Abstract

We outline the process for determining the morphology of subsurface epithelial cell nuclei using depth-resolved light scattering measurements. The measurements are accomplished using a second generation angle-resolved low coherence interferometry system. The new system greatly improves data acquisition and analysis times compared to the initial prototype system. The calibration of the new system is demonstrated in scattering studies to determine the size distribution of polystyrene microspheres in a turbid sample. The process for determining the size of cell nuclei is discussed by analyzing measurements of basal cells in a sub-surface layer of intact, unstained epithelial tissue.

© 2003 Optical Society of America

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References

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  1. Backman, V., V. Gopal, M. Kalashnikov, K. Badizadegan, R. Gurjar, A. Wax, I. Georgakoudi, M. Mueller, C.W. Boone, R.R. Dasari, and M.S. Feld, "Measuring cellular structure at submicrometer scale with light scattering spectroscopy," IEEE. J. Sel. Top. Quantum Electron. 7, 887-893 (2001).
    [CrossRef]
  2. Sokolov, K., J. Galvan, A. Myakov, A. Lacy, R. Lotan, and R. Richards-Kortum, "Realistic three-dimensional epithelial tissue phantoms for biomedical optics," J. Biomed. Opt. 7, 148-156 (2002).
    [CrossRef] [PubMed]
  3. Mourant, J.R., T.M. Johnson, S. Carpenter, A. Guerra, T. Aida, and J.P. Freyer, "Polarized angular dependent spectroscopy of epithelial cells and epithelial cell nuclei to determine the size scale of scattering structures," J. Biomed. Opt. 7, 378-387 (2002).
    [CrossRef] [PubMed]
  4. Wax, A., C. Yang, V. Backman, K. Badizadegan, C.W. Boone, R.R. Dasari, and M.S. Feld, "Cell organization and sub-structure measured using angle-resolved low coherence interferometry," Biophys. J. 82, 2256-2264 (2002).
    [CrossRef] [PubMed]
  5. Backman, V., M.B. Wallace, L.T. Perelman, J.T. Arendt, R. Gurjar, M.G. Muller, Q. Zhang, G. Zonios, E. Kline, T. McGillican, S. Shapshay, T. Valdez, K. Badizadegan, J.M. Crawford, M. Fitzmaurice, S. Kabani, H.S. Levin, M. Seiler, R.R. Dasari, I. Itzkan, J. Van Dam, and M.S. Feld, "Detection of preinvasive cancer cells," Nature 406, 35-36 (2000).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef]

Biophys. J.

Wax, A., C. Yang, V. Backman, K. Badizadegan, C.W. Boone, R.R. Dasari, and M.S. Feld, "Cell organization and sub-structure measured using angle-resolved low coherence interferometry," Biophys. J. 82, 2256-2264 (2002).
[CrossRef] [PubMed]

Cancer Research

Wax, A., C.H. Yang, M.G. Muller, R. Nines, C.W. Boone, V.E. Steele, G.D. Stoner, R.R. Dasari, and M.S. Feld, "In situ detection of neoplastic transformation and chemopreventive effects in rat esophagus epithelium using angle-resolved low-coherence interferometry," Cancer Research 63, 3556-3559 (2003).
[PubMed]

Gastroenterology

Wallace, M.B., L.T. Perelman, V. Backman, J.M. Crawford, M. Fitzmaurice, M. Seiler, K. Badizadegan, S.J. Shields, I. Itzkan, R.R. Dasari, J. Van Dam, and M.S. Feld, "Endoscopic detection of dysplasia in patients with Barrett's esophagus using light-scattering spectroscopy," Gastroenterology 119, 677-682 (2000).
[CrossRef] [PubMed]

Gastrointest. Endosc.

Lovat, L.B., D. Pickard, M. Novelli, P.M. Ripley, H. Francis, I.J. Bigio, and S.G. Bown, "A novel optical biopsy technique using elastic scattering spectroscopy for dysplasia and cancer in Barrett's esophagus," Gastrointest. Endosc. 51, 4919 (2000).

IEEE. J. Sel. Top. Quantum Electron.

