# Radiation Damage of Polydimethylsiloxane and Polyimide by X-ray Free-Electron Laser
## 3 Results
### Observation of Radiation Damage on PDMS- and PI-Based Microfluidics Chips  
The SFX with the mix-and-inject technique is significant for elucidating the time-resolved molecular mechanism for interactions (or reactions) between the substrate (or inhibitor) and macromolecule crystals [55]. Various mix-and-inject systems have been developed, such as the injector [56; 57; 58] and conveyer belt [35] approach. Among them, the microfluidic method has been developed for time-resolved SX using synchrotron X-rays [32]. The advantage of this method is that the crystals and ligand solutions are mixed in the channel and precisely exposed to X-rays at the desired time. In addition, as in the sample delivery method using capillaries [30] or single-channel microfluidics [34], all samples pass through the inner channel aligned with the X-ray. In a previous serial synchrotron crystallography (SSX) experiment, synchrotron X-rays directed onto a microfluidic device made of PI, and no physical radiation damage was observed on the device [32]. Meanwhile, in the XFEL experiment, there has been no report of a case in which diffraction data were collected by directly exposing the microfluidic chip channel. Based on our results for XFEL-induced radiation damage on PDMS and PI films, we believed that a microfluidic device made of PDMS and PI would potentially be physically damaged at the position of  
Figure 3: Time-lapse snapshot of the PDMS film during continuous XFEL exposure. The verticallinear features developed around the beam transmission area, due to radiation damage, are indicated by red arrows.  
XFEL penetration. We expected the occurrence of large holes in PDMS-based microfluidics and leakage of crystals or solutions. In PI-based microfluidic devices, holes may occur in the channel area through which the XFEL passes; however, the hole size is small. Therefore, we expected that, due to surface tension, no leakage of the solution or crystals would occur.  
To demonstrate our hypothesis, we exposed a microfluidic device fabricated using PDMS and PI to XFELs during the flow of the solution. When a PDMS-based microfluidic chip was exposed to XFEL, a large hole (>100 \(\upmu\)m) was created at the XFEL penetration position, as for the PDMS film (Figure 4A). Moreover, we observed physical and structural distortions of the PDMS-based microfluidic device. Accordingly, the solution did leak out from the hole generated by the XFEL radiation damage in the microfluidic device. Consequently, the PDMS-based microfluidic device is considered to have no application in XFEL experiments where the material is exposed to the XFEL.  
For PI-based microfluidics, a hole was generated in the area through which the XFEL passed (Figure 4B). However, in the PI-based microfluidic device, unlike in the PDMS microfluidic chip, the flowing solution in the channel did not exit the hole generated by the XFEL. Instead, bubbles owing to radiation damage formed at the point exposed to XFEL (Figure 4B and Supplementary Video S4). Upon exposure to XFEL, these bubbles formed continuously and moved through the channel in the opposite direction to gravity, changing the flow path of the solution at the top. Continuous small bubbles accumulated to form large bubbles in the inner channel of the microfluidic device, and eventually, large bubbles were trapped inside the channel of the microfluidic device (Figure 4B and Supplementary Video S4). As these bubbles did not cover the entire area inside the channel, they were not washed out by the flowing solution. Consequently, these bubbles interfered with the path of the solution or crystal sample. This will be applicable for delivering solutions or crystal samples used in typical X-ray diffraction or scattering experiments. For performing a time-resolved SFX experiment with a mix-and-inject approach using microfluidics, the issue arises of microcrystals not being delivered at the desired time, owing to the change in the sample path.