Chemical staining of particulate organic matter for improved contrast in soil X-ray µCT images

Degradability of organic matter (OM) in soil depends on its spatial location in the soil matrix. A recent breakthrough in 3D-localization of OM combined dual-energy X-ray CT-scanning with OsO4 staining of OM. The necessity for synchrotron-based µCT and the use of highly toxic OsO4 severely limit applications in soil biological experiments. Here, we evaluated the potential of alternative staining agents (silver nitrate, phosphomolybdenic acid (PMA), lead nitrate, lead acetate) to selectively enhance X-ray attenuation and contrast of OM in CT volumes of soils containing specific mineral soil particle fractions, obtained via lab-based X-ray µCT. In comparison with OsO4, administration of Ag+ and Pb2+ resulted in insufficient contrast enhancement of OM versus fine silt (< 20 µm) or clay (< 2 µm) mineral particles. The perfusion procedure used in this work induced changes in soil structure. In contrast, PMA staining resulted in a selective increase of OM’s attenuation contrast, which was comparable to OsO4. However, OM discrimination from other soil phases remained a challenge. Further development of segmentation algorithms accounting for grey value patterns and shape of stained particulate OM may enable its automated identification. If successful in undisturbed soils, PMA staining may form an alternative to OsO4 in non-synchrotron based POM detection.