Recent progress with chemistry tools to deliver into living cells has seen a shift of attention from counterion-mediated uptake of cell-penetrating peptides (CPPs) and their mimics, particularly the Schmuck cation, toward thiol-mediated uptake with cell-penetrating poly(disulfide)s (CPDs) and cyclic oligochalcogenides (COCs), here exemplified by asparagusic acid. A persistent challenge in this evolution is the simultaneous and quantitative detection of cytosolic delivery and cytotoxicity in a high-throughput format. Here, we show that the combination of the HaloTag-based chloroalkane penetration assay (CAPA) with automated high-content (HC) microscopy can satisfy this need. The automated imaging of thousands of cells per condition in multiwell plates allows us to obtain quantitative data on not only the fluorescence intensity but also on the localization in a very short time. Quantitative and statistically relevant results can be obtained from dose–response curves of the targeted delivery to selected cells and the cytotoxicity in the same experiment, even with poorly optimized cellular systems.

中文翻译：

---

从Schmuck阳离子到最新的环状寡硫属化合物，细胞摄取的自动化高含量成像。

化学工具传递到活细胞中的最新进展已将注意力从抗衡离子介导的细胞穿透肽（CPPs）及其模拟物（尤其是Schmuck阳离子）的吸收转移到了硫醇介导的细胞穿透性聚二硫（二硫化物）的吸收）（CPD）和环状寡硫属元素化物（COC），此处以天冬氨酸为例。在这种发展过程中，一个持续的挑战是以高通量形式同时定量检测胞质传递和细胞毒性。在这里，我们表明基于HaloTag的氯代烷烃渗透测定（CAPA）与自动高含量（HC）显微镜的结合可以满足这一需求。多孔板中每种条件下成千上万个细胞的自动成像使我们不仅可以在很短的时间内获得关于荧光强度的定量数据，还可以获得关于定位的定量数据。即使在细胞系统优化不佳的情况下，也可以从目标递送至所选细胞的剂量反应曲线和细胞毒性中获得定量和统计学相关的结果。