Novel silica stabilization method for the analysis of fine nanocrystals using coherent X-ray diffraction imaging

High-energy X-ray Bragg coherent diffraction imaging (BCDI) is a well established synchrotron-based technique used to quantitatively reconstruct the three-dimensional morphology and strain distribution in nanocrystals. The BCDI technique has become a powerful analytical tool for quantitative investigations of nanocrystals, nanotubes, nanorods and more recently biological systems. BCDI has however typically failed for fine nanocrystals in sub-100 nm size regimes – a size routinely achievable by chemical synthesis – despite the spatial resolution of the BCDI technique being 20–30 nm. The limitations of this technique arise from the movement of nanocrystals under illumination by the highly coherent beam, which prevents full diffraction data sets from being acquired. A solution is provided here to overcome this problem and extend the size limit of the BCDI technique, through the design of a novel stabilization method by embedding the fine nanocrystals into a silica matrix. Chemically synthesized FePt nanocrystals of maximum dimension 20 nm and AuPd nanocrystals in the size range 60–65 nm were investigated with BCDI measurement at beamline 34-ID-C of the APS, Argonne National Laboratory. Novel experimental methodologies to elucidate the presence of strain in fine nanocrystals are a necessary pre-requisite in order to better understand strain profiles in engineered nanocrystals for novel device development

The use of DAPI fluorescence lifetime imaging for investigating chromatin condensation in human chromosomes

Chromatin undergoes dramatic condensation and decondensation as cells transition between the different phases of the cell cycle. The organization of chromatin in chromosomes is still one of the key challenges in structural biology. Fluorescence lifetime imaging (FLIM), a technique which utilizes a fluorophore’s fluorescence lifetime to probe changes in its environment, was used to investigate variations in chromatin compaction in fixed human chromosomes. Fixed human metaphase and interphase chromosomes were labeled with the DNA minor groove binder, DAPI, followed by measurement and imaging of the fluorescence lifetime using multiphoton excitation. DAPI lifetime variations in metaphase chromosome spreads allowed mapping of the differentially compacted regions of chromatin along the length of the chromosomes. The heteromorphic regions of chromosomes 1, 9, 15, 16, and Y, which consist of highly condensed constitutive heterochromatin, showed statistically significant shorter DAPI lifetime values than the rest of the chromosomes. Differences in the DAPI lifetimes for the heteromorphic regions suggest differences in the structures of these regions. DAPI lifetime variations across interphase nuclei showed variation in chromatin compaction in interphase and the formation of chromosome territories. The successful probing of differences in chromatin compaction suggests that FLIM has enormous potential for application in structural and diagnostic studies

Platinum blue staining of cells grown in electrospun scaffolds

Fibroblast cells grown in electrospun polymer scaffolds were stained with platinum blue, a heavy metal stain, and imaged using scanning electron microscopy. Good contrast on the cells was achieved compared with samples that were gold sputter coated. The cell morphology could be clearly observed, and the cells could be distinguished from the scaffold fibers. Here we optimized the required concentration of platinum blue for imaging cells grown in scaffolds and show that a higher concentration causes platinum aggregation. Overall, platinum blue is a useful stain for imaging cells because of its enhanced contrast using scanning electron microscopy (SEM). In the future it would be useful to investigate cell growth and morphology using three-dimensional imaging methods