Electron Microprobe Analysis and Histochemical Examination of the Calcium Distribution in Human Bone Trabeculae: A Methodological Study Using Biopsy Specimens from Post-Traumatic Osteopenia

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Abstract
This work was supported by grants from the Swedish Medical Research Council (project No. 12X—718) and the Alfred Österlund and the Greta and Johan Kock Research Foundations. Energy dispersive X-ray microanalysis (EDX) (or electron microprobe analysis) of the relative intensity for calcium in different bone trabeculae from the tibia epiphysis, and in different parts of one and the same trabecula, was performed on 3 patients who had earlier had a fracture of the ipsilateral tibia-diaphysis. The variation in intensity was compared with the histochemical patterns obtained with both the Goldner and the von Kóssa staining techniques for detecting calcium in tissues. Previously reported calcium distribution features, found to be typical for posttraumatic osteopenia, such as striated mineralization patterns in individual trabeculae and large differences in mineralization level between different trabeculae, could be verified both by means of the two histochemical procedures and from the electron microprobe analysis. A pronounced difference was observed, however, between the two histochemical staining techniques as regards their sensitivity to detect calcium. The two methods have different turnover points, from negative to positive, along a gradient change of calcium concentrations. To judge from the values obtained from the EDX measurements, the sensitivity of the Goldner technique should be more than ten times higher than that of von Kossa. The EDX measurements gave more detailed information than either of the two histochemical techniques: great variations in the intensity of the calcium peak were found in trabeculae stained as unmineral-ized as well as mineralized.