Quantifying Changes in Fibrin Fiber Network Morphology

Abstract

During blood clotting, thrombin and fibrinogen interact, whereby thrombin cleaves the fibrinogen molecule into two peptides, the fibrinopeptides, ultimately forming fibrin monomers. These fibrinogen monomers assemble to form a fibrin network that may be studied using ultrastructural techniques. This study investigates the use of a grid, placed onto a micrograph, to quantify changes in morphology. The fibrin fiber micrographs of a healthy donor were compared to a database of donors and were shown to be a true representative of a typical healthy individual. Eighteen micrographs of this single donor were taken at 40,000× machine magnification, and a grid was placed over the micrographs. The grid dimensions were calculated by using the scale bar inserted onto the micrograph. Each grid block was equal to 0.5 by 0.5 µm for a total grid area of 28 µm(2). A percentage changed fibrin fiber morphology was then calculated for each 28 µm(2) of fibrin clot produced in the laboratory. It is concluded that this effortless and simple grid technique to quantify changes in ultrastructure of fibrin clot morphology may provide a method to statistically quantify changes in fibrin fiber ultrastructure when studying conditions affecting hemostasis.

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“…A representative micrograph of each sample was selected, and subsequently a 10x5 grid was superimposed onto the picture. One fiber was randomly selected out of every block of the grid, ensuring that fiber measurements were not duplicated, and that fibers were systematically assessed to prevent observer bias [29,30]. Fibrin fiber thickness was measured with ImageJ (ImageJ is a public domain, Java-based image processing program developed at the National Institutes of Health: http://rsbweb.nih.gov/ij/).…”