Abstract

Polarization-resolved, second harmonic generation (P-SHG) microscopy at single pixel resolution is utilized for medical diagnosis of pathological skin dermis. In analyzing the large area, pixel by pixel, second-order susceptibility of normal and pathological skin dermis, we found that P-SHG can be used to distinguish normal and dermal pathological conditions of keloid, morphea, and dermal elastolysis. Specifically, we found that the second order susceptibility tensor ratio of d33/d31 for normal skins is 1.27±0.20, while the corresponding values for keloid, morphea, and dermal elastolysis are respectively 1.67±0.29, 1.79±0.30, and 1.75±0.31. We also found that the histograms of the d33/d31 ratio for the pathological skins contain two peak values and are 1.5 times wider than that of the normal case, suggesting that the pathological dermal collagen fibers tend to be more structurally heterogeneous. Our work demonstrates that pixel-resolved, second-order susceptibility microscopy is effective for detecting heterogeneity in spatial distribution of collagen fibers and maybe used for future clinical diagnosis and in vivo studies of collagen pathological conditions.

© 2009 Optical Society of America

1. Introduction

Collagenous diseases of skin dermis often involve excessive production of collagen fibers and dissolution of elastic fibers as these processes are commonly observed in three common types of skin dermis pathologies of keloid, morphea, and dermal elastolysis. Morphologically, keloid results in thick collagen fibers resulting from the wound healing processes that extends beyond the boundaries of the original wound. The abnormal proliferation of collagen fibers can result in the formation of benign tumors [1-3]. On the contrary, morphea is caused by autoimmune deficiencies, resulting in the thickening of collagen fibers [4]. Finally, dermal elastolysis is associated with skin softening and wrinkle formatison resulting from the decrease of elasticity at the affected sites. One explanation for the cause of this condition is the enzymatic degradation of elastic fiber in the dermal layer resulting from genetic mutation [5, 6]. Clearly, developing the appropriate imaging modality capable of resolving differences in collagen content between normal and pathological skin dermis will be of tremendous value for applications in clinical diagnosis of skin pathologies and in vivo investigations of these diseases in animal models.

In recent years, SHG imaging has been effectively used to investigate biomedical problems from studying tumor biology to detecting cornea pathologies [79]. Due to its coherent nature, the polarization dependence of SHG is sensitive to the structure and arrangement of the molecules that produces the signal [1012]. Since type I collagen is a major component of skin dermis and is known to produce strong second harmonic generation signal, SHG microscopy is a promising technique for diagnosing dermal abnormalities. In particular, by measuring the SHG intensity variation with respect to the polarization of the incident laser light, the second order susceptibility tensor element ratios can be determined [13, 14]. In this work, we investigate the potential of using excitation polarization-resolved, second harmonic generation (P-SHG) microscopy to characterize the changes in the dermal collagen of human skin with the pathological conditions of keloid, morphea, and dermal elastolysis.

2. Materials and methods

2.1 Second order susceptibility image analysis

In general, polarization P of a material induced by an external field E can be written as:

P=χ(1)E+χ(2)EE+χ(3)EEE+...,

where the second order susceptibility tensor χ (2) determines the strength and property of the second harmonic generation signal [31]. Since collagen fiber is composed of triple helical polypeptide chain, aligned parallel in forming bundles [15], we assume, on the scale of SHG imaging resolution, that the χ (2) tensor has cylindrical symmetry. This assumption reduces the number of independent χ (2) tensors to 4 elements [10, 12]. Under the experimental condition that the fiber lies in the x-z plane and the laser is incident along the +y directions (Fig. 1), the induced polarization for SHG can be modeled as

{Px=(d15sin2φ)E2Pz=(d31sin2φ+d33cos2φ)E2

where the contracted notation dij is used to describe the susceptibility tensor [31] with d31=χzxx/2, d15=χ xzx/2 and d33=χzzz/2. In addition, ϕ is the angle between the excitation laser polarization and fiber orientation, ϕ=θL-θF [10, 12]. However, since it was shown that the fiber orientations is equal to the principal axis of the second order nonlinear susceptibility tensor [16], therefore, ϕ is also the angle between the principal axis of the tensor and the polarization of the excitation source. From this analysis, the total SHG produced along the x and z axes can be expressed as

IPx2+Pz2{(d15d31sin2ϕ)2+(sin2ϕ+d33d31cos2ϕ)2}E4

In Eq. (3), the parameters to be determined are d15/d31, d33/d31, the fiber orientation (θF), and the overall proportionality constant. By measuring the dependence of the SHG intensity on the polarization of incident light, these parameters can be determined by curve fitting. Experimentally, this is accomplished by acquiring SHG images at different polarization angles θL of the incident laser light. However, for each set of data, there are two sets of parameters with θF differ by 90 degrees that produce the same SHG polarization dependence. This degeneracy can be resolved since it is known that the minimum SHG intensity results when the excitation polarization and collagen fiber orientation are approximately perpendicular [10, 1214].

