# A Novel Method for Estimating the Dosage of Cold Atmospheric Plasmas in Plasma Medical Applications
## 2 Materials and Methods
### CAP Bio-Med Platform and Treatment of Cells  
As shown in Figure 2a, the CAP Bio-Med Platform employed in this study is a newly designed all-in-one device developed by the Plasma Health Scientech Group (PHSG) of the Tsinghua University of China. The upper part of the device has double doors to form a confined space. The UV lamp attached to the ceiling can be applied for atmosphere sterilization and disinfection. The CAP Bio-Med Platform is a multi-function device and mainly consists of three units:  
1. The CAP unit: It includes a co-axial type of atmospheric pressure dielectric barrier discharge (AP-DBD) plasma generator and the high-voltage alternating-current (HVAC) power supply module. As shown in Figure 2b, a tungsten wire confined inside a quartz tube works as the powered electrode, whereas a copper slice works as the \begin{table}
\begin{tabular}{c c c} \hline \hline  & **Materials** & **Sources of Procurement** \\ \hline \multirow{2}{*}{Cell line} & Human Embryonic Kidney & Cell Center of Peking Union Medical \\  & 293 (HEK293) cells & College, Beijing, China \\ \hline \multirow{6}{*}{Reagents} & Dulbecco’s Modified Eagle & Thermo Fisher Scientific Co., Ltd., \\  & Medium (DMEM) & Waltham, MA, USA \\  & Fetal bovine serum & Thermo Fisher Scientific Co., Ltd., \\  & Penicillin–Streptomycin & Waltham, MA, USA \\  & CO\({}_{2}\) & Beijing Praxair Practical Gas Co., Ltd., \\  & MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazol-bromide) solution & Beijing League Biotech Co., Ltd., \\  & Dimethyl sulfoxide (DMSO) & Beijing League Biotech Co., Ltd., \\ \cline{1-1} \cline{2-3}  & & Beijing, China \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Materials and their sources of procurement used in this study. grounded electrode. An acrylic tube is connected to the exit of the CAP generator to keep the treatment distance fixed at 2.0 cm and simultaneously protect the CAP jet from the entrainment of the surrounding air. The plasma jet shown in Figure 2c is uniform and stable with a gas temperature close to the room temperature, which meets the basic requirements of plasma medical applications. 2. The control unit: It includes the helium gas flow rate control module (0-20 slpm), the driving frequency control module (0-25 kHz), the discharge voltage control module (0-15 kV), and the discharge time control module (0-999 s). These control modules can ensure that the experimental parameters can be conveniently adjusted in a wide range. Moreover, the discharge will automatically stop when the pre-set discharge time is up. The HVAC power supply and the preceding control modules are integrated inside the lower part of the device as illustrated in Figure 2a. 3. The positioning unit: It includes a customized holder that supports the AP-DBD generator and the three-dimensional (3D) precision translational stages (LWX60-L200 and LWE4090, Misi Automation Equipment Co., Ltd., Dongguan, China). The CAP generator holder is used to fix the generator and to keep it vertical to the treated materials. When treating the cells, a 96-well plate can be put on the horizontal plane attached to the precision translational stage as shown in Figure 2a, so that the distance between the plasma jet exit and the upper surface of the 96-well plate can be adjusted precisely. For the cell treatment, the plasma dosages were varied by changing the treatment time and simultaneously keeping all the other parameters unchanged. The discharge frequency was fixed at \(f=23\) kHz and the pure helium (99.999%) gas flow rate was \(Q=8.0\) slpm. The amplitude of the discharge voltage was fixed to be \(V_{\text{d}}=3.5\) kV measured by a digital oscilloscope (DPO4034,  
Figure 2: **(a)** Picture of the CAP Bio-Med Platform; (b) schematic of the geometrical configuration of the CAP generator; and (c) typical image of the CAP jet. Tektronix, Beaverton, OR, USA) and a voltage probe (P6015A, Tektronix, Beaverton, OR, USA). Under such conditions, the free CAP jet was approximately 5.0 cm long, determined by a visible image processing technique [27; 28].