Dynamic analyses of vibrational motion in cell membranes provide a lot of information on the complex dynamic motilities of a red blood cell (RBC). Here, we present the correlation properties of membrane fluctuation in discocyte and spherocyte RBCs by using quantitative phase microscopy (QPM). Since QPM can provide nanometer sensitivity in thickness measurement within a millisecond time scale, we were able to observe the membrane flicking of an RBC in nanometer resolution up to the bandwidth of 50 Hz. The correlation properties of the vibrational motion were analyzed with the detrended fluctuation analysis (DFA) method. Fractal scaling exponent α in the DFA method was calculated for the vibrational motion of a cell surface at various surface points for normal discocyte and abnormal spherocyte RBCs. Measured α values for normal RBCs are distributed between 0.7 and 1.0, whereas those for abnormal spherocyte RBCs are within a range from 0.85 to 1.2. We have also verified that the vibrational motion of background fluid outside of a cell has an α value close to 0.5, which is a typical property of an uncorrelated white noise.
We present the real-time quantitative analysis of Vibrio vulnificus-infected host cells using quantitative phase microscopy (QPM) based on interferometric techniques. This provides the ability to retrieve the phase or optical path-length distribution over the cell with nanometer path-length sensitivity from a single interferogram image. We have used QPM to study dynamic cell morphologic changes and to noninvasively quantify the cell volumes of rat basophilic leukemia RBL-2H3 cells infected with V. vulnificus strains: wild type (MO6-24/O) and RtxA1 toxin mutant (CMM770). During the process of V. vulnificus infection in RBL-2H3 cells, the dynamic changes of quantitative phase images, cell volumes, and areas were observed in real time using QPM. In contrast, dramatic changes were not detected in RBL-2H3 cells infected with the noncytotoxic RtxA1 toxin mutant. The results showed good correlation between QPM analysis and biochemical assays, such as lactate dehydrogenase assay or β-hexosaminidase release assay. We suggest that QPM is a powerful quantitative method to study the dynamic process of host cells infected with pathogens in a noninvasive manner.
We present the study of the correlation properties of RBC flickering using double-path interferometric quantitative
phase microscopy (QPM) using detrended fluctuation analysis (DFA). For DFA of RBC membrane fluctuations, we
have measured time series thickness variations of a normal RBC for 20 seconds. The amplitude of membrane
fluctuations in RBC have showed significantly larger than the background noise level without a RBC. We have
demonstrated a practical DFA application for QPM by studying the correlation property of RBC membrane fluctuations
in a noninvasive manner. By measuring the fractal scaling exponents of the time series RBC thickness variations
obtained from QPM, we have analyzed the correlation properties of RBC membrane fluctuations and the background
noise without a sample. The exponents for a normal RBC revealed the long-range correlation property in time series
during 20 seconds. However, the averaged exponent for background noise outside a cell was close to the exponent of
white noise.
Monitoring a degranulation process in a live mast cell is a quite important issue in immunology and pharmacology. Because the size of a granule is normally much smaller than the resolution limit of an optical microscope system, there is no direct real-time live cell imaging technique for observing degranulation processes except for fluorescence imaging techniques. In this research, we propose optical quantitative phase microscopy (QPM) as a new observation tool to study degranulation processes in a live mast cell without any fluorescence labeling. We measure the cell volumes and the cross sectional profiles (x-z plane) of an RBL-2H3 cell and a HeLa cell, before and after they are exposed to calcium ionophore A23187 and silver nanoparticles (AgNPs). We verify that the volume and the cross sectional line profile of the RBL-2H3 cell were changed significantly when it was exposed to A23187. When 50 µg/mL of AgNP is used instead of A23187, the measurements of cell volume and cross sectional profiles indicate that RBL-2H3 cells also follow degranulation processes. Degranulation processes for these cells are verified by monitoring the increase of intracellular calcium ([Ca2+]i) and histamine with fluorescent methods.