 

Fig. 1. Graphical illustration showing the polarization direction of the laser relative to the orientation of the fiber. The laser is incident along the +y direction. θL is the angle of the excitation polarization and θF is the angle of fiber orientation measured with respect to the z-axis. The angle between excitation polarization and fiber orientation is ϕ=θL-θF.

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2.2 Excitation polarization resolved second harmonic generation microscope

The microscope system used to obtain P-SHG images (Fig. 2) is similar to one previously described [17, 18]. It is based on an upright microscope (E800, Nikon, Japan) with a femtosecond pulsed light source from a Ti:sapphire laser (Tsunami, Spectra Physics) pumped by a diode-pumped, solid-state(DPSS) laser (Millennia X, Spectra Physics, Mountain View, CA). A half-wave-plate and a linear polarizer were used to control the 780 nm output of the laser source, and an additional half and quarter-wave-plate combination were used to compensate for the depolarization effect of the main dichroic mirror (700dcspxruv-2p, Chroma Technology, Rockingham, Vermont) [18]. The laser light is guided by a set of galvanometer controlled scanning mirrors (Model 6220, Cambridge Technology) onto the back aperture of a 20× water immersion objective (Plan Fluor, NA 0.75, Nikon, Japan). The average laser power at the sample was 10 mW and the scanned field size was 206×206 µm2 composed of 240×240 pixels. Furthermore, the pixel dwell-time was 50 µsec. As previously discussed, the use of a low NA objectives prevents further depolarization of the excitation source [19]. The backward SHG is collected with the same objective, and filtered by a secondary dichroic mirror (435DCXR, Chroma Technology) and a narrow band pass filter (HQ390/20, Chroma Technology). Single-photon counting photomultiplier tubes (R7400P, Hamamatsu, Japan) were used as detectors. In our experiments, we elected to collect backward SHG signal for analysis. This choice is primarily motivated by the fact that in clinical and in vivo animal model studies, the backward SHG signal is much more accessible. Furthermore, since it was shown that the backward SHG signal can be dominant over the forward SHG signal due to the characteristic interaction length of collagen [20], backward SHG detection in collagen-containing tissues may be more efficient for diagnostic purposes. Finally, since our backward SHG signal was not polarization-resolved, tissue scattering should not affect our analysis. However, since our methodology requires the linearity of the excitation polarization to be maintained, unless tissue depolarization effects are corrected for, this approach may not be applicable to tissues beyond the imaging depths of around 20 µm [21].

In fitting for the d15/d31 and d33/d31 ratios, images at 21 excitation polarizations over 180 degrees were obtained. At each excitation polarization, 3 images were acquired for determination of the average value and standard deviation of d15/d31 and d33/d31. However, in fitting for the d15/d31 and d33/d31 ratios, we only analyzed pixels with an average SHG photon count (over all 21 excitation polarization angles) equal to or above 2. This was done to avoid poor fitting from the low data dynamic range. Pixel values which were not analyzed were assigned the background pseudocolor in the d15/d31 and d33/d31 images. Fitting and plotting of the results were performed using an IDL program (ITT Visual Information Solutions) and NIH ImageJ software.

 

Fig. 2. Schematic diagram of the P-SHG microscope. Half and quarter wave plates were used to compensate the depolarization caused by the main dichroic mirror.

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2.3 Specimen preparation

Both normal and pathological skins were obtained from the National Taiwan University Hospital. The study protocol was approved by the University’s Institutional Review Board. Four normal human skin specimens were obtained from four different patients, and three types of human pathological skins, keloid, morphea, and dermal elastolysis, were obtained from 13 patients. All specimens were formalin fixed, paraffin embedded, and sectioned to 5 µm thick. Common characteristics of these pathological tissues include the excessive production of collagen fibers and the dissolution of elastic fibers. The reticular dermis area of each sample was selected for imaging.

3. Results and discussions

A representative analysis for obtaining ratios of the second order susceptibility-resolved image is shown in Fig. 3. Figure 3(a) shows the normal skin SHG image with intensity average over 21 incident polarization angles. The polarization dependence for a selected pixel is shown in Fig. 3(b). The error bars represent the standard deviations by averaging over the polarization resolved SHG intensities of 3 repeated images, and the solid curve is the result of model fitting using Eq. (3). Performing the analysis at the single pixel level allows us to determine the d15/d31 and d33/d31 ratios for each pixel, and display the result as images. Figure 3(d) shows the d33/d31 ratio image as a false color scale is used to represent the range of the d33/d31 values. Furthermore, as the d33/d31 histogram in Fig. 3(d) shows, the single-pixel resolved analysis also allows us to obtain statistical data on the distribution of the fitted parameter d33/d31.

