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Biomedical Imaging and Instrumentation

Biomedical Imaging and Instrumentation - Poster Session B

Poster F8 - Developing A Protocol for Microscopy with UV Surface Excitation (MUSE) Imaging of the Kidney in Pre-Clinical Investigation of Diabetic Kidney Disease

Thursday, October 24, 2024

2:45 PM – 3:45 PM EST

Location: Exhibit Hall E, F & G

Presenting Author(s)

Preye Porbeni

Student
Joint Department of Biomedical Engineering at UNC and NC State, United States

Primary Investigator(s)

Kennita Johnson, PhD (she/her/hers)

Assistant Professor
Joint Department of Biomedical Engineering at UNC and NC State
Chapel Hill, North Carolina, United States

Introduction: Histology is used to examine tissues on the cellular level and is the gold standard for diagnosing disease. Unfortunately, histology can be very time-intensive and not conducive for quick examination during experimentation. Microscopy with UV Surface Excitation (MUSE) imaging allows for quick examination of tissues because it uses ultraviolet (UV) light instead of white light. UV light penetrates deeper into the tissue than white light providing a better view of the tissue microstructures. Tissues can be stained very quickly (less than a minute) with fluorescent dyes that are excited by the UV light. This staining highlights different structures within the tissue, which allows for the simulation and imitation of standard histology. The goal of this research is to develop a protocol for MUSE imaging of rodent kidney. To meet this goal, we met the following objectives: 1) To refine the MUSE microscope set-up, 2) Determine fluorescent dye concentrations suitable for imaging, and 3) Characterize the combination of fluorescent dyes that created the most detailed image of the tissue.

Materials and

Methods: We constructed a custom MUSE microscope with a 5-megapixel camera, 4X objective, and 3-axis motion stage. We designed a camera stand to fit under the specimen to capture the reflected UV light off the specimen on top of the quartz glass stage. We tested four concentrations (100%, 10%, 1%, 0.1% of solubility in water) for: DAPI, rhodamine, eosin Y, hoechst, fluorescein, and light green. Fluorescent dyes were pipetted onto cotton rounds and were imaged under UV light at the same camera settings to determine which concentrations were suitable for imaging. Chosen dyes were combined to stain porcine and rodent kidney tissue and images were visually inspected to discern kidney structures.

Results, Conclusions, and Discussions: The 3D printed scaffolding for the camera and specimen stage provided stability to capture clear images of kidney tissues. All dyes were oversaturated at 100% solubility in water. They provided satisfactory illumination at 1% and 0.1% solubility in water. Fluorescent dye concentrations were combined to delineate structures in the kidney (Figure 1). Staining procedures were 40 seconds per dye. We were able to distinguish differences in the cortex and outer medulla of control and diabetic kidneys. This imaging provided new information that we could not get from our experimentation with ultrasound or histology. MUSE imaging offers a quick intermediate step between the other two imaging modalities in our studies.

Acknowledgements (Optional):