Lipophilic rather than hydrophilic photosensitizers show strong adherence to standard cell culture microplates under cell-free conditions

Analysis of photosensitizer (PS) uptake kinetics into tumor cells is a standard cell culture experiment in photodynamic therapy (PDT) – usually performed in plastic microplates or petri dishes. Organic substances such as PS can potentially interact with the plastic surfaces. In this study, we provide a qualitative comparison of three lipophilic PS (hypericin, Foscan® and Photofrin®) and two rather hydrophilic PS formulations (PVP-hypericin and aluminum (III) phthalocyanine tetrasulfonate chloride) regarding their adherence to the surfaces of 96-well microplates obtained from four different manufacturers. For estimation of the relevance of PS adherence for cellular uptake studies we compared the fluorescence signal of the respective PS in microplates containing A431 human epithelial carcinoma cells with microplates incubated with the respective PS under cell-free conditions. We demonstrate that lipophilic PS substances show a strong adherence to microplates – in case of direct lysis and fluorescence measurement resulting in 50% up to 90% of the overall signal to be caused by adherence of the substances to the plastic materials in a cellular uptake experiment. For the hydrophilic compounds, adherence is negligible. Interestingly, adherence of PS agents to microplates takes place in a time-dependent and thus kinetic-like manner, requiring up to several hours to reach a plateau of the fluorescence signal. Furthermore, PS adherence is a function of the PS concentration applied and no saturation effect was observed for the concentrations used in this study. Taken together, this study provides a systematic analysis under which conditions PS adherence to cell culture plates may contribute to the overall fluorescence signal in – for example – PS uptake experiments.

► We studied six photosensitizers (PS) for cell-free adherence to cell culture plates.
► In absence of serum, lipophilic but not hydrophilic PSs show strong adherence.
► PS adherence to microplates is a function of the time and concentration applied.
► In cell studies adherence may account for up to 90% of the overall PS fluorescence.
► Cellular PS uptake studies must consider adherence to cell culture material.