 

Fig. 3. Single-pixel resolved, P-SHG image analysis in normal skin dermis. (a). SHG intensity image. (b). Plot of SHG intensity versus the relative angle between laser and fiber for the selected pixel (red square) in A. The fitting parameters determined with Eq. (3) are d33/d31=1.41±0.05, d15/d31=0.78±0.09. (c) Image and (d) histogram of d33/d31 for the normal dermis image shown in (a). Inhomogeneous distribution of d33/d31 is revealed in (c) and (d). Pk and Sd stand for the peak value and standard deviation of the histogram. Scale bar is 50µm.

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We performed P-SHG analysis on seventeen different human skin samples. Figure 4 shows the false-color map of the d33/d31 second order susceptibility tensor element ratio for each of the sample. The palette at the upper right hand corner maps the pseudo colors corresponding to different numerical values of d33/d31. The images on the first row (Fig. 4(a)4(d)) are the results of the four normal skin samples, and the subsequent three rows correspond to the skin samples with the pathological conditions of keloid (Fig. 4(e)4(h)), morphea (Fig. 4(i)4(l)), and dermal elastolysis (Fig. 4(m)4(q)). The pixel-resolved map allows convenient inspection of the heterogeneous spatial distributions of d33/d31.

 

Fig. 4. Spatially resolved d33/d31 maps for normal and pathological dermis are shown. (a)~(d): normal, E~H: keloid, I~L: morphea, and M~Q: dermal elastolysis. The false color palette mapping the corresponding d33/d31 values is displayed on the upper right corner. Each image size is 206×206 µm2.

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In addition, the histograms for the d33/d31 ratios for the normal and the three pathological cases studied are shown in Fig. 5(a). Comparison of the distributions shows that the value of d33/d31 for normal skin has lower values and a narrower distribution than that of the pathological cases. However, the histograms for the d15/d31 (Fig. 5(b)) do not show sufficient sensitivity to distinguish between normal and the pathological cases. Detailed analysis of the specimen variations of d33/d31 are displayed in Fig. 6(a) and the corresponding standard deviations are shown in Fig. 6(b).

 

Fig. 5. Histograms of (a) d33/d31 and (b) d15/d31 for normal, keloid, morphea, and dermal elastolysis skin dermis are shown with the peak value (Pk) and the standard deviation (Sd) of each distribution. Histograms of d33/d31 and d15/d31 for normal and pathological skin dermis range respectively from 0.64 to 2.25 and 0.1 to 1.5. Note that the d33/d31 distributions for the pathological samples all have wider spread than that of the normal samples. Such dependence was not observed in the d15/d31 results.

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Fig. 6. (a). Peak values and (b) the standard deviation of the distribution of d33/d31 for each sample. The letter denotes the sample type and the number corresponds to the specimen number. N: normal dermis, K: keloid, M: morphea,, and D: dermal elastolysis.

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Furthermore, our results are numerically summarized in Table 1. From the table, we see that although the d15/d31 ratio does not show differences between the skin samples studied, the d33/d31 ratio can be used to distinguish between normal and the pathological conditions of keloid, morphea, and dermal elastolysis. Specifically, the d15/d31 ratio for normal skin is 0.63± 0.22 while the average d15/d31 value for all the pathological cases is 0.66±0.23 On the other hand, d33/d31 for normal skins average over four samples was measured to be 1.27±0.20, while that for the pathological skins averaged over thirteen samples was determined to be 1.74±0.30. Since histochemical studies of human skin dermis have estimated the composition of normal dermal collagen to be of 80% type I collagen and 20% type III collagen [22, 23], we expect our results to reflect the SHG properties of mainly type I collagen. Data from previous determination of d33/d31 for type I collagen using rat tail tendon and chicken skin ranges from 1.3 to 1.6 and are consistent with our measurements [24, 25]. For the pathological samples, the deviation from the normal d33/d31 value suggests an alteration of collagen properties. These changes would result in a more heterogeneous distribution of the dermal collagen. Moreover, the d33/d31 histograms of normal, keloid, morphea, and dermal elastolysis (Fig. 5(a)) show that two distinct peaks are present in the pathological tissues, suggesting that two populations of collagen are indeed present in pathological dermis. Additional evidence for the increase in the heterogeneity of pathological dermal collagen is observed in the approximately 1.5 times broader distribution of the d33/d31 ratio for pathological dermis as compared to that of the normal skin. The observation that the ratio d15/d31 remains relatively constant in all our samples can be understood in terms of a model for calculating the bulk susceptibility of collagen molecule, where collagen is approximated by a symmetric cylindrical distribution of molecules with single preferred axis of hyperpolarizability [12]. This approximation leads to a constant value for d15/d31, though at a different value from our measurement. Despite this difference, the model provides an intuitive physical interpretation that relates the value of d33/d31 to the pitch angle of the collagen molecule. Larger d33/d31 indicates the collagen fiber has larger axial polarizability.

