Adapting Image J to perform Bone Histomorphometry

Goal: Develop method by which Bone histomorphometry can be performed without cost by use of free software from the NIH.

Bone histomorphometry uses stained bone slides to measure various bone parameters

Commercial software is used to perform the measurements But these are all based on

basic measurements of length, area, and number

Software specifically tailored towards bone field simplifies the measurements, but little competition leads to high prices

Image J can perform the basic same measurements (Area, length and number)

LengthArea

Image J calculates area accurately Compared Image J calculated vs manually

calculated measurements of area from various samples.

Control sample was 2 * 2 square inch box Started creating void areas within the box to

make sure that image J can measure complicated images

Image J introduces error with distance measurements of circles On a

computer screen, circles are made of square pixels

Comparing to manual calculations, image J introduced an average error of 5.4%

Image J vs Manual calculation of Perimeter

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Diameter of circle (mm)

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Difference between numbers vs % error

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Diameter of Circle (mm)

Difference betw eenvalues

% error

Correcting for error introduced Error was consistent in that Image J was over

estimating the distance and a little over 5% Excel equation can automatically remove the error

introduced by Image J. Average error was reduced from 5.4% to .29%

Original vs adjusted % error

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Diameter of Circle (mm)

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Original % error

Variable Adjusted %error

Average adjusted %error

Next steps

All bone histomorphometry is performed on undecalcified bone samples embedded in methyl methacrylate We have some bone samples, we

need to get the materials to do the embedding

Also need to figure out proper cutting angles for the polycut microtome

Analysis is performed on samples stained using standard H/E and Goldners Trichrome H/E allows for measurement of bone

and tissue volumes Goldner trichrome reveals

unmineralized bone matrix, allowing measurements of osteoid volumes