Backman, V., V. Gopal, M. Kalashnikov, K. Badizadegan, R. Gurjar, A. Wax, I. Georgakoudi, M. Mueller, C.W. Boone, R.R. Dasari, and M.S. Feld, "Measuring cellular structure at submicrometer scale with light scattering spectroscopy," IEEE. J. Sel. Top. Quantum Electron. 7, 887-893 (2001).
[CrossRef]

J. Biomed. Opt.

Sokolov, K., J. Galvan, A. Myakov, A. Lacy, R. Lotan, and R. Richards-Kortum, "Realistic three-dimensional epithelial tissue phantoms for biomedical optics," J. Biomed. Opt. 7, 148-156 (2002).
[CrossRef] [PubMed]

Mourant, J.R., T.M. Johnson, S. Carpenter, A. Guerra, T. Aida, and J.P. Freyer, "Polarized angular dependent spectroscopy of epithelial cells and epithelial cell nuclei to determine the size scale of scattering structures," J. Biomed. Opt. 7, 378-387 (2002).
[CrossRef] [PubMed]

J. Opt. Soc. Am. A

Nature

Backman, V., M.B. Wallace, L.T. Perelman, J.T. Arendt, R. Gurjar, M.G. Muller, Q. Zhang, G. Zonios, E. Kline, T. McGillican, S. Shapshay, T. Valdez, K. Badizadegan, J.M. Crawford, M. Fitzmaurice, S. Kabani, H.S. Levin, M. Seiler, R.R. Dasari, I. Itzkan, J. Van Dam, and M.S. Feld, "Detection of preinvasive cancer cells," Nature 406, 35-36 (2000).
[CrossRef] [PubMed]

Opt. Lett.

Phys. Rev. Lett.

Wax, A., S. Bali, and J.E. Thomas, "Time-resolved phase-space distributions for light backscattered from a disordered medium," Phys. Rev. Lett. 85, 66-69 (2000).
[CrossRef] [PubMed]

Reports on Progress in Physics

Fercher, A.F., W. Drexler, C.K. Hitzenberger, and T. Lasser, "Optical coherence tomography �?? principles and applications," Reports on Progress in Physics 66, 239-303 (2003).
[CrossRef]

Other

van de Hulst, H.C., Light scattering by small particles. 1957, New York: Dover Publications.

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Figures (13)

Fig. 1.
Fig. 1.

(a). Schematic of new a/LCI system. L1-L5 - lenses, BS1,BS2 - beamsplitters, D1,D2 - balanced detectors, RR - retroreflector. Shaded area shows scattered light. (b) Diagram illustrating angle selectivity by scanning lens L2 perpendicular to the beam. Light at the top of the collimated beam entering L3 is directed to the detector in the top figure. Light at the center is directed to the detector in the bottom figure.

Fig. 2. (a)
Fig. 2. (a)

Contour plot showing angular scattering by suspension of polystyrene microspheres as a function of depth. Light areas indicate increased intensity on a log scale. (b) Angular distributions vs. depth. Top left 0–100 µm, top right 100–200 µm, bottom left, 200–300 µm, bottom right 300–400 µm.

Fig. 3.
Fig. 3.

Fitted angular distribution from surface layer

Fig. 4.
Fig. 4.

Minimization of chi-squared value to determine best fit.

Fig. 5.
Fig. 5.

Raw and filtered scattering data from basal cell nuclei

Fig. 6.
Fig. 6.

Filtered data from Fig.5 with low order fit

Fig. 7.
Fig. 7.

Oscillatory component of data with best fit.

Fig. 8.
Fig. 8.

Chi-squared minimization to find best fit. The values are parabolic (solid line) near the minimum. The uncertainty is given by double the minimum value.

Fig 9.
Fig 9.

Original microsphere data and data adjusted by adding a low order polynomial

Fig. 10.
Fig. 10.

Best fit yields incorrect size when an incorrect slowly varying background is present

Fig. 11.
Fig. 11.

Best fit yields correct size when slowly varying background is removed using a low order polynomial prior to fitting. Readjusted data corrects the slowly varying background.

Fig. 12.
Fig. 12.

Micrograph of microspheres (bars=10, 50 µm)

Fig. 13.
Fig. 13.

Fourier transform of a portion of image in Fig. 12

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