Tables Icon

Table 1. Numerical summary of second-order susceptibility analysis for normal and pathological dermis

In conclusion, P-SHG microscopy with single-pixel resolution mapping of χ (2) tensor element ratio d33/d31 can provide a quantitative method for diagnosing dermal pathological condition of keloid, morphea, and dermal elastolysis. The minimally invasive nature of our technique and the backward signal collection configuration allow extension of this method to clinical applications and in vivo animal studies. In addition, the sensitivity of P-SHG to molecular conformation and structural arrangements enable the potential application of the technique to other biomedical studies such as thermal injury [26, 27], diabetes [28], photoaging [29, 30], and tumor invasion [7].

Acknowledgments

We like to acknowledge the support of the National Research Program for Genomic Medicine (NRPGM) of the National Science Council (NSC) in Taiwan for this work. This work was completed in the Optical Molecular Imaging Microscopy Core Facility (A5) of NRPGM.

References and links

1. A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006). [CrossRef]  

2. V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008). [CrossRef]  

3. A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006). [CrossRef]  

4. B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990). [CrossRef]  

5. K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004). [CrossRef]  

6. K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004). [CrossRef]  

7. X. Han, R. M. Burke, M. L. Zettel, P. Tang, and E. B. Brown, “Second harmonic properties of tumor collagen: determining the structural relationship between reactive stroma and healthy stroma,” Opt. Express 16, 1846–1859 (2008). [CrossRef]   [PubMed]  

8. H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007). [CrossRef]  

9. W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008). [CrossRef]  

10. P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002). [CrossRef]  

11. R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005). [CrossRef]  

12. S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006). [CrossRef]  

13. S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004). [CrossRef]   [PubMed]  

14. F. Tiaho, G. Recher, and D. Rouede, “Estimation of helical angles of myosin and collagen by second harmonic generation imaging microscopy,” Opt. Express 15, 12286–12295 (2007). [CrossRef]   [PubMed]  

15. K. Beck and B. Brodsky, “Supercoiled protein motifs: The collagen triple-helix and the alpha-helical coiled coil,” J. of Struct. Biol. 122, 17–29 (1998). [CrossRef]  

16. C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008). [CrossRef]  

17. W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009). [CrossRef]  

18. C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008). [CrossRef]   [PubMed]  

19. B. Richards and E. Wolf, “Electromagnetic Diffraction in Optical Systems.2. Structure of the Image Field in an Aplanatic System,” Proc. Roy. Soc. London Ser. A-Math. Phys. Scie. 253, 358–379 (1959). [CrossRef]  

20. S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009). [CrossRef]   [PubMed]  

21. J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008). [CrossRef]   [PubMed]  

22. W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977). [CrossRef]   [PubMed]  

23. E. H. Epstein, “[Alpha1(Iii)]3 Human Skin Collagen - Release by Pepsin Digestion and Preponderance in Fetal Life,” J. Biol. Chem. 249, 3225–3231 (1974). [PubMed]  

24. P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002). [CrossRef]   [PubMed]  

25. C. Odin, T. Guilbert, A. Alkilani, O. P. Boryskina, V. Fleury, and Y. Le Grand, “Collagen and myosin characterization by orientation field second harmonic microscopy,” Opt. Express 16, 16151–16165 (2008). [CrossRef]   [PubMed]  

26. S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005). [CrossRef]   [PubMed]  

27. Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006). [CrossRef]   [PubMed]  

28. B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000). [CrossRef]   [PubMed]  

29. S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005). [CrossRef]   [PubMed]  

30. M. J. Koehler, K. Konig, P. Elsner, R. Buckle, and M. Kaatz, “In vivo assessment of human skin aging by multiphoton laser scanning tomography,” Opt. Lett. 31, 2879–2881 (2006). [CrossRef]   [PubMed]  

