Source: Arvin H. Soepriatna¹, Kelsey A. Bullens², and Craig J. Goergen¹

¹ Weldon School of Biomedical Engineering, Purdue University, West Lafayette, Indiana

² Department of Biochemistry, Purdue University, West Lafayette, Indiana

Near-infrared fluorescence (NIRF) imaging is an exciting optical technique that utilizes fluorescent probes to visualize complex biomolecular assemblies in tissues. NIRF imaging has many advantages over conventional imaging methods for noninvasive imaging of diseases. Unlike single photon emission computed tomography (SPECT) and positron emission tomography (PET), NIRF imaging is rapid, high-throughput, and does not involve ionizing radiation. Furthermore, recent developments in engineering target-specific and activatable fluorescent probes provide NIRF with high specificity and sensitivity, making it an attractive modality in studying cancer and cardiovascular disease. The presented procedure is designed to demonstrate the principles behind NIRF imaging and how to conduct in vivo and ex vivo experiments in small animals to study a variety of diseases. The specific example shown here employs an activatable fluorescent probe for matrix metalloproteinase-2 (MMP2) to study its uptake in two different rodent models of abdominal aortic aneurysms (AAAs).

The following procedure provides detailed steps needed to collect in vivo and ex vivo NIRF images from small animals:

1. Experimental Setup

1. Connect a fiber optic light source to the fluorescence imaging system using a fiber optic light guide.
2. Select the appropriate excitation filter for the experiment. The excitation filter determines the wavelength of light to be delivered to the sample and should be chosen to match the excitation spect

Representative in vivo and ex vivo NIRF images taken from rodents with abdominal aortic aneurysms (AAAs) are shown in Figures 1-2. An activatable fluorescent probe was injected systemically via the tail vein to visualize matrix metalloproteinase-2 (MMP2) activity. MMP2 is an elastolytic enzyme involved in the degradation of the extracellular matrix that plays a major role in the initiation and progression of AAA. All images were acquired using a 625 nm excitation filter, a 700 nm emission filter, and 60

NIRF imaging relies on fluorescent probes to quantify and visualize biomolecular assemblies in tissues. Absorbed photon energy from near-infrared light excites fluorescent molecules to a higher energy state, and the emitted light with a longer characteristic wavelength is captured by a fluorescence imaging system. Here, the application of NIRF imaging to study MMP2 activity in abdominal aortic aneurysms was demonstrated in vivo and ex vivo. Unlike SPECT or PET, which are