scholarly journals Automated Confocal Laser Scanning Microscopy and Semiautomated Image Processing for Analysis of Biofilms

1998 ◽  
Vol 64 (11) ◽  
pp. 4115-4127 ◽  
Author(s):  
Martin Kuehn ◽  
Martina Hausner ◽  
Hans-Joachim Bungartz ◽  
Michael Wagner ◽  
Peter A. Wilderer ◽  
...  
Keyword(s):  
Image Analysis ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Image Acquisition ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Three Dimensions ◽  
Confocal Laser ◽  
Cross Sectional ◽  
In Situ Conditions

ABSTRACT The purpose of this study was to develop and apply a quantitative optical method suitable for routine measurements of biofilm structures under in situ conditions. A computer program was designed to perform automated investigations of biofilms by using image acquisition and image analysis techniques. To obtain a representative profile of a growing biofilm, a nondestructive procedure was created to study and quantify undisturbed microbial populations within the physical environment of a glass flow cell. Key components of the computer-controlled processing described in this paper are the on-line collection of confocal two-dimensional (2D) cross-sectional images from a preset 3D domain of interest followed by the off-line analysis of these 2D images. With the quantitative extraction of information contained in each image, a three-dimensional reconstruction of the principal biological events can be achieved. The program is convenient to handle and was generated to determine biovolumes and thus facilitate the examination of dynamic processes within biofilms. In the present study, Pseudomonas fluorescens or a green fluorescent protein-expressing Escherichia coli strain, EC12, was inoculated into glass flow cells and the respective monoculture biofilms were analyzed in three dimensions. In this paper we describe a method for the routine measurements of biofilms by using automated image acquisition and semiautomated image analysis.

scholarly journals Confocal Laser Scanning Microscopy and Image Analysis for Elucidating Crumb and Crust Microstructure of Bran-Enriched South African Fried Dough and Batter

Foods ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 605 ◽  
Author(s):  
Oluwatoyin O. Onipe ◽  
Daniso Beswa ◽  
Afam I. O. Jideani
Keyword(s):  
Image Analysis ◽  
Laser Scanning ◽  
Polynomial Regression ◽  
Initial Moisture Content ◽  
Soxhlet Extraction ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Cross Sectional ◽  
Linear Effect ◽  
Staining Protocol

A double staining protocol for image acquisition using confocal microscopy (CLSM) coupled with image analysis was employed to elucidate the crust and cross-sectional properties of fried dough. Penetrated oil by image analysis (POia), porosity and pore features were quantified from the cross-section micrographs. Crust surface roughness was measured using fractal metrics and fat content was determined by solvent extraction using the American Association of Cereal Chemists method. Crumb porosity ranged between 54.94%–81.84% and reduced (p < 0.05) with bran addition. Crumb pore sizes ranged from 0–475 µm with <1 circularity, indicating elliptical shape. POia values were notably higher (p < 0.05) than PO by Soxhlet extraction (POsox), except for wheat bran (WB) fried dough where the values of POia and POsox were closely ranked. The linear effect of initial moisture content and bran concentration showed a significant impact on the image properties. The mean fractal dimension (FD) decreased as initial moisture increased. The addition of WB caused a significant reduction in the FD of fried dough, while the opposite effect was noted for its oat bran counterpart. Due to non-collinearity of image properties (FD, POia and porosity), data were fitted to cubic polynomial regression with R2 values > 0.70. CLSM and image analysis were effective in measuring oil absorption and interpreting crumb properties of fried dough. The protocol used in this study can be applied to other thick deep-fried foods for qualitative observation and quantitative measurement of a specific physical or chemical property.

Confocal laser scanning microscopy and Raman imagery of the late Neoproterozoic Chichkan microbiota of South Kazakhstan

2010 ◽  
Vol 84 (3) ◽  
pp. 402-416 ◽  
Author(s):  
J. William Schopf ◽  
Anatoliy B. Kudryavtsev ◽  
Vladimir N. Sergeev
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Three Dimensional ◽  
Laser Scanning Microscopy ◽  
Three Dimensions ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser ◽  
Depth Study ◽  
Late Neoproterozoic

