In vitro experiments using radiolabeled molecules is fundamental for Positron emission tomography (PET) or single photon emission computed tomography (SPECT) tracer development and various metabolic assays, but no consensus on appropriate incubation conditions exists. Specifically, the use of shaking versus non-shaking conditions, cell number to medium volume and the choice of cell plating material may unintentionally influence cellular oxygenation and medium composition. This is problematic when testing the oxygen-dependence of tracers including 18F-fluoro-2-deoxyglucose ([18F]FDG) and hypoxia-selective 2-nitroimidazoles (e.g., 18F-fluoroazomycin-arabinoside, [18F]FAZA) or when doing prolonged experiments. The purpose of this study was to assess the influence of various experimental conditions on tracer retention. Tumor cells were seeded in a) Glass or standard Polystyrene Petri dishes or as b) discrete droplets in polystyrene Petri dishes or on 9 mm glass coverslips positioned in glass Petri dishes. When confluent, cells were pre-equilibrated for 2 h to 21%, 0.5% or 0% O2 and [18F] FDG or [18F] FAZA was added, followed by cell harvest and analysis of radioactivity 1 h ([18F]FDG) or 3 h ([18F]FAZA) after. Experiments were conducted with/without orbital shaking. The influence of hypoxia on tracer retention varied widely among cell lines, but shaking-induced convection did not influence uptake. In contrast, hypoxia-driven [18F] FAZA, and to some extent [18F] FDG, retention was much lower in cells grown on polyethylene than glass. Scaling-down the number of cells did not compromise accuracy. Tracer retention was similar under stagnant and forced convection conditions suggesting that the former approach may be appropriate even when accurate control of oxygen and tracer availability is required. In contrast, conventional plasticware should be used with caution when studying tracers and drugs that are metabolized and retained or activated at low O2 levels. Downscaling of cell number, by reducing the effective growth area, was feasible, without compromising accuracy.

中文翻译：

---

[18F] FDG和[18F] FAZA保留的体外低氧反应性：振动与停滞条件，玻璃与聚苯乙烯基质和细胞数量缩小的影响。

使用放射性标记分子的体外实验是正电子发射断层扫描（PET）或单光子发射计算机断层扫描（SPECT）示踪剂开发和各种代谢测定的基础，但在合适的培养条件上尚无共识。具体而言，使用摇动与非摇动条件，细胞数至中等体积以及细胞铺板材料的选择可能会无意影响细胞氧合和培养基组成。当测试包括18F-氟-2-脱氧葡萄糖（[18F] FDG）和缺氧选择性2-硝基咪唑（例如18F-氟阿霉素-阿拉伯糖苷，[18F] FAZA）的示踪剂的氧依赖性时，或长时间进行测试时，这是有问题的实验。这项研究的目的是评估各种实验条件对示踪剂保留的影响。将肿瘤细胞接种在a）玻璃或标准聚苯乙烯陪替氏培养皿中，或作为b）聚苯乙烯陪替氏培养皿中的离散液滴或置于玻璃陪替氏培养皿中的9 mm玻璃盖玻片上。汇合后，将细胞预平衡2 h至21％，0.5％或0％O2，并添加[18F] FDG或[18F] FAZA，然后收获细胞并分析1 h的放射性（[18F] FDG）或3小时（[18F] FAZA）之后。在有/没有轨道摇动的情况下进行实验。缺氧对示踪剂保留的影响在细胞系中差异很大，但摇动引起的对流不影响摄取。相反，缺氧驱动的[18F] FAZA，在某种程度上[18F] FDG，在聚乙烯上生长的细胞中的保留率要比玻璃低得多。缩小像元数量不会影响准确性。示踪剂在停滞和强制对流条件下的保留情况相似，这表明即使需要精确控制氧气和示踪剂的可用性，前一种方法也可能适用。相反，当研究在低O2水平下被代谢并保留或激活的示踪剂和药物时，应谨慎使用常规塑料制品。通过减小有效生长面积来缩小细胞数量是可行的，而不会影响准确性。