Research ArticleCANCER

Fast label-free multilayered histology-like imaging of human breast cancer by photoacoustic microscopy

+ See all authors and affiliations

Science Advances  17 May 2017:
Vol. 3, no. 5, e1602168
DOI: 10.1126/sciadv.1602168
  • Fig. 1 Schematic of the UV-PAM system for surgical margin imaging.

    The UV laser beam is first spatially filtered and expanded by a pair of lenses and a pinhole. The beam is then focused through an aspherical lens onto the bottom of the breast tissue specimen (sample), which is placed inside a water tank on top of a sample holder. Some generated acoustic waves propagate through the tissue and reach a focused ultrasonic transducer. The received acoustic pressure is transduced into an electric signal, which is then amplified and recorded by a data acquisition (DAQ) card. During data acquisition, a maximum amplitude projection (MAP) image from the measured B-scan data is displayed on a computer screen within approximately 1 s. By raster-scanning the sample holder, a MAP image from the C-scan data is also displayed.

  • Fig. 2 Imaging of thin breast tissue slices without and with H&E staining.

    (A) Unstained paraffin-embedded breast tissue slice imaged by the UV-PAM system. (B) H&E-stained deparaffinized breast tissue slice imaged by a standard microscope. The green dashed lines in (A) and (B) outline the boundaries of the normal and tumor regions. The upper-right area is the tumor region, and the bottom-left area is the normal region. (C and D) Zoomed-in UV-PAM and H&E-stained images of the normal regions (red dashed regions) in (A) and (B), respectively. The blue dashed lines and arrows on the top right-hand corners in (C) and (D) label a representative local deformation. (E) Overlay image of (D) on (C). (F and G) Zoomed-in UV-PAM and H&E-stained images of the tumor regions (yellow dashed regions) in (A) and (B), respectively. (H) Overlay image of (G) on (F).

  • Fig. 3 Imaging of a breast tumor from the first patient.

    (A) UV-PAM image of the fixed, unprocessed breast tumor. (B) H&E-stained histologic image of the same area shown in (A) acquired after processing, sectioning, and staining the excised breast tissue. The blue dashed lines in (A) and (B) outline the interface between the normal and tumor regions. (C and D) Zoomed-in UV-PAM and H&E-stained images of the red dashed regions in (A) and (B), respectively. (E and F) Zoomed-in UV-PAM and H&E images of the yellow dashed regions in (A) and (B), respectively. IDC, invasive ductal carcinoma; DCIS, ductal carcinoma in situ. (G) Zoomed-in UV-PAM image of the orange dashed region in (A). CN, cell nuclei.

  • Fig. 4 Imaging of a breast tumor from the second patient.

    (A) UV-PAM image of the fixed, unprocessed breast tumor. (B) H&E-stained histologic image of the same area shown in (A) acquired after processing, sectioning, and staining the excised breast tissue. (C and D) Zoomed-in UV-PAM and H&E images of the red dashed regions in (A) and (B), respectively. LCN, lymphocyte cell nucleus; TCN, tumor cell nucleus. (E and F) Zoomed-in UV-PAM and H&E-stained images of the yellow dashed regions in (A) and (B), respectively. D, duct. The two ducts are surrounded by invasive ductal carcinoma.

  • Fig. 5 Distributions of cell nuclear area values and internuclear distances in the breast tumor specimens (Figs. 3 and 4), where bin interval = 8 and n = 30 for each distribution.

    (A) Histogram of the cell nuclear cross-sectional areas imaged by UV-PAM (Figs. 3C and 4C). The green dashed line is a Gaussian fit for lymphocytes, with a mean of 32.8 μm2 and an SD of 7 μm2. The red dashed line is a Gaussian fit for cancer cells, with a mean of 67.6 μm2 and an SD of 11 μm2. (B) Histogram of the cell nuclear cross-sectional areas imaged by histology (Figs. 3D and 4D). The green dashed line is a Gaussian fit for lymphocytes, with a mean of 30.1 μm2 and an SD of 6.7 μm2. The red dashed line is a Gaussian fit for cancer cells, with a mean of 66 μm2 and an SD of 9.2 μm2. (C) Histogram of the internuclear distances imaged by UV-PAM (Figs. 3C and 4C). The green dashed line is a Gaussian fit for lymphocytes, with a mean of 13.1 μm and an SD of 3.8 μm. The red dashed line is a Gaussian fit for cancer cells, with a mean of 15.4 μm and an SD of 2.4 μm. (D) Histogram of the internuclear distances imaged by histology (Figs. 3D and 4D). The green dashed line is a Gaussian fit for lymphocytes, with a mean of 13.2 μm and an SD of 4.1 μm. The red dashed line is a Gaussian fit for cancer cells, with a mean of 17.2 μm and an SD of 2.9 μm.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/3/5/e1602168/DC1

    fig. S1. Illustrations of negative and positive margins.

    fig. S2. Experimentally measured spatial and axial resolutions of the UV-PAM system.

    fig. S3. Procedure for obtaining thin breast tissue slices without and with H&E staining.

    fig. S4. Imaging of a breast tumor from the third patient.

    fig. S5. Representative xz-projected human breast tumor image acquired over a 10 mm × 4.2 mm area from the first patient specimen.

    fig. S6. Photographs of the breast tissue specimens.

    fig. S7. Reproducibility of the UV-PAM system for breast tumor tissue imaging.

    movie S1. Closeup images of a row in the ROI of the first patient’s breast tumor specimen imaged by UV-PAM.

    movie S2. Series of depth-resolved images of the first patient’s breast tumor specimen imaged by UV-PAM.

  • Supplementary Materials

    This PDF file includes:

    • fig. S1. Illustrations of negative and positive margins.
    • fig. S2. Experimentally measured spatial and axial resolutions of the UV-PAM system.
    • fig. S3. Procedure for obtaining thin breast tissue slices without and with H&E staining.
    • fig. S4. Imaging of a breast tumor from the third patient.
    • fig. S5. Representative xz-projected human breast tumor image acquired over a 10 mm × 4.2 mm area from the first patient specimen.
    • fig. S6. Photographs of the breast tissue specimens.
    • fig. S7. Reproducibility of the UV-PAM system for breast tumor tissue imaging.
    • Legends for movies S1 and S2

    Download PDF

    Other Supplementary Material for this manuscript includes the following:

    • movie S1 (.mp4 format). Closeup images of a row in the ROI of the first patient’s breast tumor specimen imaged by UV-PAM.
    • movie S2 (.mp4 format). Series of depth-resolved images of the first patient’s breast tumor specimen imaged by UV-PAM.

    Files in this Data Supplement:

Related Content