Albert Einstein College of Medicine
November 22, 2011
Forward and inverse methods applied to imaging the topography and reflectivity of the ventral surface of fixed and live cells using confocal interference microscopy
In spite of the advantages associated with the molecular specificity of fluorescence imaging, there is still a need to augment these approaches with label-free imaging. Therefore, we have implemented a form of confocal interference microscopy based upon phase-shifted, laser-feedback interferometry so that we can determine the topography and reflectivity of the ventral surface of fixed and live cells grown on glass coverslips. In order to separate the contribution of the elastically scattered light by sub-cellular structures from the reflection at the coverslip-buffer interface, we address the forward problem such that, given a priori information we can predict the experimentally observed topography and reflectivity. We also developed a solution to the relevant inverse problem because the majority of applications lack a priori information regarding reflectivity and shape. This approach offers an opportunity to probe protein aggregation, index of refraction variations and structure. We have applied this approach to imaging the topography and reflectivity of: (1) calibration spheres nanometers above a glass surface; (2) integrin adhesions at the ventral surface and (3) invadopodia in both fixed and live cells.