Near-infrared molecular probes for in vivo imaging

Abstract

Cellular and tissue imaging in the near-infrared (NIR) wavelengths between 700 and 900 nm is advantageous for in vivo imaging because of the low absorption of biological molecules in this region. This unit presents protocols for small animal imaging using planar and fluorescence lifetime imaging techniques. Included is an overview of NIR fluorescence imaging of cells and small animals using NIR organic fluorophores, nanoparticles, and multimodal imaging probes. The development, advantages, and application of NIR fluorescent probes that have been used for in vivo imaging are also summarized. The use of NIR agents in conjunction with visible dyes and considerations in selecting imaging agents are discussed. We conclude with practical considerations for the use of these dyes in cell and small animal imaging applications.

Figure 1

(A) Simplified schematic of a preclinical planar fluorescence reflectance imaging system with light source, imaging chamber and digital camera for detection. The wavelengths of excitation and emission collection can be selected with optical bandpass filters. (B) Brightfield image of mouse with subcutaneous tumor in left flank. (C) Fluorescence image of same mouse showing signal from NIR fluorescent molecular probe IntegriSense 680 (center wavelenth of filters: excitation 630 nm, emission 700 nm)

Figure 2

Structures of a) cyanine, b) ICG, c) SIDAG, d) PPCy.

Figure 3

New NIR cyanines. a) cyanine with RGD peptide, b) pH-sensitive cyanines, c) ROS cyanine.

Figure 4

Structures of representative miscellaneous NIR dyes. a) m-THPC, b) squaraine, c) BODIPY, d) push-pull-type NIR dye

Figure 5

Representative nanoparticle-based NIR probes for biological imaging.

Figure 6

Schematic illustration of self-illuminating NIR QDs.

Figure 7

Lung cancer cells treated with LS166 and costained with mitotracker green. Panel A shows the 780 nm channel alone, panel B shows the 488 nm and 780 nm panels merged. Yellow areas indicate co-localization. (Ye, et al., 2008))

Figure 8

Anticipated results from time-domain diffuse optical imaging. (A) fluorescence decay curves are collected for each measurement point in a raster scan of selected 2D region. (B) The fluorescence lifetime can be determined by fitting the tail of the decay curve. (C) Fluorescence intensity maps can be created by integration of fluorescence counts. Regions of interest are selected manually to determine average fluorescence intensity from dietary contents in the gastrointestinal (GI) tract (S1) or tumor (S2). (D) Fluorescence lifetime maps are created from fitting of the decay curves for each point. This map shows that the fluorescence from the GI (S3) can be distinguished from that of the fluorescent molecular probe in the tumor (S4) by differences in lifetime.