Syllabus
1. Introduction to quantitative plant anatomy definition of scientific field, historical milestones, tradition in the Department of Plant Physiology, trends in current development of the field, fractal geometry, theory of Chaos, stereology, image analysis, application of quantitative plant anatomy in research
2. Traditional morphometric methods I. Length measurements in 2D, calibration of measurements, thickness measurements, area estimation (point-counting method and method of linear integration). Examples - leaf thickness, diameter of root metaxylem vessels, stomatal density, etc.
3. Traditional morphometric methods II. Measurements of 2D curve (line-intercept method, modified Buffon method, difference between isotrpic and anisotropic structures), counting of particles in 2D. Systematic uniform random sampling. Examples - measurement of length of leaf venation, stamatal density, Salisbury's index.
4. Stereological methods I. Introduction to stereological methods, geometrical selection, types of geometrical selection.
5. Stereological methods II. Cavalieri´s estimator, estimation of object volume and volumetric proportions by point-counting method. Examples: organ volume, e.g. leaf volume, proportion of tissues in an organ)
6. Stereological methods III. Particle counting in 3D, volume of irregularly shaped particles, (disector principle). Examples - estimation of the cell number and the mean cell volume of leaf mesophyll cells.
7. Stereological methods IV. Estimation of surface area (method of vertical sections and orientator). Examples - generation of isotropic sections, organ surface, inner leaf surface.
8. Stereological methods VI. Modern methods (nukleator, selector, fakir, etc.) and current trends of development.
9. Measurement of root length Curve measurement in 3D (method of total vertical projections, comparison with image analysis). Examples -measurement of root length by image analysis, line-intercept method and method of total vertical projection.
10. Introduction to image analysis Basic terms, human image perception, historical milestones in development of computer technology and image processing and analysis, basic processes of image analysis, basic terms of operations of methematical morphology. Image J - basics for users. Ellipse - image processing software.
11. Confocal microscopy, 3D image analysis Excursion to confocal microscopy in Physiological Institute of CAS, demonstration of applications using confocal microscopy, stereological methods and 3D image analysis - space reconstructions.
12. Design of experiment, technical aspects of usage of stereological methods General requirements during plannig of biological experiment. Unbiasedness of estimators, systematical uniform random sampling, sources of material variability, statistical measures of variability, example during application of point-counting method - determination of density and distribution of test system, number of sampling fields, etc., generalization of directions applied when planning experimental design. Orientation in literature, requirements for examination, final discussion
Annotation
Course deals with practical review of available methods for quantitative description of biological anatomical and morphological structures with demonstration of their application in biological research. Traditional morphometric and stereological methods, introduction to image analysis, experimental design, principles of statistical evaluation of data.
Practical training includes hand-on examples of methods, way of data acquiring and processing, calculations. Image analysis is demonstrated on J-image and Ellipse software programs.