Precambrian microbiotas, such as that permineralized in bedded and stromatolitic cherts of the late Neoproterozoic, 750- to 800-Ma-old, Chichkan Formation of South Kazakhstan, have traditionally been studied by optical microscopy only. Such studies, however, are incapable of documenting accurately either the three-dimensional morphology of such fossils or their chemical composition and that of their embedding minerals. As shown here by analyses of fossils of the Chichkan Lagerstätte, the solution to these long-standing problems is provided by two techniques recently introduced to paleontology: confocal laser scanning microscopy (CLSM) and Raman imagery. The two techniques are used together to characterize, in situ and at micron-scale resolution, the cellular and organismal morphology of the thin section-embedded organic-walled Chichkan fossils. In addition, Raman imagery is used to analyze the molecular-structural composition of the carbonaceous fossils and of their embedding mineral matrix, identify the composition of intracellular inclusions, and quantitatively assess the geochemical maturity of the Chichkan organic matter.CLSM and Raman imagery are both broadly applicable to the study of fossils, whether megascopic or microscopic and regardless of mode of preservation, and both are non-intrusive and non-destructive, factors that permit their use for analyses of archived specimens. They are especially useful for the study of microscopic fossils, as is demonstrated in this first in-depth study of diverse taxa of a single Precambrian microbiota for which they provide information in three dimensions at high spatial resolution about their organismal morphology, cellular anatomy, kerogenous composition, mode of preservation, and taphonomy and fidelity of preservation.

“Grid-mapped” freeze-fracture: Three-Dimensional confocal laser scanning microscopy for directed fracturing and histological mapping of neurons in spinal cord and brain

1993 ◽  
Vol 51 ◽  
pp. 64-65
Author(s):  
J. E. Rash ◽  
L. R. Whalen ◽  
P. B. Guthrie ◽  
M. Morita ◽  
R. Dillman ◽  
...  
Keyword(s):  
Motor Neurons ◽  
Laser Scanning ◽  
Three Dimensional ◽  
Freeze Fracture ◽  
Laser Scanning Microscopy ◽  
Three Dimensions ◽  
Male Rats ◽  
Whole Body ◽  
Confocal Laser ◽  
Fracture Plane

A new correlative microscopic technique, “grid-mapped” freeze fracture, is introduced. This technique allows individual cells in histological slices to be freeze fractured, and their ultrastructural details to be correlated with conventional histological and gross anatomical features. Adult male rats were anesthetized, the sciatic nerve was exposed and crushed, and rhodaminefilled latex microspheres (Lumafluor, Inc) were injected at the crush site to label motor neurons. After 3-7 days, rats were fixed by whole-body perfusion. The brains and spinal cords were removed, embedded in 5% gelatin, and 50 or 100 μm thick slices were cut with a Vibratome. Slices were mapped in three-dimensions (Figs. 1-2 and 3-4) using a Molecular Dynamics Multiport 2001 confocal microscope, and the depths of selected cells were measured (±2μm) from the cut surfaces. After freezing on gold specimen supports, the fracture plane was directed through selected neurons using the precise planar microtome of the JEOL JFD-9000-CR freeze-fracture machine.

scholarly journals Potential role of nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase in apoptosis and oxidative stress

2001 ◽  
Vol 114 (9) ◽  
pp. 1643-1653 ◽  
Author(s):  
Z. Dastoor ◽  
J.L. Dreyer
Keyword(s):  
Oxidative Stress ◽  
Laser Scanning ◽  
Fluorescent Protein ◽  
Nuclear Translocation ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Apoptotic Cells ◽  
Transfected Cells ◽  
And Oxidative Stress

Recent studies indicating a role of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in apoptosis or oxidative stress has been reported. Using confocal laser-scanning microscopy, we have investigated the cellular distribution of GAPDH in central nervous system (CNS)-derived cells (neuroblastoma mNB41A3), in non-CNS derived cells (R6 fibroblast) and in an apoptosis-resistant Bcl2 overexpressing cell line (R6-Bcl2). Induction of apoptosis by staurosporine or MG132 and oxidative stress by H(2)O(2) or FeCN enhanced the nuclear translocation of endogenous GAPDH in all cell types, as detected by immunocytochemistry. In apoptotic cells, GAPDH expression is three times higher than in non-apoptotic cells. Consistent with a role for GAPDH in apoptosis, overexpression of a GAPDH-green fluorescent protein (GAPDH-GFP) hybrid increased nuclear import of GAPDH-GFP into transfected cells and the number of apoptotic cells, and made them more sensitive to agents that induce apoptosis. Bcl2 overexpression prevents nuclear translocation of GAPDH and apoptosis in untransfected cells, but not in transfected cells that overexpress GAPDH-GFP. Our observations indicate that nuclear translocation of GAPDH may play a role in apoptosis and oxidative stress, probably related to the activity of GAPDH as a DNA repair enzyme or as a nuclear carrier for pro-apoptotic molecules.

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