Figure 2 | Scientific Reports

Figure 2

From: Surpassing light-induced cell damage in vitro with novel cell culture media

Figure 2

In vitro photo-toxicity can be resolved with a new culture medium. (a) Setup of experiment to test media for photo-reactive components. Step (i) media is treated ± light stimulation (W = 0.3 mW/mm2, t = 5 ms and f = 1 Hz) for 20 hours delivering 108 kJ/m2 of light, step (ii) after which viable cells are placed in ± pre-irradiated media for 24 hours before step (iii) analysis by PI exclusion assay. (b) Viabilities of OPCs 24 hours after placement into DMEM + SATO treated ± light (108 kJ/m2). Note: the robust loss of OPC viability in pre-irradiated media is similar to irradiating cells directly with light (see Fig. 1h). (c and d) Significant improvement (p = 0.0021) of OPC viability in photo-inert media MEMO + SATO compared to DMEM + SATO when treated with light (180 kJ/m2), but both conditions retain significant loss of viabilities compared to cells kept in dark (p = 0.0003 and p = 0.022 respectively), and addition of riboflavin (Rb.), a component absent in MEMO, restores highly significant loss (p = 0.0005) of OPC viability when treating cells with light. (e and f) Increasing light dose to 360 kJ/m2 induces robust loss of OPC viability in MEMO + SATO, however, combining MEMO + SOS protects OPC viabilities during light doses of 360 kJ/m2 and up to 2160 kJ/m2 (W = 0.6 mW/mm2, t = 0.5 ms and f = 10 Hz), a dose 20 fold greater than the 108 kJ/m2 dose that induced almost complete loss of OPC viability in standard conditions of DMEM + SATO (see Fig. 1h). All above histograms are normalized to controls with data representing means ± s.e.m. of a number of biological replicates (value within or above each histogram) and p values from Student’s two tailed unequal variance t-test. Black and blue histograms represent conditions kept in the dark or exposed to light respectively. Light doses in kJ/m2 are shown above histograms and as insets within representative images of irradiated experiments. Cell culture media conditions are described above images and under respective histograms.

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