David Wertheim

Segmentation of microscope images of living cells

This paper describes a segmentation method combining a texture based technique with a contour based method. The technique is designed to enable the study of cell behaviour over time by segmenting brightfield microscope image sequences. The technique was tested on artificial images, based on images of living cells and on real sequences acquired from microscope observations of neutrophils and lymphocytes as well as on a sequence of MRI images. The results of the segmentation are compared with the results of the watershed and snake segmentation methods. The results show that the method is both effective and practical.

SwanMesh: A Multicast Enabled Dual-Radio Wireless Mesh Network for Emergency and Disaster Recovery Services

In this paper we present the design and implementation of our multicast enabled dual-radio wireless mesh network for emergency communications. We have developed a novel implementation of a Multicast extension to the AODV (MAODV) protocol in Linux Kernel 2.6 user space to support multicast operation of our SwanMesh testbed. This paper presents the architecture of our SwanMesh testbed. Furthermore, we present unicast communication throughput test results. We observed that SwanMesh can efficiently deliver data services such as broadband internet in emergency situations using its unicast functionality although there were noticeable throughput drops after each hop. Therefore the SwanMesh multicast operation may be suitable for delivery of real time applications in emergency communication, such as audio and video teleconferencing. We also performed validation tests to ensure correct multicast functionality of our MAODV implementation. The possible application of WMN in emergency is discussed.

Infrared imaging in diabetic foot ulceration

Diabetic foot ulcers present a difficult problem in clinical management because of increased risk of soft-tissue infection in diabetes plus impaired local blood supply due to diabetic vascular disease. Infection of diabetic foot ulcers has particular risk of involvement of the adjacent bone resulting in the serious complication of osteomyelitis. This needs early aggressive antibiotic therapy to avoid even more serious secondary long-term complications, but unfortunately clinical diagnosis and radiological examination may be unhelpful in early osteomyelitis, when antibiotic therapy is most effective. Furthermore the large number of patients plus the chronic nature of diabetic foot ulceration precludes routine investigation for early osteomyelitis by X-ray or isotope bone scanning in every case, for logistic, radiation protection, and cost reasons. This preliminary study has shown significantly increased temperature on infrared imaging, not only around the ulcer, but in the entire foot in patients subsequently confirmed radiologically as having early osteomyelitis.

Measurement of forces associated with compression therapy

Compression therapy is the principal treatment for leg ulcers associated with venous disease. The efficacy of compression therapy can be variable, which may in part be owing to the degree of compression applied. However, if the mechanism of action of this treatment could be better understood, it might be possible to improve its efficacy. It is not clear whether assessment of the degree of compression should be made under static or dynamic conditions, or both. A review of methods used previously suggests the need for a new method of assessment allowing continuous monitoring, even during movement. A system for continuous static and dynamic measurements of compression is described. Using an air chamber and manometer to test the system, agreement within ±3 mmHg is observed. The system is applied to investigate changes in forces, expressed as pressure, under bandages and compression stockings. Application of five bandage systems by experienced nurses to a volunteer shows a marked variation in applied pressure. During short periods of walking, rapid changes in pressure under compression stockings are observed, including some transients of less than 0.25 s. The method is simple to apply and may help to understand further the mechanism of action of compression therapy.

Colocalization analysis of sialomucins CD34 and CD164.

Flow cytometric protocols are employed to identify and characterize hemopoietic stem/progenitor populations before transplantation. Cell surface antigens, including CD34, are employed in this process and widely used in harvest protocols, which largely ignores the potential functional role of such antigens. Transmembrane glycoprotein sialomucins, including CD34 and CD164, have been implicated in cell‐to‐cell interactions and activation. CD164, also expressed on early hemopoietic populations, was reported to have a possible function facilitating CD34+ cells to adhere to bone marrow stroma. In this study, we employed high‐definition laser‐scanning confocal microscopy to investigate CD34 and CD164 surface co‐localization patterns on bone marrow and cord blood cells and to compare the expression patterns using a three‐dimensional computer‐generated method developed in house. Differential interference microscopy analysis revealed bone marrow membrane activity was higher than the corresponding cord blood counterpart, perhaps indicating the marrow microenvironmental nature. Fluorescence analysis of CD34 and CD164 antigens showed both were expressed first in a halo‐like pattern and second in antigen‐dense pockets. Three‐dimensional computer analyses further revealed that this pocketing corresponded to dense crest‐like surface structures appearing to rise from the point of adherence on the slide. Further, it was found that CD34 and CD164 display strong colocalization patterns on cells expressing both antigens. The dual nature of the CD34 and CD164 antigens discovered here lends further evidence to the previous literature implicating a strong functional link between these two sialomucins, which should be considered in the transplantation arena and in the function of such sialomucins as negative regulators of cell proliferation.

