Contact Microscopy using a Compact Laser Produced Plasma Soft X-Ray Source

Soft X-ray contact microscopy potentially allows imaging of wet living biological specimens at a spatial resolution higher than optical microscopy and without many of the constraints of scanning electron microscopy. In this paper, we present the development of a laboratory scale contact microscope that uses a laser produced plasma soft X-ray source. The source is based on a double-stream gas-puff target approach and it operates in the "water window" spectral range which enables to capture images of biological samples with a natural contrast. In the preliminary experiments the contact microscope system has been used for imaging of fixed and dried non-malignant HCV29 human bladder cell lines cultured on polymethyl methacrylate photoresists. The samples were exposed with 150 pulses of soft X-rays as an initial test to demonstrate the possibility of image formation. The soft X-ray contact images registered in the photoresists exhibit high resolution in the atomic force microscopy topography which indicates the potential application of soft X-ray contact microscopy in life science to examine small features as small as few tens of nm. The technique could also be used for living cell imaging with further optimization of the microscope system and development of a special specimen holder. The details of the soft X-ray contact microscopy technique and the experimental results are presented and discussed

Differentiation between single bladder cancer cells using principal component analysis of time-of-flight secondary ion mass spectrometry

Time-of-flight-secondary ion mass spectrometry (TOF-SIMS) mass spectra measurements combined with an appropriate sample preparation protocol are the powerful tools to obtain unique information about the chemical composition of biological materials. In our studies, two questions were addressed, i.e., whether it is possible to develop a fixative-based sample preparation protocol and whether it allows one to distinguish between cells originating from various stages of cancer progression. Therefore, four human bladder cancer cell lines (with distinct malignancy degree) have been investigated. A chemical fixation protocol has been used for TOF-SIMS measurements, and mass spectra were obtained using a Bi_{3}+ primary ion beam. The principal component analysis (PCA) has been applied to analyze the whole range of mass spectra (without preselection of any particular masses) using two approaches of data preprocessing, namely, mean centering and autoscaling. The PC3 versus PC2 plot has showed significant differences between nonmalignant cancer cells and the cancerous ones for both of preprocessing approaches. The analysis of mass spectra of human bladder cells allows one to find a list of mass peaks with intensities significantly larger in cancerous bladder cells compared to nonmalignant cell cancer of the ureter (HCV29 cells). These findings show that TOF-SIMS in combination with PCA can be used to identify reference, human bladder cells from cancerous ones