Near-infrared gold nanocages as a new class of tracers for photoacoustic sentinel lymph node mapping on a rat model

Abstract

This work demonstrated the use of Au nanocages as a new class of lymph node tracers for noninvasive photoacoustic (PA) imaging of a sentinel lymph node (SLN). Current SLN mapping methods based on blue dye and/or nanometer-sized radioactive colloid injection are intraoperative due to the need for visual detection of the blue dye and low spatial resolution of Geiger counters in detecting radioactive colloids. Compared to the current methods, PA mapping based on Au nanocages shows a number of attractive features: noninvasiveness, strong optical absorption in the near-infrared region (for deep penetration), and the accumulation of Au nanocages with a higher concentration than the initial solution for the injection. In an animal model, these features allowed us to identify SLNs containing Au nanocages as deep as 33 mm below the skin surface with good contrast. Most importantly, compared to methylene blue Au nanocages can be easily bioconjugated with antibodies for targeting specific receptors, potentially eliminating the need for invasive axillary staging procedures in addition to providing noninvasive SLN mapping.

Figures

Figure 1.

Absorption spectrum of Au nanocages. Wavelength at peak optical absorption was ~735 nm. Inset shows a typical SEM image of the Au nanocages with the scale bar being 50 nm. The average edge length was 50.3 ± 6.7 nm (standard deviation). The Au nanocages were stabilized with polyvinyl pyrrolidone (PVP).

Figure 2.

In vivo noninvasive photoacoustic time-course sagittal MAP images. (A) Photograph of axillary region with hair removed. (B) Photograph with skin and fatty tissue removed after the photoacoustic images had been recorded. The dark blue dyed lymphatic channel on the bottom of the SLN is shown in the inset. Photoacoustic images acquired before (C) and after (D–H) the nanocage injection: (D) 5 min (SLN started to appear), (E) 59 min, (F) 140 min, (G) 194 min. All images were acquired without signal averaging. (H) Accumulations of nanocages in a SLN over time, in terms of the amplitude changes of PA signals. After the injection, PA signals increased with time, which means gradual accumulations of nanocages. Peak accumulation occurred at ~140 min after the injection. PA signals from the SLN were normalized by those from adjacent blood vessels (the dotted box in Figure 2E) to minimize the ultrasonic focal effect, and normalized by maximum. Error bar is standard error. BV, blood vessels; SLN, sentinel lymph node. Color bar represents optical absorptions.

Figure 3.

(A) Experimental setup for PA signal measurements from SLNs and gelatin phantoms containing nanocages. (B) PA signal from the gelatin phantom containing nanocages with various concentrations (0–8 nM). All measured values were scaled down to 0 dB gain. Errorbar indicates standard deviation. (C) Ex vivo photoacoustic images and photographs of dissected SLNs containing Au nanocages for PA signal measurement. A photoacoustic image (1) and a corresponding photograph (2) of the highly accumulated SLN. A photoacoustic image (3) and a corresponding photography (4) of the low accumulated SLN. In the photographs, dark blue represents the accumulation of nanocages. SLN, sentinel lymph node.

Figure 4.

Depth capability of noninvasive photoacoustic SLN mapping in another rat, demonstrating potential for clinical use. Photoacoustic sagittal MAP images before (A) and after (B–E) the injection: (A) control image; (B) 28 min; (C) 126 min with a layer of chicken breast tissue placed on axillary region, total imaging depth was 10 mm; (D) 165 min with the second layer of chicken breast tissue, total imaging depth was 21 mm; (E) 226 min with the third layer of chicken breast tissue, total imaging depth was 33 mm. (F) Photoacoustic B-scan with 20 times signal average, showing the SLN located 33 mm deep. Memory of the acquisition system limited the record length in depth. (G) The amplitude variations of photoacoustic signals over imaging depths. Data were scaled down to 0 dB and normalized by the maximum. Errorbar represents standard deviation. All images were acquired without signal averaging except the B-scan (F). Colorbars represent the optical absorption. BV, blood vessel; SLN, sentinel lymph node.