## 4 Discussion  
The use of cold physical plasma to selectively target cancer cells has been a topic of interest in recent years [23; 24; 25]. Our study provides further evidence of its potential effectiveness by investigating physical plasma's effects in tongue cancer and breast cancer cell lines and found a clear dose-dependent reduction in cell viability in all cell lines tested. The IC50 values for physical plasma exposure indicate that even relatively short exposure times can induce significant cytotoxicity in cancer cells. Several studies have investigated physical plasma effects on various cancer cell types, including brain, lung, blood, cervical, melanoma, breast, colorectal, head and neck, hepatocellular, prostate, ovarian, osteosarcoma, pancreatic, bladder, gastric, thyroid, uterine, and epidermal cancer cells [26]. These studies have shown promising results in inducing cancer cell death which is in line with our results. [PERSON] et al. investigated the effect of physical plasma treatment on triple-negative breast cancer cells, which are known to be particularly aggressive and difficult to treat. They found that physical plasma treatment was able to induce cell death in these cancer cells through ROS-dependent cellular pathways [27]. [PERSON] et al. investigated the effect of physical plasma treatment on triple-negative breast cancer cells. The researchers found that physical plasma treatment was able to induce cell death in these cancer cells, as well as inhibit their migration and invasion abilities both in vitro and in vivo [28]. [PERSON] et al. investigated the effect of physical plasma treatment on Oral squamous cell carcinoma and found that physical plasma treatment was able to inhibit cell growth in OSCC cells in a manner that is dependent on the dosage and triggers apoptosis by generating reactive oxygen and nitrogen species. The induced apoptosis is linked to an increase in Bax expression and a decrease in Bcl-2 expression. Additionally, plasma treatment of cancer cells was found to cause cell cycle arrest through the ATM/P53/P21 signaling pathway, and cellular senescence [29]. Novel treatment options need to be not only effective but also safe. One aspect of assessing safety is studying genotoxic effects [30]. We have previously established a high-throughput, high-content imaging micronucleus assay incorporating the requirements of OECD guidelines on the CBMN assay and utilizing the power of algorithm-driven object segmentation for unsupervised analysis and quantification of micronucleeli in binucleated cells [19]. Several studies have demonstrated that cold physical plasma-induced reactive oxygen species (ROS) are responsible for plasma-induced toxicity, have been associated with some hallmarks of cancer and are thought to spur malignancy and DNA damage when applied chronically at higher concentrations [31]. However, we could not find any genotoxic effects of gas plasma exposure in human HaCaT keratinocytes. These findings align with recent studies indicating that physical plasma did not cause DNA damage or abnormalities like micronucleeli [32; 33]. The absence of genotoxic effects in physical plasma-treated cells underline the favorable safety profile for human use. Our study also evaluated the potential toxicity of argon gas and ddH\({}_{2}\)O as negative controls and found no reduction in metabolic activity after 24 h. This suggests that the observed toxicity in the physical plasma experiments is attributed to ROS rather than gas flow. To investigate the effects of physical plasma on the patient-derived tumor samples, punch biopsies of BC and OSCC tissues were generated ex vivo and were investigated by immunofluorescence after cold plasma exposure. The results acquired from TUNEL staining demonstrate that physical plasma treatment significantly impacts cancerous tissue by inducing apoptosis.