DescriptionLocation: Room 1.12, MRC Building Advantages: Speed, resolution, variety of laser lines, wide field of view, improved clarity of thick samples, flexible photo-bleaching and photo-activation Disadvantages: Lower sensitivity; The Vox 3i is a spinning disc confocal microscope designed for rapid acquisition of 3D and 4D data. The microscope consists of a Nikon TiE Zeiss AxioObserver Z1 inverted stand attached to a Yokogawa CSU-X1 W1 spinning disc scan head. The detector is a Hamamatsu C9100-13 EMCCD camera that multiplies the signal on the chip before reading out. This makes it more sensitive than a regular CCD but a large pixel size means that resolution is relatively low. The Ultraview Focus Drive (Prior NanoscanZ piezo focus) is generally used for acquiring stacks because it is fast and does not limit the rate of acquisition. It has a maximum range of 250 µm so any specimens thicker than this will have to be imaged using the microscope internal focus motor. The TiE stand includes the Nikon Perfect Focus System (PFS)There are two cameras on the system, a Photometrics Prime BSI Scientific CMOS and a Photometrics Prime 95B Scientific CMOS. The Prime BSI has smaller pixels for higher resolution, and the 95B has larger pixels for higher sensitivity. The Z1 stand includes Zeiss' Definite Focus system, which can maintain samples in precise focus indefinitely. This is most suitable for samples mounted at a glass/aqueous interface but there are some exceptions. Please check specimen and objective lens compatibility before using the PFSautofocus. The Vox also has a Photokinesis Unit (PKU) photomanipulation unit for spot or ROI illumination in techniques such as FRAP and photo-activation/photo-switching. Suitable specimens: All culture vessels must have glass bottoms with the same thickness as a standard #1.5 cover-slip (170 µm). There are inserts for slides, Ibidi µSlides, Lab-Tek chambered cover slips (note - use Lab-Tek I not Lab-Tek II), 35 mm dishes, multi-well plates with square not chamfered corners. If you have any other requirements then you must speak to a member of the light microscopy facility before you start preparing your specimen. Note that CO2 supply is not available for all configurations so check what vessel you need before you start setting up your specimen. In general spinning disc confocal systems perform optimally with adherent cells in culture and other relatively thin specimens. Thicker specimens with a large amount of out-of-focus fluorescence can be difficult to image because of a phenomenon called pinhole crosstalk. The CSU-W1 has a an additional 25µm spinning disk for use with thicker samples at lower magnification. - Using the Nikon TiE microscope
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