Drug Discovery Science, Boehringer Ingelheim Pharma GmbH & Co. KG, Biberach an der Riss 88397, Germany.
Biotechniques. 2020 May;68(5):245-250. doi: 10.2144/btn-2020-0003. Epub 2020 Feb 21.
Phagocytosis is a fundamental mechanism of innate immunity and its impairment is associated with severe chronic diseases, for example, chronic obstructive pulmonary disease. Investigating phagocytosis requires flexible tools and assay conditions, such as different fluorescent particle types, detection colors and readouts. We comprehensively evaluated and optimized phagocytosis assays using particles labeled with fluorescent pH-sensitive pHrodo dyes, facilitating the specific detection of phagocytosed particles. Beads, bacterial and yeast particles labeled with pHrodo red and green were tested for their uptake by THP-1 cells and primary human macrophages by flow cytometry and high-content imaging. Whereas the latter allowed kinetic phagocytosis measurement, the former demonstrated the feasibility of using cell sorting for periods of up to 6 h, enabling downstream applications such as pooled genetic screens.
吞噬作用是先天免疫的一种基本机制，其功能障碍与严重的慢性疾病有关，例如慢性阻塞性肺疾病。研究吞噬作用需要灵活的工具和检测条件，例如不同类型的荧光颗粒、检测颜色和读数。我们使用荧光 pH 敏感 pHrodo 染料标记的颗粒全面评估和优化了吞噬作用检测，这有利于对吞噬颗粒的特异性检测。用 pHrodo 红色和绿色标记的珠子、细菌和酵母颗粒，通过流式细胞术和高内涵成像来检测 THP-1 细胞和原代人巨噬细胞的摄取情况。虽然后者允许进行动力学吞噬作用测量，但前者证明了使用细胞分选长达 6 小时的可行性，这使得可以进行 pooled genetic screens 等下游应用。