Extracting respiratory data from pulse oximeter plethysmogram traces in newborn infants

To investigate whether valid respiratory data can be extracted from the pulse oximeter plethysmographic (pleth) trace in healthy newborn infants, pleth data were collected from the foot, and respiratory airflow was simultaneously measured using a facemask. The pleth waveform was analysed using fast Fourier transform (FFT), low-pass filtering (LPF), and by plotting the peak-to-peak amplitude variation (PtP). Using FFT in 14 term infants, the median (range) respiratory rate from the pleth signal was 43 (30–65) breaths/min, and from the flow signal it was 44 (30–67) breaths/min (median difference 0.01 breaths/min, p>0.05). Both LPF and PtP analysis yielded waveforms with a frequency similar to the respiratory rate. Respiratory information, including respiratory rate and a respiratory-like waveform, can reliably be extracted from the pleth trace of a standard pulse oximeter in newborn infants. Such analysis may be clinically useful for non-invasive assessment of respiratory problems in infants and young children.

Micro-CT analysis of matrix-type drug delivery devices and correlation with protein release behaviour

A series of matrix-type drug delivery devices comprising a continuous phase of microporous poly(epsilon-caprolactone) (PCL) and a dispersed phase of protein particles (gelatin) with defined size ranges (45-90, 90-125 and 125-250 microm) were produced by rapidly cooling suspensions in dry ice followed by solvent extraction from the hardened material. High protein loadings (38-44%, w/w) were achieved and highly efficient protein release (90% of the initial load) was obtained over time periods of 3-11 days depending on particle loading and size range. The duration of protein release was extended from 3 to 11 days by reducing the protein load. Quantitative analysis of Micro-CT images identified a three to four times increase in the population of sub-40 microm pores in those matrices which gave rise to accelerated protein release in 24 h (40% rising to 80%) and reduced duration of protein release (11-3 days). Formation of a high density of channels and fissures (connects) between the particles is indicated, which facilitate fluid ingress and diffusion of solubilised protein molecules. Micro-CT analysis also confirmed the uniformity of particle distribution in the matrices and provided measurements of macroporosity within 5-30% of the theoretical value for materials displaying irregular shaped macropores larger than 90 microm. These findings demonstrate the utility of Micro-CT for optimising the formulation and performance of matrix-type delivery devices for macromolecular entities.

A new method of imaging particle tracks in solid state nuclear track detectors.

Solid state nuclear track detectors are used to determine the concentration of α particles in the environment. The standard method for assessing exposed detectors involves 2D image analysis. However 3D imaging has the potential to provide additional information relating to angle as well as to differentiate clustered hit sequences and possibly energy of α particles but this could be time consuming. Here we describe a new method for rapid high‐resolution 3D imaging of solid state nuclear track detectors. A ‘LEXT’ OLS3100 confocal laser scanning microscope (Olympus Corporation, Tokyo, Japan) was used in confocal mode to successfully obtain 3D image data on four CR‐39 plastic detectors. Three‐dimensional visualization and image analysis enabled characterization of track features. This method may provide a means of rapid and detailed 3D analysis of solid state nuclear track detectors.

Micro-CT in drug delivery.

Micro-computed tomography (micro-CT) has not to date been fully exploited in the area of controlled drug delivery despite its capability for providing detailed, 3-D images of morphology and the opportunity this presents for exploring the relationships between delivery device formulation, structure and performance. Micro-CT was used to characterize the internal structure of polycaprolactone (PCL) matrix-type devices incorporating soluble particulates (lactose Mw 342.30, gelatin Mw 20-25kDa) as models of hydrophilic bioactives or pore-forming excipients. Micro-CT images confirmed that the lactose and gelatin particles were uniformly dispersed throughout the PCL phase and that efficient delivery of 95-100% of each species in 9days involved transport from the matrix core. Quantitative analysis of micro-CT images provided values for matrix macroporosity, which were within 15% of the theoretical value and revealed uniform porosity throughout the samples. Total release of protein occurred in 9days (PBS, 37 degrees C) from matrices containing a high protein load (44%w/w) and was independent of particle size. Measurements of equivalent pore diameter and frequency distribution identified a large population of sub-40microm pores in each material, indicative of a high density of connecting channels between particles which facilitates protein transport through the matrices.

Interactive image processing system for assessment of cell movement

The study of cancer cell motility is considered to be important in understanding cancer metastasis. The movement behaviour of cells within clustered cell colonies is of particular interest. Changes in cell movement, area and velocity can be an indicator of cell spreading. The aim of the study is to develop and apply a computerised interactive image processing system to quantify the movement of cells within cell clusters. A semi-automatic boundary description method based on two-dimensional rendering is devised. The system is later combined with image-processing methods that facilitate the relocation of the cell boundary over time; this forms a new approach to assessing cell movement. These methods are incorporated into a software system, enabling an interactive procedure to define and monitor the movement of single cells in cell clusters from digitised microscope images. Validation of the method shows a maximum error of 10% in defining the area through a cubic spline interpolation. The system is applied to analyse the movement and area of HT115 human colon cancer cells. The system provides tools for the analysis of movement, area and velocity of single cells in cancer cell colonies and may thus be of value in further understanding cancer cell motility.