31. R. W. Boyd, Nonlinear Optics (Academic Press, San Diego, CA., 1992), Chap. 1.

References

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  1. A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
    [CrossRef]
  2. V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
    [CrossRef]
  3. A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006).
    [CrossRef]
  4. B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990).
    [CrossRef]
  5. K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
    [CrossRef]
  6. K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
    [CrossRef]
  7. X. Han, R. M. Burke, M. L. Zettel, P. Tang, and E. B. Brown, “Second harmonic properties of tumor collagen: determining the structural relationship between reactive stroma and healthy stroma,” Opt. Express 16, 1846–1859 (2008).
    [CrossRef] [PubMed]
  8. H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
    [CrossRef]
  9. W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
    [CrossRef]
  10. P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
    [CrossRef]
  11. R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005).
    [CrossRef]
  12. S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
    [CrossRef]
  13. S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
    [CrossRef] [PubMed]
  14. F. Tiaho, G. Recher, and D. Rouede, “Estimation of helical angles of myosin and collagen by second harmonic generation imaging microscopy,” Opt. Express 15, 12286–12295 (2007).
    [CrossRef] [PubMed]
  15. K. Beck and B. Brodsky, “Supercoiled protein motifs: The collagen triple-helix and the alpha-helical coiled coil,” J. of Struct. Biol. 122, 17–29 (1998).
    [CrossRef]
  16. C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
    [CrossRef]
  17. W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
    [CrossRef]
  18. C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
    [CrossRef] [PubMed]
  19. B. Richards and E. Wolf, “Electromagnetic Diffraction in Optical Systems.2. Structure of the Image Field in an Aplanatic System,” Proc. Roy. Soc. London Ser. A-Math. Phys. Scie. 253, 358–379 (1959).
    [CrossRef]
  20. S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
    [CrossRef] [PubMed]
  21. J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
    [CrossRef] [PubMed]
  22. W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
    [CrossRef] [PubMed]
  23. E. H. Epstein, “[Alpha1(Iii)]3 Human Skin Collagen - Release by Pepsin Digestion and Preponderance in Fetal Life,” J. Biol. Chem. 249, 3225–3231 (1974).
    [PubMed]
  24. P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
    [CrossRef] [PubMed]
  25. C. Odin, T. Guilbert, A. Alkilani, O. P. Boryskina, V. Fleury, and Y. Le Grand, “Collagen and myosin characterization by orientation field second harmonic microscopy,” Opt. Express 16, 16151–16165 (2008).
    [CrossRef] [PubMed]
  26. S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
    [CrossRef] [PubMed]
  27. Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
    [CrossRef] [PubMed]
  28. B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
    [CrossRef] [PubMed]
  29. S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
    [CrossRef] [PubMed]
  30. M. J. Koehler, K. Konig, P. Elsner, R. Buckle, and M. Kaatz, “In vivo assessment of human skin aging by multiphoton laser scanning tomography,” Opt. Lett. 31, 2879–2881 (2006).
    [CrossRef] [PubMed]
  31. R. W. Boyd, Nonlinear Optics (Academic Press, San Diego, CA., 1992), Chap. 1.

2009 (2)

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

2008 (7)

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

C. Odin, T. Guilbert, A. Alkilani, O. P. Boryskina, V. Fleury, and Y. Le Grand, “Collagen and myosin characterization by orientation field second harmonic microscopy,” Opt. Express 16, 16151–16165 (2008).
[CrossRef] [PubMed]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

X. Han, R. M. Burke, M. L. Zettel, P. Tang, and E. B. Brown, “Second harmonic properties of tumor collagen: determining the structural relationship between reactive stroma and healthy stroma,” Opt. Express 16, 1846–1859 (2008).
[CrossRef] [PubMed]

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

2007 (2)

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

F. Tiaho, G. Recher, and D. Rouede, “Estimation of helical angles of myosin and collagen by second harmonic generation imaging microscopy,” Opt. Express 15, 12286–12295 (2007).
[CrossRef] [PubMed]

2006 (5)

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

M. J. Koehler, K. Konig, P. Elsner, R. Buckle, and M. Kaatz, “In vivo assessment of human skin aging by multiphoton laser scanning tomography,” Opt. Lett. 31, 2879–2881 (2006).
[CrossRef] [PubMed]

2005 (3)

2004 (3)

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

2002 (2)

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

2000 (1)

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

1998 (1)

K. Beck and B. Brodsky, “Supercoiled protein motifs: The collagen triple-helix and the alpha-helical coiled coil,” J. of Struct. Biol. 122, 17–29 (1998).
[CrossRef]

1992 (1)

R. W. Boyd, Nonlinear Optics (Academic Press, San Diego, CA., 1992), Chap. 1.

1990 (1)

B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990).
[CrossRef]

1977 (1)

W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
[CrossRef] [PubMed]

1974 (1)

E. H. Epstein, “[Alpha1(Iii)]3 Human Skin Collagen - Release by Pepsin Digestion and Preponderance in Fetal Life,” J. Biol. Chem. 249, 3225–3231 (1974).
[PubMed]

1959 (1)

B. Richards and E. Wolf, “Electromagnetic Diffraction in Optical Systems.2. Structure of the Image Field in an Aplanatic System,” Proc. Roy. Soc. London Ser. A-Math. Phys. Scie. 253, 358–379 (1959).
[CrossRef]

Al-Attar, A.

