Scanning electron microscopy in bone pathology: review of methods, potential and applications

This article reviews the applications of SEM methods to human bone pathologies referring to studies made at UCL. We consider the methods which may be most suitable; these prove to be not "routine" in the context of most bio-medical applications of SEM. Valuable information can be obtained from a bone sample if its edges are ground flat, before making either a matrix surface preparation by washing away all the cells or a mineralizing front preparation, by also dissolving the osteoid-for which hydrogen peroxide is recommended to produce a robust specimen. BSE contrast from a cut block surface can be used to measure bone phase volume. SE contrasts from natural surfaces (trabeculae, canals and lacunae) can be used to study forming, resting and resorbing surfaces both qualitatively and quantitatively (except in the case of histological osteomalacia, where the existence of osteoid will go undetected and reversal lines will be difficult to distinguish from recently resorbed surfaces). We also recommend the use of PMMA embedded bone blocks, which can be used as obtained from the pathologist, but are better embedded by a more rigorous procedure. BSE image analysis can be used to quantitate bone density fractions opening up a completely new investigative method for the future. Osteoid can be measured automatically using CL if the bone sample is block stained with brilliant sulphaflavine before embedding or if a scintillant is added to the embeddant. We give examples of observations made from a number of bone diseases: vitamin D resistant rickets, osteogenesis imperfecta; osteomalacia; osteoporosis; hyperparathyroidism; fluorosis; Paget's disease; tumour metastasis to bone.