## 6 Viability of A549, A431 and HEK 293 Cells after CAP Jet Treatment  
In the indirect treatment of cells with the irradiated medium the various sensitivity of different types of cells was observed. The human cell lines, A431 and HEK293 were seeded in 96-well flat-bottom plates at a density of 5000-10,000 cells in 100 uL of DMEM F12 media per well. The same cell-free DMEM F12 media in 12-well plates was exposed to the argon CAP jet during 5 min, 10 min and 15 min at \(U\) = 4.9 kV and v = 4 L/min. The indirect treatment of cells with the irradiated medium was carried out according to the scheme: 1:1, 50% of the initial medium and 50% of the medium treated with plasma. Therefore, 100 uL of treated media was added to each of 96-well flat-bottom plates with cells 15 min after the plasma treatment. To analyze the effect, MMT-Assay of Cell Viability was applied after the treatment. The viability of A431 cells and HEK 293 cells shown in Figure 12 was quantified 24 h after the media was added. A greater than 90% reduction in A431 cell viability was registered for the case of indirect treatment with the media exposure to the argon CAP jet during 5 min (see Figure 12a). For the cases of 10 min and 15 min, A431 cells viability is almost the same. In contrast to A431 cellsviability, the HEK 293 cells shown in Figure 12b were less sensitive to the indirect CAP treatment. Indeed, the data showed 80% live cells after the treatment with the media exposed to CAP jet during 5 min. For the cases of 10 min and 15 min, the viability of HEK 293 cells decreases to 40% and 38%, respectively. The direct treatment with the CAP jet was carried out with A549 cells. In the experiment, A549 human lung adenocarcinoma cells were exposed to argon and helium CAP jets for different voltages, working gas flow rates and with and without the presence of the grounded substrate beneath the plates with the media and cells. MTT assay results in Figure 13 show the viability of A549 cells 24 and 48 h after the direct argon CAP treatment with and without the grounded substrate. The cells were exposed to the plasma jet during 1 min at \(U=3.6\), 4.2 and 4.9 kV. For a lower voltage plasma treatment, at \(U=3.6\) kV, 60% cells survive 24 h after the treatment and approximately 35% after 48 h for both cases. So, no visible effect of the grounded substrate was registered. However, at higher voltages, at \(U=4.2\) kV and 4.9 kV, the use of the grounded substrate significantly enhances the effect of plasma treatment. In this case, the number of survivals drops to almost zero, whereas without the grounded substrate the statistics of cell viabili  
Figure 12: Live cells in % 24 h after the indirect argon CAP treatment of A431 human skin carcinoma (**a**) and HEK 293 human embryonic cells (**b**). Irradiation of media was 5, 10 and 15 min with CAP, without the grounded substrate, at \(U=4.9\) kV and v = 4 L/min. Figure 13: Number of survivals of A549 cells in % for cases with and without the grounded substrate after 24 and 48 h of direct argon CAP treatment, 1 min, at \(U=3.6\), 4.2 and 4.9 kV, gas flow rate v = 4 L/min. all voltages \(U\) = 3.6, 4.2 and 4.9 kV. It is seen that for the case of an electrically isolated target an increase of the input power in the plasma device does not enhance an efficiency of the plasma treatment. The cell viability was also monitored with iCELLigence instrument to quantify cell proliferation. 30,000 cells were planted per a well of an iCELLigence plate the day before the experiment. A549 cells in an 8-well E-plates were treated for 2 min with argon and helium CAP jets. Taking different parameters of CAP, we wanted to show that the treatment does not work if the plasma parameters are chosen without optimization. First we made experiment with helium plasma jet parameters which are far from the optimal values found in Section 3, \(U\) = 4.2 kV and \(\
u\) = 5 L/min. For argon plasma jet the parameters of the plasma source \(U\) = 3.6 kV and \(\
u\) = 3 L/min.