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

Alkilani, A.

Baffet, G.

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

Beck, K.

K. Beck and B. Brodsky, “Supercoiled protein motifs: The collagen triple-helix and the alpha-helical coiled coil,” J. of Struct. Biol. 122, 17–29 (1998).
[CrossRef]

Bercovitch, L.

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

Berman, B.

B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990).
[CrossRef]

Boryskina, O. P.

Boyd, R. W.

R. W. Boyd, Nonlinear Optics (Academic Press, San Diego, CA., 1992), Chap. 1.

Brodsky, B.

K. Beck and B. Brodsky, “Supercoiled protein motifs: The collagen triple-helix and the alpha-helical coiled coil,” J. of Struct. Biol. 122, 17–29 (1998).
[CrossRef]

Brown, E. B.

Brown, J. J.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Buckle, R.

Burke, R. M.

Campagnola, P. J.

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

Celliers, P. M.

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

Chen, J. S.

Chen, S. Y.

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Chen, W. L.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

Chen, Y. C.

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Chen, Y. F.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

Chern, G. W.

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Chou, C. K.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

Chu, S. W.

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Da Costa, V.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Da Silva, L. B.

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

Davison, S. P.

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

Dill, S. W.

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

Dong, C. Y.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
[CrossRef] [PubMed]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

Duncan, M. R.

B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990).
[CrossRef]

Eichler, J.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

Elsner, P.

Epstein, E. H.

E. H. Epstein, “[Alpha1(Iii)]3 Human Skin Collagen - Release by Pepsin Digestion and Preponderance in Fetal Life,” J. Biol. Chem. 249, 3225–3231 (1974).
[PubMed]

Fleury, V.

Fwu, P. T.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

Gailhouste, L.

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

Gay, S.

W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
[CrossRef] [PubMed]

Guilbert, T.

Han, X.

Hsiao, C. H.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Hsiao, C. Y.

Hsu, C. J.

Huang, S. C. M.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Jan, G. J.

Jee, S. H.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
[CrossRef] [PubMed]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

Kaatz, M.

Kauffman, C. L.

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

Kim, B. M.

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

Kim, D.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

Kirschner, R. E.

A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006).
[CrossRef]

Koehler, M. J.

Konig, K.

Le Grand, Y.

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

C. Odin, T. Guilbert, A. Alkilani, O. P. Boryskina, V. Fleury, and Y. Le Grand, “Collagen and myosin characterization by orientation field second harmonic microscopy,” Opt. Express 16, 16151–16165 (2008).
[CrossRef] [PubMed]

Lee, H. S.

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

Lewis, K. G.

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

Li, T. H.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

Lim, R.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Lin, B. L.

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Lin, C. H.

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

Lin, H. C.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Lin, L. C.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

Lin, S. J.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
[CrossRef] [PubMed]

Lin, W. C.

Liu, T. M.

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

Lo, W.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
[CrossRef] [PubMed]

Ma, D. H. K.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Mansfield, J. C.

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

Matcher, S. J.

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

Meigel, W. N.

W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
[CrossRef] [PubMed]

Mess, S.

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

Millard, A. C.

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

Moger, J.

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

Mohler, W. A.

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

Odin, C.

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

C. Odin, T. Guilbert, A. Alkilani, O. P. Boryskina, V. Fleury, and Y. Le Grand, “Collagen and myosin characterization by orientation field second harmonic microscopy,” Opt. Express 16, 16151–16165 (2008).
[CrossRef] [PubMed]

Plotnikov, S. V.

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

Recher, G.

Reiser, K. M.

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

Renault, A.

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

Richards, B.

B. Richards and E. Wolf, “Electromagnetic Diffraction in Optical Systems.2. Structure of the Image Field in an Aplanatic System,” Proc. Roy. Soc. London Ser. A-Math. Phys. Scie. 253, 358–379 (1959).
[CrossRef]

Robinson-Bostom, L.

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

Rouede, D.

Rubenchik, A. M.

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, “Collagen structure and nonlinear susceptibility: Effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity,” Lasers Surg. Med. 27, 329–335 (2000).
[CrossRef] [PubMed]

Slemp, A. E.

A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006).
[CrossRef]

So, P. T. C.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

Stoller, P.

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

Su, P. J.

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

Sun, C. K.

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Sun, C-H.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Sun, Y.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, “Monitoring the thermally induced structural transitions of collagen by use of second-harmonic generation microscopy,” Opt. Lett. 30, 622–624 (2005).
[CrossRef] [PubMed]

Tai, S. P.

