Table 1.
The limitations of transmission electron microscopy, Raman spectroscopy, near-IR fluorescence microscopy, and bright-field optical microscopy and the potential advantages of photoacoustic microscopy in detection, mapping, and quantification of SWCNTs in histological specimens.
Figure 1.
Acoustic-resolution photoacoustic microscopy (AR-PAM) of single walled carbon nanotubes (SWCNTs).
PAM images at 4 different concentrations (a) 400 ng, (b) 800 ng, (c) 10 µg and (d) 40 µg of the SWCNTs spread across a given area on a glass slide, (e) an unknown concentration of SWCNTs spread on the surface of a human breast cancer tissue section and (f) unstained human breast cancer tissue. All scale bars are 1 mm.
Figure 2.
The linear-fitted standard curve of photoacoustic signals obtained from 400 ng, 800 ng, 10 µg and 40 µg of single walled carbon nanotubes.
For each concentration, 3 spots were spotted in a small area on a glass slide and air dried. The photoacoustic amplitude of the nanotubes distributed in each spot is integrated and represents the total photoacoustic amplitude. Each data point in Fig. 2 represents the average photoacoustic amplitude of 3 spots at each concentration, and the error bar represents the standard deviation. The R2 value for the linear-fitting curve is 0.992.
Table 2.
Concentration-dependent photoacoustic signals produced by single walled carbon nanotubes (SWCNTs).
Figure 3.
Comparison between conventional transmission-mode optical microscopy and optical-resolution photoacoustic microscopy (OR-PAM) of human breast cancer tissues in the absence and presence of single walled carbon nanotubes (SWCNTs).
Optical images of H&E stained breast cancer tissue sections without (a) and with (c) SWCNTs. OR-PAM images of the same human breast cancer tissue without (b) and with (d) SWCNTs. In panel c, artifacts generated during the preparation of histological specimens such as micro- and nano-sized bubbles (represented by the letters B) nuclear stains (green arrows) and, dust particles (blue arrows) are shown and appear black along with SWCNTs aggregates (red arrows). In panel d, only the SWCNT aggregates are detected (red arrows) and the artifacts are not detected.. The scale bar in (a) is 200 µm and applies to all the panels.
Figure 4.
Confocal Raman (a–c) and NIR luminescence (d–f) imaging of stained and unstained breast cancer tissue with SWCNTs.
Optical transmission microscopy before (a) and after (b) laser excitation during Raman imaging of SWCNTs in H&E stained breast cancer tissue. (c) Raman spectrum obtained on the same tissue using a WITec Alpha 300 Confocal Raman Microscope excited with a 532 nm laser. The laser intensity used damages the tissue under investigation and creates artifacts represented by letter ‘B’. The Raman spectra of SWCNTs showing G (tangential oscillations of carbon atoms along the nanotube axis) and D (vibrational modes of disordered carbon) bands. (d) Confocal images of breast cancer tissue containing SWCNTs in the bright field. (e) NIR Confocal fluorescence image of the tissue with SWCNTs (λex = 650 nm, λem = 750–900 nm. (f) Overlay of optical and NIR images of breast cancer tissue with SWCNT. Circles show the SWCNT aggregates.
Figure 5.
Histological sections of poly(propylene fumarate) (PPF) scaffold implanted rabbit subcutaneous tissue stained with methylene blue.
Images are presented at 2× (a), 4× (b) and 10× (c) magnification. The connective tissue (CT) and PPF scaffold (P) is present in all the sections. Inflammatory cells (IC) and blood vessel (BV) are also observed in the images (a) and (c). The red arrows in all the images represent regions of intense staining and the black arrows represent air bubbles and stain spots.
Figure 6.
Comparison between conventional transmission-mode optical microscopy and optical-resolution photoacoustic microscopy (OR-PAM) of subcutaneous rabbit tissue implanted with poly(propylene fumarate) (PPF) scaffold for 12 weeks and labeled with SWCNTs.
(a) The optical image and (b) the OR-PAM image of methylene blue and basic fuchsin stained subcutaneous rabbit tissue with SWCNTs. (c) Image processed using the SCANCO software by subtracting the background signal of the breast tissue. The SWCNTs appear as black aggregates (NT), the blue stained areas represent the connective tissue CT, and ‘P’ represents the PPF scaffold in the optical images.