# Imaging of Cell Structures Using Optimized Soft X-ray Contact Microscopy
## 4 Conclusions  
Soft X-ray contact microscopy may provide a powerful tool for studying cell structures. What distinguishes SXCM from other techniques used to visualize a biological sample (electron microscopy, super-resolution fluorescence microscopy) is the possibility of high-resolution imaging of internal cell structures without the need for the expensive and lengthy procedures of sample preparation. Optimizing the SXR exposure energy and PMMA development conditions affects the resolution and visibility of subcellular structures. In this work, good-quality images of biological cells were obtained using an easy to operate desk-top contact SXR microscopy system based on a double stream gas puff target. However, the energy used must be adapted directly to the cell line. We have demonstrated that more pulses are required to visualize internal structures in the DU-145 prostate carcinoma cells than for HCC38 breast cancer cells. The imaging conditions (the number of pulses, development conditions) should be adjusted adequately to the cell type and internal structure, which are to be observed. Therefore, if adjusted correctly and properly optimized, the contact soft X-ray microscopy can efficiently provide various information concerning cells' biology, complementing the other existing microscopy techniques. For further research, we plan to assemble a small compact  
Figure 8: Soft X-ray contact microscopy images of various cell lines: (**a**) DU145, (**b**) HCC38, and (**c**) hMSC. Images obtained with SXR exposure of 800 pulses. Cells fixed with 3.7% paraformaldehyde in phosphate-buffered saline (PBS). Results were collected using AFM (di CPII, Veeco) working in tapping mode. Scan size: \(80\times 80\) μm\({}^{2}\). Insets: fragments of cells showing detailed cellular structures (image size \(20\times 20\) μm\({}^{2}\)).  
device with an implemented fluorescence microscope. Such a device will identify individual cell structures imaged by SXCM with immunofluorescence methods with accurate specificity.  
The following are available online at [[http://www.mdpi.com/2076-3417/10/19/6895/s1](http://www.mdpi.com/2076-3417/10/19/6895/s1)]([http://www.mdpi.com/2076-3417/10/19/6895/s1](http://www.mdpi.com/2076-3417/10/19/6895/s1)), Figure S1: AFM images of DU145 cells saved on the PMMA photoresists by soft X-ray contact microscopy. Images were obtained after the exposure to 400 SXR pulses. Imprints were recorded by AFM (di CPII, Veeco) working in tapping mode. Scan size of \(25\times 25\) um\({}^{2}\). Insets: fragments of the cell showing detailed structures (image size \(5\times 5\) um\({}^{2}\)).  
Conceptualization, P.N.O., P.W. and A.N.-S.; methodology, P.N.O., P.W. and A.N.-S.; validation, A.B. and J.C.; investigation, P.N.O., P.W. and A.N.-S.; writing--original draft preparation, P.N.O., A.N.-S. and P.W.; writing--review and editing, E.A.T., M.L.; visualization, M.L., K.G., P.N.O. and A.N.-S.; supervision, E.A.T. and H.F.; funding acquisition, H.F. and P.W. All authors have read and agreed to the published version of the manuscript.  
This research was funded by the National Science Centre, Poland under the Beethoven Programme, grant number 2016/23/G/ST2/04319 and the European Union under the Horizon 2020 Programme Laserlab-Europe Project, grant number 871124.  
The authors declare no conflict of interest