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

Tan, H. Y.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. and Tech. 71, 83–85 (2008).
[CrossRef]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

Tang, P.

Teng, S. W.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

Thomassen, J. M.

A. Al-Attar, S. Mess, J. M. Thomassen, C. L. Kauffman, and S. P. Davison, “Keloid pathogenesis and treatment,” Plastic Reconstruct. Surg. 117, 286–300 (2006).
[CrossRef]

Tiaho, F.

Tsai, T. H.

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

Webb, W. W.

R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005).
[CrossRef]

Weber, L.

W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
[CrossRef] [PubMed]

Wei, R.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Wilkel, C. S.

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

Williams, R. M.

R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005).
[CrossRef]

Winlove, C. P.

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

Wolf, E.

B. Richards and E. Wolf, “Electromagnetic Diffraction in Optical Systems.2. Structure of the Image Field in an Aplanatic System,” Proc. Roy. Soc. London Ser. A-Math. Phys. Scie. 253, 358–379 (1959).
[CrossRef]

Wong, B. J-F.

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Wu, R. J.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30, 2275–2277 (2005).
[CrossRef] [PubMed]

Young, T. H.

Zettel, M. L.

Zipfel, W. R.

R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005).
[CrossRef]

Appl. Phy. Lett. (1)

W. L. Chen, T. H. Li, P. J. Su, C. K. Chou, P. T. Fwu, S. J. Lin, D. Kim, P. T. C. So, and C. Y. Dong, “Second harmonic generation chi tensor microscopy for tissue imaging,” Appl. Phy. Lett. 94, 183902(2009).
[CrossRef]

Arc. Facial Plastic Surg. (1)

V. Da Costa, R. Wei, R. Lim, C-H. Sun, J. J. Brown, and B. J-F. Wong, “Nondestructive imaging of live human keloid and facial tissue using multiphoton microscopy,” Arc. Facial Plastic Surg. 10, 38–43 (2008).
[CrossRef]

Arch. Dermatol. Res. (1)

W. N. Meigel, S. Gay, and L. Weber, “Dermal Architecture and Collagen Type Distribution,” Arch. Dermatol. Res. 259, 1–10 (1977).
[CrossRef] [PubMed]

Biophys. J. (5)

P. Stoller, K. M. Reiser, P. M. Celliers, and A. M. Rubenchik, “Polarization-modulated second harmonic generation in collagen,” Biophys. J. 82, 3330–3342 (2002).
[CrossRef] [PubMed]

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. C. So, and C. Y. Dong, “Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging,” Biophys. J. 91, 2620–2625 (2006).
[CrossRef] [PubMed]

R. M. Williams, W. R. Zipfel, and W. W. Webb, “Interpreting second-harmonic generation images of collagen I fibrils,” Biophys. J. 88, 1377–1386 (2005).
[CrossRef]

S. V. Plotnikov, A. C. Millard, P. J. Campagnola, and W. A. Mohler, “Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres,” Biophys. J. 90, 693–703 (2006).
[CrossRef]

S. W. Chu, S. Y. Chen, G. W. Chern, T. H. Tsai, Y. C. Chen, B. L. Lin, and C. K. Sun, “Studies of x((2))/x((3)) tensors in submicron-scaled bio-tissues by polarization harmonics optical microscopy,” Biophys. J. 86, 3914–3922 (2004).
[CrossRef] [PubMed]

British J. Dermatology (1)

B. Berman and M. R. Duncan, “Pentoxifylline Inhibits the Proliferation of Human Fibroblasts Derived from Keloid, Scleroderma and Morphea Skin and Their Production of Collagen, Glycosaminoglycans and Fibronectin,” British J. Dermatology 123, 339–346 (1990).
[CrossRef]

Chap. 1. (1)

R. W. Boyd, Nonlinear Optics (Academic Press, San Diego, CA., 1992), Chap. 1.

Curr. Opin. Ped. (1)

A. E. Slemp and R. E. Kirschner, “Keloids and scars: a review of keloids and scars, their pathogenesis, risk factors, and management,” Curr. Opin. Ped. 18, 396–402 (2006).
[CrossRef]

J. Am. Acad. Dermatology (1)

K. G. Lewis, L. Bercovitch, S. W. Dill, and L. Robinson-Bostom, “Acquired disorders of elastic tissue: Part II. Decreased elastic tissue,” J. Am. Acad. Dermatology 51, 165–185 (2004).
[CrossRef]

J. Biol. Chem. (1)

E. H. Epstein, “[Alpha1(Iii)]3 Human Skin Collagen - Release by Pepsin Digestion and Preponderance in Fetal Life,” J. Biol. Chem. 249, 3225–3231 (1974).
[PubMed]

