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Fluoromodule-based reporter/probes designed for in vivo fluorescence imaging
Ming Zhang, … , Marcel P. Bruchez, Alan S. Waggoner
Ming Zhang, … , Marcel P. Bruchez, Alan S. Waggoner
Published September 8, 2015
Citation Information: J Clin Invest. 2015;125(10):3915-3927. https://doi.org/10.1172/JCI81086.
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Technical Advance Oncology

Fluoromodule-based reporter/probes designed for in vivo fluorescence imaging

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Abstract

Optical imaging of whole, living animals has proven to be a powerful tool in multiple areas of preclinical research and has allowed noninvasive monitoring of immune responses, tumor and pathogen growth, and treatment responses in longitudinal studies. However, fluorescence-based studies in animals are challenging because tissue absorbs and autofluoresces strongly in the visible light spectrum. These optical properties drive development and use of fluorescent labels that absorb and emit at longer wavelengths. Here, we present a far-red absorbing fluoromodule–based reporter/probe system and show that this system can be used for imaging in living mice. The probe we developed is a fluorogenic dye called SC1 that is dark in solution but highly fluorescent when bound to its cognate reporter, Mars1. The reporter/probe complex, or fluoromodule, produced peak emission near 730 nm. Mars1 was able to bind a variety of structurally similar probes that differ in color and membrane permeability. We demonstrated that a tool kit of multiple probes can be used to label extracellular and intracellular reporter–tagged receptor pools with 2 colors. Imaging studies may benefit from this far-red excited reporter/probe system, which features tight coupling between probe fluorescence and reporter binding and offers the option of using an expandable family of fluorogenic probes with a single reporter gene.

Authors

Ming Zhang, Subhasish K. Chakraborty, Padma Sampath, Juan J. Rojas, Weizhou Hou, Saumya Saurabh, Steve H. Thorne, Marcel P. Bruchez, Alan S. Waggoner

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Figure 4

Application and quantitation of fluorescence from fluoromodules in living mice.

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Application and quantitation of fluorescence from fluoromodules in livin...
(A) Example of a nude mouse implanted with 300 cells of iRFP713-expressing HCT116 cells in the left flank and an equal number of Mars1Cy-expressing cells, prelabeled with SC1, in the right flank. Image is representative of data quantified in B. The fluoromodule-hosting cells used here express less protein than the cells hosting iRFP713 (Supplemental Figure 5). (B) Fluorescence measured from mice injected and imaged as described in A, using different numbers of cells. Points denote means; bars indicate SEM (n = 3: 1 × 105 Mars1Cy-expressing cells, n = 4: others, total animals treated and imaged: 16). Analysis of the same data in terms of relative fluoromodule signal yields similar results (Supplemental Figure 6). (C) Fluorescence image of a tumor section (3.2 by 1.7 mm) composed of HCT116 cells that express Mars1Cy on the cell surface, labeled by SC1 given i.p. Scale bar: 16 μm. Field is near the center of the section. (D and E) s.c. FAP-bearing tumors that express Mars1Cy on the cell surface were labeled by SC1 (400 nM final) administered i.p. and i.v. (tail vein), respectively. (F and G) s.c. tumors as in D and E were labeled with SCi1 (400 nM final), injected i.p. or i.v., respectively. (H) i.p. injection of SC1 produces no significant background signal when FAP-bearing cells are not present. (I and J) Mars1Cy-bearing tumors can be imaged in the i.p. cavity of mice after delivery of SC1 or SCi1, respectively, via i.p. injection. Data presented in C–J represent results obtained from 3 samples/animals. Scales in mouse images are given in millimeters.

Copyright © 2023 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)

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