Acquisition of a Leica Stellaris 5 Upright Confocal Microscope for In Vivo Imaging Applications

Project Summary/Abstract
The purpose of this shared S10 instrumentation grant application is the purchase of a Leica upright Stellaris 5
confocal microscope to replace an obsolete, end-of-life Leica SP-5 confocal microscope dedicated for live
animal imaging. The main aim is to continue to provide high-resolution intravital confocal imaging capabilities
to the University of Miami UM research community by upgrading and replacing the existing instrument (now 18
years old) that has reached the end of its life and can no longer be repaired or serviced. The requested
microscope fills a critical need and will support the ongoing efforts of the NIH-funded research of 9 Major User
groups representing 5 Departments at the University of Miami (UM). Other than the microscope we are
replacing, there are no other available, open access microscopes at UM configured for live animal imaging
studies that meet the needs of the existing and future User Groups. The replacement Leica Stellaris 5 system
is configured with a white light laser (WLL) having an excitation range from 495 to 790 and a detection range
up to 850 nm. It also has a 405 nm laser for near-UV excitation. The extended excitation wavelengths
provided by the WLL allows for additional fluorescent reporters to be imaged simultaneously in a single
experiment, thereby providing additional information compared with the older technology that was limited to the
detection of 4 or 5 fluorescent reporters at best. The Stellaris 5 system enabling the multiplexing of fluorescent
reporters in a single experiment also reduces the utilization of research animals consistently with the 3Rs of
animal research (Replacement, Reduction, Refinement). The microscope is equipped with a 5x and a long-
distance 10X dry objects and with apochromatic water-immersion objectives (16x, 25x, and 40x) with high
numerical aperture and large free working distances that are essential for intravital imaging. The upright
configuration of the objective lenses is required for proper access to the live animal tissue to be imaged and
positioning of the objectives to regions of interest. The upright configuration and long working distance
objectives are necessary to accommodate the anesthesia apparatus and stereotaxic head-holder, which
require marked clearance from the microscope body and objectives mounting turret to allow positioning of the
animal on the stage and under the objective. The instrument will be housed and maintained at the open access
Analytical Imaging Core Facility (AICF), where the entire UM community has access to instrumentation on a
fee-for-service basis. The existing confocal microscopes in the AICF and other imaging facilities at the
University, other than the obsolete Leica SP-5 system, allow for high precision state-of-art imaging of ex vivo
tissues and live cells in culture vessels; however, the inverted configuration of those systems and the objective
lenses they are equipped with are not amenable for live animal imaging, for the reasons elaborated above.
Therefore, the replacement of the existing upright SP-5 system that reached its end-of-life is critically needed.

Grant Number: 1S10OD038196-01
NIH Institute/Center: NIH
Principal Investigator: Midhat Abdulreda