Histomorphometry Outline What is morphometry Why histomorphometry Major challenge
Ward, Terry, Host of “All Things Considered”, Reporter has reference to this Academic Journal, PHwiki organized this Journal Histomorphometry Or How to get numbers out of slides Stephen Greenwald Pathology Group, Institute of Cell & Molecular Science Barts & The London School of Medicine & Dentistry Outline What is morphometry Why histomorphometry Measurement methods Manual Computerised St in addition to ard processes in computerised histomorph. Image capture Enhancement Thresholding Measuring Micro CT of stented arteries What is morphometry A body of methods as long as obtaining numerical in as long as mation about the shape in addition to size of a structure in terms of quantities such as: volume surface area relative amounts of each component orientation, interconnections distribution of substructures etc.
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Why histomorphometry When applied to biological tissue examined microscopically it useful in correlating structure in addition to function e.g. Alveolar or gut villus surface area Arterial composition in addition to elasticity Quantification of Hyperplasia, dysplasia, hypertrophy Immunohistochemical or flourescent markers Area or intensity Major challenge To extract in as long as mation about large 3-D structures from microscopic measurements on thin 2-D sections To do this histomorphometry uses the: Delesse Principle Delesse Principle In a rock composed of a number of minerals, the area occupied by any given mineral is proportional to the volume of the mineral in the rock Repeated determinations of the area fraction will yield an estimate of the volume fraction. The more determinations; the better the estimate. Delesse A. (1847) Procede mechanique pour determines la composition des roches. Comptes Rendus de lAcademie des Science (Paris) 25, 544)
How to estimate area fraction paper cutting in addition to weighing planimetry Planimeter How to estimate area fraction paper cutting in addition to weighing planimetry dot counting
Dot counting Nuclear area/cell area = number of dots in nuclei/number of dots in cell Absolute area of a structure = number of dots in structure x area of dot square How to estimate area fraction paper cutting in addition to weighing planimetry dot counting square counting Square counting 34 squares 7 squares
How to estimate area fraction paper cutting in addition to weighing planimetry dot counting square counting pixel counting in a digital image semi- or fully automatic system Computerised histomorphometry Main processor Video memory ADC LUT Image processor Storage ADC LUT User. mouse, light-pen Microscope TV camera Pixel counting
Pixel counting Recognising objects by colour More difficult measurements length seminiferous tubules surface area alveoli, gut villi etc. counting discrete objects cells, nuclei, alveoli, elastic lamellae etc. Size distribution cells, nuclei, tumours etc.
How to estimate length (Buffons needle problem) If you drop a nail/needle on the floor, what is the probability it will come to rest over a crack between the floor boards Louis le Clerc, Compte de Buffon, (1707 -1788) French naturalist & polymath Buffon needle problem The probability (p) of the needle (or nail) l in addition to ing on a join depends on the length of the needle (l), the width of the boards (d) in addition to the angle it makes with the direction of the boards (). The angle determines the projected (i.e. effective) length of the nail, (lproj) d l Inverse problem; i.e. throw the grid at the nails Imagine a contour of length L composed of small elements, l Now throw the grid (spacing, d) at the nails (i.e. the small elements)
Villus in addition to crypt length measurements How to measure surface area Measure absolute volume (V) of entire organ Archimedes, weight (knowing density) Estimate tissue volume fraction from area fraction calculate tissue volume Count intercepts (Nint) using grid of total length (L) Pattern recognition normal v abnormal morphology displasia, metaplasia counting poorly stained structures nuclei, nuclear organelles, leucocytes
St in addition to ard processes Image capture Enhancement contrast/colour background correction Thresholding (identifying structures of interest) colour intensity shape Measurement area, perimeter, counting 1.5MB 10kB 1kB Image enhancement: shade correction Uneven background illumination Image enhancement: shade correction Original image with uneven illumination
Image enhancement: shade correction Shade corrected image Contrast enhancement: Original
And rendering Conclusions Histomorphometry is useful as long as counting in addition to measuring clearly defined structures Limited by a lack of intelligent software Extremely difficult to better the human eye-brain combination as long as pattern recognition/diagnosis For histopathologists, may be valuable as long as quantifying prognosis Measuring ratio or distribution of different tumour markers No immediate cause as long as alarm amongst histopathologists but watch this space.
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