Heracell co2 incubator

The cell culture of rat C6 gliomas was obtained from the Russian Collection of Cell Cultures, Institute of Cytology, Russian Academy of Sciences (Saint-Petersburg, Russia). C6 rat glioma cells were cultured at 2.5 × 10⁵/mL in Petri dishes (d = 35 mm, Nunc, Allerod, Denmark) or 96-well flat-bottomed plates (TPP, Geneva, Switzerland, 1 × 10⁴/well) in Dulbecco’s Modified Eagle Medium (DMEM, Sigma-Aldrich, Darmstadt, Germany) containing 10% fetal bovine serum (Sigma-Aldrich) and 10⁻⁴ g/mL gentamicin sulfate (Shandong Weifang Pharmaceutical Factory Co., Liaocheng, China) in a Heracell CO₂ incubator (Thermo Fisher, Saint-Petersburg, Russia) at 37 °C with 95% humidity and 5% CO₂ for 1–2 days [44 ]. Fibroblast cell culture was obtained from the cell bank of Pokrovskaya Hospital (Saint-Petersburg, Russia). Human fibroblasts were cultured at 96-well flat-bottomed plates (TPP, 2 × 10⁴/well) in DMEM containing 10% fetal bovine serum and penicillin streptomycin 10⁻⁴ g/mL in a CO₂ incubator at 37 °C, 95% humidity and 5% CO₂ for 2 days [45 (link)].

Cells exposed to light were cultivated in a Multitron incubator with CO₂ supply (Infors HT, Switzerland). For white light, the built-in LED lights were used for a homogenous illumination from above, resulting in a light intensity of 80 µmol m⁻² s⁻¹ (2 mW/cm²) at the height of the cell layer. This intensity was chosen to be as low as possible to minimize the impact on mammalian cells while still allow photosynthetic oxygen production. Red and blue light were provided by an LED strip (LLV-shop, Germany), also providing homogenous illumination from above in the same intensity (measured in µmol m⁻¹ s⁻¹). Samples cultivated in darkness were kept in a HERAcell CO₂ incubator (Thermo Fisher Scientific). Standard cell culture conditions (37 °C, 5% CO₂) were applied in both incubators. Light spectra were measured using an LI-180 spectrometer (LI-COR Biosciences GmbH, Germany) and are shown in Fig. 1a.

Worms were isolated from umbilical areas of infected cattle skin as previously described by Cho-Ngwa et al. [11 (link)]. Briefly, cattle skin containing palpable nodules obtained from the butchery in Douala Cameroon were washed with soap and rinsed with distilled water. The skin was then sterilized with 70% ethanol after which the nodules were carefully opened and the entire nodular content removed and submerged in 2 mL of complete culture medium (CCM) comprising of RPMI-1640 (SIGMA cat: R0883), supplemented with 25 mM HEPES, 2 g/L sodium bicarbonate, 2 mM L-glutamine, 5% heat inactivated new born calf serum (SIGMA Cat: N4762), 200 units/mL penicillin/ 200 μg/mL streptomycin and 0.25 μg/mL amphotericin B, pH 7.4 in 12-well culture plates. The worms were left in cultures in a HERACELL-CO₂ incubator (Thermo Fisher, UK) overnight and checked for any contamination before drugs were added.

Human skin fibroblasts were cultured in DMEM supplemented with 10% fetal bovine serum (FBS) and 1% penicillin‐streptomycin. The cells were incubated in a 37°C Heracell CO₂ incubator (Thermo Fisher Scientific Co., Ltd.) at 5% CO₂, and the medium was changed every 2 days. All tests were performed on cells obtained between 4th and 6th passages.
Next, 100 μl of the HSF cell suspension (8 × 10⁴–1 × 10⁵ cells/ml) was added to each well of a 96‐well plate. After 12 h, the culture medium was replaced with the DOP and FDOP in sample wells and UVA control wells, respectively, and the plates were irradiated at 5 mW/cm2 under a UVA lamp (Spectronics Corporation, Ltd) for 2 h. The cells were then incubated in a constant temperature and humidity incubator at 37°C and 5% CO₂ for 24 h.