In this study, we extend on the three parameter analysis approach of utilizing a noninvasive dual-liquid-crystal-based polarization-resolved second harmonic generation (SHG) microscopy to facilitate the quantitative characterization of collagen types I and II in fracture healing tissues. The SHG images under various linear and circular polarization states are analyzed and quantified in terms of the peptide pitch angle (PA), SHG-circular dichroism (CD), and anisotropy parameter (AP). The results show that the collagen PA has a value of 49.26° after 2 weeks of fracture healing (collagen type II domination) and 49.05° after 4 weeks (collagen type I domination). Moreover, the SHG-CD and AP values of the different collagen types differ by 0.05. The change tendencies of the extracted PA, SHG-CD, and AP parameters over the healing time are consistent with the collagen properties of healthy nonfractured bone. Thus, the feasibility of the proposed dual-liquid-crystal-based polarization-SHG method for differentiating between collagen types I and II in bone fracture healing tissue is confirmed.
© 2022. The Author(s).