We present the real time quantitative analysis of Vibriovulnificus-infected host cells using high stability quantitative
phase microscopy (HSQPM). It provides the ability to retrieve the phase or optical path length distribution over the cell
from a single interferogram image, which has been measured with nanometer path length sensitivity for long periods of
time. We have applied HSQPM to study dynamic cell morphologic changes and to quantify noninvasively cell volumes
of rat basophilic leukemia RBL-2H3 cells infected with pathogenic bacteria V. vulnificus strains, wild type (MO6-24/O)
and RTX toxin mutant (CMM770). During the process of V. vulnificus wild type infection to RBL-2H3 cells, the
dynamic changes of quantitative phase images, cell volumes and areas were observed in real time using HSQPM. In
contrast, the dramatic changes were not detected in RBL-2H3 cells infected with RTX toxin mutant. The results showed
the good correlation between HSQPM analysis and biochemical assays such as lactate dehydrogenase (LDH) assay and
β-hexosaminidase release assay. We suggest that HSQPM is useful real time quantitative method to study the dynamic
process of host cells infected with pathogen in a noninvasive manner.
The use of AgNP is becoming more and more widespread in biomedical field. But compared with the promising
bactericidal function, other physiological effects of AgNP on cells are relatively scant. In this research, we propose
quantitative phase microscopy (QPM) as a new method to study the degranulation, and AgNP-induced RBL-2H3 cell
degranulation is studied as well. Firstly, HeLa cells as the cell control and PBS as the solvent control, we measured the
cell volume and cross section profile (x-z plane) with QPM. The results showed that the volume and cross section profile
changed only the RBL-2H3 cells exposed to calcium ionophore A23187, which demonstrates the validity of QPM in
degranulation research. Secondly, 50μg/mL of AgNP was used instead of A23187, and the measurement of cell volume
and cross section profile was carried out again. RBL-2H3 cell volume increased immediately after AgNP was added, and
cross section profile showed that the cell surface became granulated, but HeLa cell was lack of that effect. Phase images
obviously indicated the RBL-2H3 cell deformation. Thirdly, stained with Fluo-3/AM, intracellular calcium Ca2+]i of
single RBL-2H3 cell treated with AgNP was observed with fluorescent microscopy; incubated with AgNP for 20min, the
supernatant of RBL-2H3 cells was collected and reacted with o-phthalaldehyde (OPA), then the fluorescent intensity of
histamine-OPA complex was assayed with spectrofluorometer. The results of Ca2+]i and histamine increase showed that
degranulation of AgNP-induced RBL-2H3 cell occurred. So, the cell volume was used as a parameter of degranulation in
our study and AgNP-induced RBL-2H3 cells degranulation was confirmed by the cell volume increment, cross section
profile change, and [Ca2+]i and histamine in supernatant increase.
We present a novel method to determine the effective elastic constant (EEC) and effective restoring
force (ERF) by using volumetric analysis of Red blood cell (RBC)s with Full field quantitative phase
microscopy (FFQPM). We use the simple harmonic oscillator model to determine EEC and ERF. We
investigate the EECs and ERFs of different shape of RBCs (discocyte, acanocyte, stomatocyte, and
spherocyte) and we investigate the effective temporal coherence of RBCs by analyzing temporal
volumetric behavior of the RBCs.
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.
We present a novel chromatic dispersion measurement method using a spectral domain interferometer for single mode
optical fiber over a wide spectral range (200 nm). This technique is based on the Mach-Zehnder interferometer using a
white light source and spectrometer. A phase was directly retrieved from a measured spectral interferogram to obtain
relative group velocity, chromatic dispersion and dispersion slope. The measured results with the proposed method were
compared with those obtained using a conventional measurement method. Those results have good agreement with each
other. Our proposed method can simply, accurately, and quickly (< 500 ms) measure chromatic information for a short
length of optical fiber as well as optical devices.
Bio-cells and tissues have intrinsic polarization characteristics, which are changed by external stimulus and internal metamorphosis in cells and tissues and some of the bio-cells and tissues have intrinsic birefringence characteristics, which are also changed by external stimulus and internal metamorphosis in cells and tissues. In this paper, we have developed the polarization microscope for measurement of relative phase which results from birefringence characteristics of materials with improved linear polarizing method and have measured relative phase distribution of onion epidermal cells. From the measurement of the relative phase distribution of onion epidermal cells, decrease of relative phase distribution of onion epidermal cells was investigated as the elapse of time. In decrease of relative phase distribution, relative phase of cell membrane in onion epidermal cells decreased radically as compared with that of cytoplasm because decline of function in cell membrane that takes charge of matter transfer in onion epidermal cells has occurred.
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