Total three-dimensional imaging of phase objects using defocusing microscopy: Application to red blood cells

We introduce Defocusing Microscopy (DM), a bright-field optical microscopy technique able to perform total three-dimensional (3D) imaging of transparent objects. By total 3D imaging, we mean the determination of the actual shapes of the upper and lower surfaces of a phase object. We propose a method...

Full description

Saved in:
Bibliographic Details
Published inApplied physics letters Vol. 104; no. 25
Main Authors Roma, P. M. S., Siman, L., Amaral, F. T., Agero, U., Mesquita, O. N.
Format Journal Article
LanguageEnglish
Published Melville American Institute of Physics 23.06.2014
Subjects
Applied physics
Blood
Defocusing
Deformation
Erythrocytes
Glass substrates
Lipids
Membranes
Microscopy
Optical microscopy
Phase objects
Refractivity
Online AccessGet full text

Cover

More Information
Summary:We introduce Defocusing Microscopy (DM), a bright-field optical microscopy technique able to perform total three-dimensional (3D) imaging of transparent objects. By total 3D imaging, we mean the determination of the actual shapes of the upper and lower surfaces of a phase object. We propose a methodology using DM and apply it to red blood cells subject to different osmolality conditions: hypotonic, isotonic, and hypertonic solutions. For each situation, the shapes of the upper and lower cell surface-membranes (lipid bilayer/cytoskeleton) are completely recovered, displaying the deformation of red blood cell (RBC) surfaces due to adhesion on the glass-substrate. The axial resolution of our technique allowed us to image surface-membranes separated by distances as small as 300 nm. Finally, we determine the volume, surface area, sphericity index, and RBC refractive index for each osmotic condition.
Bibliography:ObjectType-Article-1
SourceType-Scholarly Journals-1
ObjectType-Feature-2
content type line 14
ISSN:0003-6951
1077-3118
DOI:10.1063/1.4884420