Biomedical subjects

K A Bartels

Publications and source records attributed to K A Bartels.

3 recordsLinked to original sources

PubMed1993

Three-dimensional distribution of damaged cells in cryopreserved pancreatic islets as determined by laser scanning confocal microscopy.

The technique of serial optical sectioning by confocal microscopy, in conjunction with off-line digital image analysis, was used to quantify the radial distribution of damaged cells in rat pancreatic islets following cryopreservation. The process consists of imaging frozen-thawed islets of Langerhans using laser scanning confocal microscopy (LSCM). The three-dimensional (3-D) distribution and analysis of the two populations of viable and damaged cells was visualized via acridine orange/propidium iodide (AO/PI) fluorescent staining. In preparation for cryopreservation, isolated and cultured rat pancreatic islets were brought to a 2 M concentration of dimethyl sulphoxide (DMSO) by serial addition at decreasing temperatures. Ice was nucleated in the islet suspension at -10 degrees C, and individual specimens were frozen to -70 degrees C at cooling rates of 1, 3, 10 and 30 degrees C/min in a programmable bulk freezer and subsequently stored in liquid nitrogen. After rapid thawing and serial dilution to remove DMSO, individual islets were prepared with AO/PI stains for imaging on the LSCM. Serial sections of the islets, 2-7 microns in thickness, were obtained and processed to obtain high-contrast images. Analysis algorithms consisted of template masking, grey-level thresholding, median filtering and 3-D blob colouring. The radial distribution of damaged cells in the islets was determined by isolating the cell and computing its distance from the centroid of the 3-D islet volume. An increase in the number of blobs corresponding to single and/or aggregates of damaged cells was observed progressively with distance from the centre towards the periphery of the islet. This pattern of freeze-induced killing of cells within the islet was found to occur consistently in the numerous individual specimens processed.

Animals↗

PubMed1993

Analysis of volumetric changes in rat pancreatic islets under osmotic stress using laser scanning confocal microscopy.

Analysis of the volumetric changes in rat pancreatic islets undergoing shrinkage and/or swelling due to osmotic stress is essential for understanding the mechanism of mass transport between cells and their environment and for optimizing cryopreservation protocols. Addition and removal of cryoprotective additives is an integral component of all cryopreservation processes. We have used laser scanning confocal microscopy (LSCM) of acridine orange/propidium iodide (AO/PI) stained Islets of Langerhans to analyze the effects of osmotic stress induced by exposure to varying concentrations of the cryoprotectant dimethyl sulfoxide (DMSO), on the islet volume at two temperatures 23 degrees C and 15 degrees C. Experiments were conducted by mounting a single islet onto a unique freeze-thaw-perfusion stage on which the system temperature and the chemical composition of the solutions can be precisely controlled. The bathing medium of the islet was rapidly changed from isotonic saline to the desired DMSO osmolality to produce a defined osmotic stress, and the islet was imaged simultaneously using 488 nm argon laser. Three to seven serial sections were obtained through each islet at increments varying between 15 microns and 20 microns. The three-dimensional (3-D) image was segmented into islet and non-islet regions using a combination of median filtering, gray level thresholding and region labeling, and the islet volume was computed by counting voxels. Further, a special analysis algorithm was applied to identify shape changes both locally and globally throughout the islet volume.

Algorithms↗

PubMed1993

Selective laser sintering for the creation of solid models from 3D microscopic images.

An exciting application of using Selective Laser Sintering (SLS) to produce solid models of microscopic specimens imaged with a laser scanning confocal microscope is presented. The SLS model, fabricated by sintering together successive layers of a fine powder with a computer controlled scanning laser, is a true 3-D magnification of the microscopic sample. The 3-D models produced are accurate representations of the data and can be handled and manipulated to evaluate surface details and morphology. The models give an extremely powerful method for 3-D data visualization and tactilization. Three dimensional digital image processing is performed on microscopic images to prepare the data for the selective laser sintering process. The image processing techniques for preparing 3-D images of both translucent and opaque specimens for laser sintering are presented. For translucent specimens, the image processing removes noise and "fills in" inclusions and cavities within the specimen data in order to produce a structurally sound model. When imaging an opaque specimen an image of the upper surface alone is acquired. Image processing is used to remove noise and to Fill the volume under the specimen surface. Sample models of a dandelion (Taraxicum officinale Compositae) pollen grain and the surface of a U.S. penny are presented. These models present examples of both the translucent and opaque data types.

Image Processing, Computer-Assisted↗