Live-Cell Imaging

Andor Revolution XD Confocal inverted microscope

• Nikon Eclipse Ti inverted microscope with Perfect Focus and DIC

• Nikon Objectives:
  • 100x Apo TIRF (Oil, NA = 1.49, WD = 0.12 mm) with Temperature Correction collar (for 23°C and 37°C)
  • 60x Plan Apo VC (Oil, NA = 1.4, WD = 0.13 mm)
  • 40x Plan Fluor (Oil, NA = 1.3, WD = 0.2 mm)
  • 20x Plan Fluor (Dry, NA = 0.5, WD = 2.1 mm)

• iXon x3 897 EM-CCD camera: back illuminated, 512×512, 16 μm pixels, 35 fps, -100°C cooling

• Neo sCMOS camera: 2560×2160, 6.45 μm pixels, 100 fps, -40°C cooling

• Yokogawa CSU-X1 confocal spinning disk head: 5000 rpm max scan speed, 1000 scans per second.  Technical overview from Andor.

• Andor Laser Combiner (ALC): Four solid state lasers at 405, 488, 561, and 640 nm with 10 millisecond switch time and active blanking control to minimize unnecessary illumination.

• Confocal emission bandpass filters:
  • DAPI: 447/60 (417-477nm)
  • GFP: 525/30 (510-540nm)
  • RFP: 607/36 (589-625nm)
  • Far-red (CY5): 685/40 (665-705nm)

• Andor TIRF module: 488 and 561 nm

• ASI motorized stage with piezo Z for rapid Z-stack acquisition.  Piezo travel range of 150 um, minimum step size of 2.2 nm.

• Okolab CO2 cage incubator: Environmental regulation of temperature, CO2, and humidity to achieve cell culture conditions.  Temperature is thermostat-regulated to ± 0.1°C from 3° above ambient temperature up to 50°C.

• Andor iQ2 Live Cell Imaging software for data capture and analysis – detailed software webinars and tutorials available online.

• Bitplane’s Imaris Imaging Workstation for advanced data display and analysis.  Microscope users may train themselves on the software and will receive free software use based on their microscope use.

Supported by the Department of Ophthalmology and Visual Sciences, the University of Wisconsin School of Medicine & Public Health, the Carl and Mildred Reeves Foundation for Macular Degeneration Research, and generous funding provided by the Schuette/Kraemer Family gift.