J. Biomed. Opt. (3)

C. K. Chou, W. L. Chen, P. T. Fwu, S. J. Lin, H. S. Lee, and C. Y. Dong, “Polarization ellipticity compensation in polarization second-harmonic generation microscopy without specimen rotation,” J. Biomed. Opt. 13, 014005 (2008).
[CrossRef] [PubMed]

S. W. Chu, S. P. Tai, T. M. Liu, C. K. Sun, and C. H. Lin, “Selective imaging in second-harmonic-generation microscopy with anisotropic radiation,” J. Biomed. Opt. 14, 010504 (2009).
[CrossRef] [PubMed]

J. C. Mansfield, C. P. Winlove, J. Moger, and S. J. Matcher, “Collagen fiber arrangement in normal and diseased cartilage studied by polarization sensitive nonlinear microscopy,” J. Biomed. Opt. 13, 044020 (2008).
[CrossRef] [PubMed]

J. Cutaneous Pathology (1)

K. G. Lewis, S. W. Dill, C. S. Wilkel, and L. Robinson-Bostom, “Mid-dermal elastolysis preceded by acute neutrophilic dermatosis,” J. Cutaneous Pathology 31, 72–76 (2004).
[CrossRef]

J. Microscopy-Oxford (1)

C. Odin, Y. Le Grand, A. Renault, L. Gailhouste, and G. Baffet, “Orientation fields of nonlinear biological fibrils by second harmonic generation microscopy,” J. Microscopy-Oxford 229, 32–38 (2008).
[CrossRef]

J. of Biomed. Opt. (2)

P. Stoller, B. M. Kim, A. M. Rubenchik, K. M. Reiser, and L. B. Da Silva, “Polarization-dependent optical second-harmonic imaging of a rat-tail tendon,” J. of Biomed. Opt. 7, 205–214 (2002).
[CrossRef]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. K. Ma, S. C. M. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. of Biomed. Opt. 12, 024013 (2007).
[CrossRef]

J. of Struct. Biol. (1)

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Figures (6)

Fig. 1.
Fig. 1.

Graphical illustration showing the polarization direction of the laser relative to the orientation of the fiber. The laser is incident along the +y direction. θL is the angle of the excitation polarization and θF is the angle of fiber orientation measured with respect to the z-axis. The angle between excitation polarization and fiber orientation is ϕ=θL -θF .

Fig. 2.
Fig. 2.

Schematic diagram of the P-SHG microscope. Half and quarter wave plates were used to compensate the depolarization caused by the main dichroic mirror.

Fig. 3.
Fig. 3.

Single-pixel resolved, P-SHG image analysis in normal skin dermis. (a). SHG intensity image. (b). Plot of SHG intensity versus the relative angle between laser and fiber for the selected pixel (red square) in A. The fitting parameters determined with Eq. (3) are d33/d31 =1.41±0.05, d15/d31 =0.78±0.09. (c) Image and (d) histogram of d33/d31 for the normal dermis image shown in (a). Inhomogeneous distribution of d33/d31 is revealed in (c) and (d). Pk and Sd stand for the peak value and standard deviation of the histogram. Scale bar is 50µm.

Fig. 4.
Fig. 4.

Spatially resolved d33/d31 maps for normal and pathological dermis are shown. (a)~(d): normal, E~H: keloid, I~L: morphea, and M~Q: dermal elastolysis. The false color palette mapping the corresponding d33/d31 values is displayed on the upper right corner. Each image size is 206×206 µm2.

Fig. 5.
Fig. 5.

Histograms of (a) d33/d31 and (b) d15/d31 for normal, keloid, morphea, and dermal elastolysis skin dermis are shown with the peak value (Pk) and the standard deviation (Sd) of each distribution. Histograms of d33/d31 and d15/d31 for normal and pathological skin dermis range respectively from 0.64 to 2.25 and 0.1 to 1.5. Note that the d33/d31 distributions for the pathological samples all have wider spread than that of the normal samples. Such dependence was not observed in the d15/d31 results.

Fig. 6.
Fig. 6.

(a). Peak values and (b) the standard deviation of the distribution of d33/d31 for each sample. The letter denotes the sample type and the number corresponds to the specimen number. N: normal dermis, K: keloid, M: morphea,, and D: dermal elastolysis.

Tables (1)

Tables Icon

Table 1. Numerical summary of second-order susceptibility analysis for normal and pathological dermis

Equations (3)

Equations on this page are rendered with MathJax. Learn more.

P=χ(1) E +χ(2) E E +χ(3)E E E + ...,
{Px=(d15sin2φ)E2Pz=(d31sin2φ+d33cos2φ)E2
I Px2 +Pz2 {(d15d31sin2ϕ)2+(sin2ϕ+d33d31cos2ϕ)2} E4

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