Drug Delivery and Translational Research ( IF 5.7 ) Pub Date : 2022-07-08 , DOI: 10.1007/s13346-022-01207-5 Anne Marzi 1 , Kai Moritz Eder 1 , Álvaro Barroso 1 , Ane Marit Wågbø 2 , Ýrr Mørch 2 , Anne Rein Hatletveit 2 , Torkild Visnes 2 , Ruth B Schmid 2 , Geir Klinkenberg 2 , Björn Kemper 1 , Jürgen Schnekenburger 1
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State-of-the-art in vitro test systems for nanomaterial toxicity assessment are based on dyes and several staining steps which can be affected by nanomaterial interference. Digital holographic microscopy (DHM), an interferometry-based variant of quantitative phase imaging (QPI), facilitates reliable proliferation quantification of native cell populations and the extraction of morphological features in a fast and label- and interference-free manner by biophysical parameters. DHM therefore has been identified as versatile tool for cytotoxicity testing in biomedical nanotechnology. In a comparative study performed at two collaborating laboratories, we investigated the interlaboratory variability and performance of DHM in nanomaterial toxicity testing, utilizing complementary standard operating procedures (SOPs). Two identical custom-built off-axis DHM systems, developed for usage in biomedical laboratories, equipped with stage-top incubation chambers were applied at different locations in Europe. Temporal dry mass development, 12-h dry mass increments and morphology changes of A549 human lung epithelial cell populations upon incubation with two variants of poly(alkyl cyanoacrylate) (PACA) nanoparticles were observed in comparison to digitonin and cell culture medium controls. Digitonin as cytotoxicity control, as well as empty and cabazitaxel-loaded PACA nanocarriers, similarly impacted 12-h dry mass development and increments as well as morphology of A549 cells at both participating laboratories. The obtained DHM data reflected the cytotoxic potential of the tested nanomaterials and are in agreement with corresponding literature on biophysical and chemical assays. Our results confirm DHM as label-free cytotoxicity assay for polymeric nanocarriers as well as the repeatability and reproducibility of the technology. In summary, the evaluated DHM assay could be efficiently implemented at different locations and facilitates interlaboratory in vitro toxicity testing of nanoparticles with prospects for application in regulatory science.
Graphical abstract
中文翻译：
基于数字全息显微镜的聚合物纳米载体无标记体外细胞毒性测试的实验室间评估
用于纳米材料毒性评估的最先进的体外测试系统基于染料和几个可能受纳米材料干扰影响的染色步骤。数字全息显微镜 (DHM) 是一种基于干涉测量的定量相位成像 (QPI) 变体，有助于对天然细胞群进行可靠的增殖量化，并通过生物物理参数以快速、无标记和无干扰的方式提取形态特征。因此，DHM 已被确定为生物医学纳米技术中细胞毒性测试的通用工具。在两个合作实验室进行的一项比较研究中，我们利用互补的标准操作程序 (SOP) 研究了 DHM 在纳米材料毒性测试中的实验室间变异性和性能。两个相同的定制离轴 DHM 系统，为生物医学实验室使用而开发，配备舞台顶部孵化室，应用于欧洲的不同地点。与洋地黄皂苷和细胞培养基对照相比，观察到与两种聚（氰基丙烯酸烷基酯）（PACA）纳米颗粒变体孵育后 A549 人肺上皮细胞群的时间干质量发展、12 小时干质量增量和形态变化。作为细胞毒性控制物的洋地黄皂苷，以及空载和装载卡巴他赛的 PACA 纳米载体，同样影响了两个参与实验室的 A549 细胞的 12 小时干物质发育和增量以及形态。获得的 DHM 数据反映了测试纳米材料的细胞毒性潜力，并且与生物物理和化学测定的相应文献一致。我们的结果证实了 DHM 作为聚合物纳米载体的无标记细胞毒性测定以及该技术的可重复性和再现性。总之，评估的 DHM 测定可以在不同地点有效实施，并促进纳米粒子的实验室间体外毒性测试，具有在监管科学中的应用前景。
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