Entire 3-Dimensional Image of Red Blood Cells using Defocusing Microscopy

Abstract

We present a simple bright-field optical microscopy technique, which we termed Defocusing Microscopy (DM) [1-3], able to actually reconstruct the 3-dimensional (3D) structure of phase objects. For this reconstruction it is only necessary to measure the object DM contrast at two different objective focal positions. We apply the developed method to red blood cells and obtain the shapes of the upper and lower surfaces, the last one deformed by the substrate. We reconstruct cells in isotonic, hypotonic conditions, as shown in the figure below. Additionally, we obtain the height fluctuations, with nanometer sensitivity, and the elastic modulii of each surface separately. For entire 3D imaging of phase objects in motion, one can use two cameras focused on two different positions. Therefore, DM technique can also be applied to obtain 3D reconstruction of cells flowing in micro-fluidic devices.[1] U. Agero, C.H. Monken, C. Ropert, R.T. Gazzinelli, and O.N. Mesquita, Phys. Rev. E 67, 051904 (2003).[2] L.G. Mesquita, U. Agero, and O.N. Mesquita, Applied Phys. Lett. 88, 133901 (2006).[3] G. Glionna, C.K. Oliveira; L.G. Siman, Moyses HW, D.M.U. Prado, C.H. Monken, and O.N. Mesquita, Applied Phys. Lett. 94, 193701 (2009).View Large Image | View Hi-Res Image | Download PowerPoint Slide