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Genomics


The DeltaVision


OMX Blaze system by GE Healthcare


between free virus and virus-infected cells in the bloodstream. The second example shows how imaging has led to new insights in the conserved spatial organisation of proteins in the ‘divisome’ of bacteria and how that relates to the cytokinetic processes essential for cell division. 3D-Structured Illumination Microscopy (3D-SIM) is a super reso- lution technique that approximately doubles the resolution in all three dimensions compared to conventional fluorescence-based optical micro- scopy methods. The result is an eight-times improvement in volume resolution. SR-SIM can achieve lateral resolution of 50-60nm and axial resolution ranging from 150-300nm. Brown et al1 used 3D super-resolution microscopy to image primary human natural killer (NK) cells. NK cells are components of the human immune system capable of secreting cytotoxic granules that can lyse and kill virus-infected or oth- erwise pathologically-transformed cells that they recognise as foreign. NK cells are part of the body’s cell-mediated immune defence system; they also secrete chemical signalling compounds called cytokines (eg, interferon) to communicate with other cells. In the study, images of NK cells after stimulation of different cell surface receptors showed the intra- cellular accumulation of lytic granules and inter- feron-gamma at actin mesh synaptic sites. The images demonstrated that while the NK cells can recognise free virus in blood (and specifically influenza particles), this recognition alone did not lead to receptor activation and opening of the actin


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mesh at the synapses unless there was co-ligation of leukocyte function-associated antigen-1 (LFA- 1). LFA-1 is an integrin that mediates cell adhesion. These findings, and the ability to visualise the remodelling of the synaptic actin using super-reso- lution imaging, led the authors to propose a novel mechanism to explain the results. They suggested that the NK cells rely on integrin recognition to differentiate between free pathogen and pathogen- infected cells in blood and proposed integrin recog- nition as a mediator of receptor activation. In the second example, Strauss et al2 described the application of 3D-SIM super-resolution microscopy to study the dynamic localisation of the FtsZ protein in two types of bacteria undergo- ing cell division: the rod-shaped Bacillus subtilis and the spherical Staphylococcus aureus. FtsZ is a tubulin-like cytoskeletal protein that polymerises to form the Z ring, which acts as a scaffold during bacterial cell division, recruits other proteins need- ed for cell division and generates a contractile force through constriction of the Z ring that is required for cytokinesis. Super-resolution 3D-SIM was used to image the architecture of Z ring structures pres- ent in the two types of bacteria.


Conventional fluorescence microscopy using green fluorescent protein (GFP) fusion proteins had previously determined that the Z ring is assembled from FtsZ precursors and is a dynamic structure that undergoes continuous subunit turnover. Conventional microscopy also led to the conclusion that the Z ring is ‘a continuous struc- ture of uniform density’. In contrast, SR-SIM


Drug Discovery World Spring 2013


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