Biomedical subjects

Keith A Nugent

Publications and source records attributed to Keith A Nugent.

8 recordsLinked to original sources

PubMed2006

Single cell volume measurement by quantitative phase microscopy (QPM): a case study of erythrocyte morphology.

The measurement of the volume of intact, viable cells presents challenging problems in many areas of experimental and diagnostic science involved in the evaluation of cellular morphology, growth and function. This investigation details the implementation of a recently developed quantitative phase microscopy (QPM) method to measure the volume of erythrocytes under a range of osmotic conditions. QPM is a computational approach which utilizes simple bright field optics to generate cell phase maps which, together with knowledge of the cellular refractive index, may be used to measure cellular volume. Rat erythrocytes incubated in imidazole-buffered solutions (22 degrees C) of graded tonicity were analysed using QPM (n=10 cells/group, x63, 0.8 NA objective). Erythrocyte refractive index (1.367) was measured using a combination of phase and morphological data obtained from cells adopting spherical geometry under hypotonic conditions. Phase-computed volume increased with decreasing solution osmolality: 42.8 +/- 2.4, 48.7 +/- 2.3, 62.6 +/- 2.3, 90.8 +/- 7.7 microm3 in solutions of 540, 400, 240, and 170 mosmol/kg respectively. These volume changes were associated with crenated, bi-concave and spherical morphological states associated with increasing tonicity. This investigation demonstrates that QPM is a valid, simple and non-destructive approach for measuring cellular phase properties and volume. QPM cell volume analysis represents a significant advance in viable cell experimental capability and provides for acquisition of 'real-time' data - an option not previously available using other approaches.

Animals↗

PubMed2006

Phase-space reconstruction of focused x-ray fields.

We apply the method of phase-space tomography to reconstruct x-ray beams focused using a compound refractive lens. We show that it is possible to decouple the effect of aberrations in the optical system from the field and hence measure both them and the original field. We recover the complex coherence function and find that it is consistent with expectations.

Journal Article↗

PubMed2005

Refractive index measurement in viable cells using quantitative phase-amplitude microscopy and confocal microscopy.

BACKGROUND: The refractive index (RI) of cellular material provides fundamental biophysical information about the composition and organizational structure of cells. Efforts to describe the refractive properties of cells have been significantly impeded by the experimental difficulties encountered in measuring viable cell RI. In this report we describe a procedure for the application of quantitative phase microscopy in conjunction with confocal microscopy to measure the RI of a cultured muscle cell specimen. METHODS: The experimental strategy involved calculation of cell thickness by using confocal optical sectioning procedures, construction of a phase map of the same cell using quantitative phase microscopy, and selection of cellular regions of interest to solve for the cell RI. RESULTS: Mean cell thickness and phase values for six cell regions (five cytoplasmic and one nuclear) were determined. The average refractive index calculated for cytoplasmic and nuclear regions was 1.360 +/- 0.004. The uncertainty in the final RI value represents the technique measurement error. CONCLUSIONS: The methodology we describe for viable cell RI measurement with this prototype cell has broad generic application in the study of cell growth and functional responses. The RI value we report may be used in optical analyses of cultured cell structure and morphology.

Cell Survival↗

PubMed2005

X-ray imaging: a generalized approach using phase-space tomography.

We discuss the role of coherence in x-ray imaging and consider how phase-space tomography can be used to extract information about partial coherence. We describe the application of phase-space tomography to x-ray imaging and recover the spatial coherence properties of a one-dimensional soft (1.5 keV) x-ray beam from a synchrotron undulator source. We present phase-space information from a Young's experiment and observe negative regions in the quasi-probability distribution. We show that, given knowledge of the coherence of the beam, we can use partially coherent diffraction data to recover fully coherent information, and we present some simple experimental demonstrations of this capability.

Algorithms↗

PubMed2004

Quantitative phase microscopy: a new tool for investigating the structure and function of unstained live cells.

1. The optical transparency of unstained live cell specimens limits the extent to which information can be recovered from bright-field microscopic images because these specimens generally lack visible amplitude-modulating components. However, visualization of the phase modulation that occurs when light traverses these specimens can provide additional information. 2. Optical phase microscopy and derivatives of this technique, such as differential interference contrast (DIC) and Hoffman modulation contrast (HMC), have been used widely in the study of cellular materials. With these techniques, enhanced contrast is achieved, which is useful in viewing specimens, but does not allow quantitative information to be extracted from the phase content available in the images. 3. An innovative computational approach to phase microscopy, which provides mathematically derived information about specimen phase-modulating characteristics, has been described recently. Known as quantitative phase microscopy (QPM), this method derives quantitative phase measurements from images captured using a bright-field microscope without phase- or interference-contrast optics. 4. The phase map generated from the bright-field images by the QPM method can be used to emulate other contrast image modes (including DIC and HMC) for qualitative viewing. Quantitative phase microscopy achieves improved discrimination of cellular detail, which permits more rigorous image analysis procedures to be undertaken compared with conventional optical methods. 5. The phase map contains information about cell thickness and refractive index and can allow quantification of cellular morphology under experimental conditions. As an example, the proliferative properties of smooth muscle cells have been evaluated using QPM to track growth and confluency of cell cultures. Quantitative phase microscopy has also been used to investigate erythrocyte cell volume and morphology in different osmotic environments. 6. Quantitative phase microscopy is a valuable, new, non-destructive, non-interventional experimental tool for structural and functional cellular investigations.

Animals↗

PubMed2004

X-ray phase vortices: theory and experiment.

We review the current work on x-ray phase vortices. We explain the role of an x-ray vortex in phase recovery and speculate on its possible applications in other fields of x-ray optical research. We present our theoretical understanding of the structure of phase vortices and test these predictions against experiment. We present experimental observations of phase vortices with charge greater than 3 and observe that their propagation appears to be consistent with our theoretical models.

Journal Article↗

PubMed2003

Optical metrology for analysis of lobster-eye x-ray optics.

A new method that uses optical microscopy to determine the physical structure of lobster-eye x-ray optics is described. This approach offers the ability to predict x-ray performance without having to take an x-ray measurement. An overlapping series of images of the entrance and exit faces of an optic are obtained and examined by purpose-built software. A 24-parameter description of each channel is obtained from which a quantitative analysis of all the major optic defects, except surface roughness, is performed. Results for a planar lobster-eye optic are used to illustrate this technique and discuss its abilities as well as directions for future enhancements.

Journal Article↗

PubMed2003

Emulated super-resolution using quantitative phase microscopy.

In the case of coherent illumination, knowledge of the phase and the amplitude of a light wave constitutes complete information. Phase and amplitude information can now be simply acquired using the technique of quantitative phase microscopy. It has been shown that this information allows other imaging modalities to be emulated. In this paper we consider how this information may be used to perform a form of super-resolution by emulating the effect of an annular pupil.

Image Processing, Computer-Assisted↗