Polarimetric second-harmonic generation (SHG) microscopy is a powerful imaging tool that enables to characterize a nanoscopic structural organization of biological materials beyond the diffraction limited resolution. The ultrastructural organization within individual voxels of the images is characterized by the nonlinear susceptibility tensor. Up to 6 observable susceptibility tensor components can be measured with double Stokes-Mueller polarimetry of SHG signal recorded with a laser scanning microscope. The polarimetric measurement is performed with a complete set (9 in total) of incoming laser beam polarization states, and for each incoming polarization state a Stokes vector is measured for outgoing SHG signal (Fig 1). The 4x9 matrix of images obtained during polarization experiments are converted to a double Mueller matrix of 4x9 dimensions. A correlation matrix X of susceptibility tensor components is obtained from the double Mueller matrix. The Hermitian X matrix represents products of complex measurable susceptibility tensor components. The X matrix provides information about the phase relations between various susceptibility components and allows to determine the degree of disorder in each of the measured voxel. The X matrix also allows to extract the orientation of the symmetry axis in the individual voxels of the imaged structure. The double Stokes-Mueller polarimetric microscopy has been applied for investigations of cardiac muscle tissue and collagen in the extracellular matrix of malignant and normal tissues of various organs. A clear differentiation between tumor and normal tissue has been obtained. The method can be readily used for histopathology investigations. Together with traditional H&E staining images, SHG polarimetric microscopy can aid pathologist in cancer diagnostics of various tissue types.

Ultrastructural imaging with second harmonic generation Stokes-Mueller polarimetric microscopy

256th National Meeting (2018)

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