technology , Solna , Sweden.
Science for Life Laboratory, Department of Applied Physics, KTH - Royal Institute of Technology, Solna, Sweden; Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.
Front Immunol. 2016 Apr 5;7:119. doi: 10.3389/fimmu.2016.00119. eCollection 2016.
Here, we report a screening platform for assessment of the cytotoxic potential of individual natural killer (NK) cells within larger populations. Human primary NK cells were distributed across a silicon-glass microchip containing 32,400 individual microwells loaded with target cells. Through fluorescence screening and automated image analysis, the numbers of NK and live or dead target cells in each well could be assessed at different time points after initial mixing. Cytotoxicity was also studied by time-lapse live-cell imaging in microwells quantifying the killing potential of individual NK cells. Although most resting NK cells (≈75%) were non-cytotoxic against the leukemia cell line K562, some NK cells were able to kill several (≥3) target cells within the 12-h long experiment. In addition, the screening approach was adapted to increase the chance to find and evaluate serial killing NK cells. Even if the cytotoxic potential varied between donors, it was evident that a small fraction of highly cytotoxic NK cells were responsible for a substantial portion of the killing. We demonstrate multiple assays where our platform can be used to enumerate and characterize cytotoxic cells, such as NK or T cells. This approach could find use in clinical applications, e.g., in the selection of donors for stem cell transplantation or generation of highly specific and cytotoxic cells for adoptive immunotherapy.
在此，我们报告了一种筛选平台，用于评估较大群体中单个自然杀伤（NK）细胞的细胞毒性潜力。人原代NK细胞分布在一个硅玻璃微芯片上，该芯片包含32400个装有靶细胞的单个微孔。通过荧光筛选和自动图像分析，可以在初始混合后的不同时间点评估每个孔中NK细胞以及活靶细胞或死靶细胞的数量。还通过微孔中的延时活细胞成像研究细胞毒性，以量化单个NK细胞的杀伤潜力。尽管大多数静息NK细胞（约75%）对白血病细胞系K562无细胞毒性，但在长达12小时的实验中，一些NK细胞能够杀死多个（≥3个）靶细胞。此外，对筛选方法进行了调整，以增加发现和评估连续杀伤NK细胞的机会。即使不同供体的细胞毒性潜力有所不同，但很明显一小部分高细胞毒性NK细胞负责了大部分的杀伤作用。我们展示了多种检测方法，利用我们的平台可以对细胞毒性细胞，如NK细胞或T细胞进行计数和表征。这种方法可用于临床应用，例如在干细胞移植供体的选择或用于过继免疫治疗的高特异性和细胞毒性细胞的生成中。
