Transport-of-intensity equation (TIE) based phase imaging in a confocal laser scanning microscope

Abstract

Conventional transport-of-intensity equation (TIE) based phase imaging is performed in wide-field microscopes. In this paper, we present phase and fluorescence dual-modality imaging in a confocal laser scanning microscopy (CLSM) system. To perform phase imaging, the depth of field (DOF) of the CLSM system was extended by using a tunable acoustic gradient index of refraction (TAG) lens. Under transmitted illumination, a few intensity images of a sample at different defocusing distances were recorded. The phase image is reconstructed from these intensity images by using transport-of-intensity equation (TIE). Fluorescence image is obtained by 3D scan of the sample, providing a 3D sectioned fluorescence image. The obtained dual-modality images with pixel-to-pixel correspondence provide for the same sample complementary information (structural/functional), to extract complex biological parameters. We demonstrate the combination of the two imaging modalities enables standalone determination of the refractive index of live cells.