We present the measurement of red blood cell (RBC) volume change induced by Ca²⁺ for a live cell imaging with full field quantitative phase microscopy (FFQPM). FFQPM is based on the Mach-Zehnder interferometer combined with an inverted microscopy system. We present the effective method to obtain a clear image and an accurate volume of the cells. An edge detection technique is used to accurately resolve the boundary between the cell line and the suspension medium. The measurement of the polystyrene bead diameter and volume has been demonstrated the validity of our proposed method. The measured phase profile can be easily converted into thickness profile. The measured polystyrene bead volume and the simulated result are about 14.74 μm³ and 14.14 μm³, respectively. The experimental results of our proposed method agree well with the simulated results within less than 4 %. We have also measured the volume variation of a single RBC on a millisecond time scale. Its mean volume is 54.02 μm³ and its standard deviation is 0.52 μm³. With the proposed system, the shape and volume changes of RBC induced by the increased intracellular Ca²⁺ are measured after adding ionophore A23187. A discocyte RBC is deformed to a spherocyte due to the increased intracellular Ca²⁺ in RBC. The volume of the spherocyte is 47.88 μm³ and its standard deviation is 0.19 μm³. We have demonstrated that the volume measurement technique is easy, accurate, and robust method with high volume sensitivity (<0.0000452 μm³) and this provides the ability to study a biological phenomenon in Hematology.

Date Published: 12 February 2009
PDF: 8 pages
Proc. SPIE 7182, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues VII, 71821P (12 February 2009); doi: 10.1117/12.809157

Published in SPIE Proceedings Vol. 7182:
Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues VII
Daniel L. Farkas; Dan V. Nicolau; Robert C. Leif, Editor(s)