EVA-enhanced embedding medium for histological analysis of 3D porous scaffold material

Abstract

When sectioning a 3D porous scaffold made of a soft elastomeric material embedded in paraffin medium, it is not easy to obtain a section because of the different mechanical properties of the paraffin and tissue/scaffold. We describe a new embedding material for histological analysis of various biomaterials that is composed of paraffin and ethylene vinyl acetate (EVA) resin (0, 3, 7, and 13 wt.%). 3D porous poly( l-lactide-ɛ-caprolactone) (PLCL) and chitosan scaffolds were fabricated to test the sectioning efficiency of the paraffin/EVA embedding material. The new embedding material was characterized by rheological analysis and solvent solubility testing in xylene and n-hexane. The hydrophilicity of the new material was assessed by contact angle measurement and its surface roughness was measured using AFM analysis. The staining efficiency of sections embedded in a paraffin/EVA mixture was determined by eosin staining of the chitosan scaffold and chitosan/collagen hybrid scaffold using a fluorescently labeled collagen. Section roughness decreased with increasing EVA content. The softening temperature of the paraffin/EVA mixture was similar to that of paraffin (50–60 °C by rheometer). The paraffin/EVA mixture dissolved completely in xylene after 30 min at 50 °C, and after 30 min in n-hexane at 60 °C. Therefore, the new embedding medium can be used for histological analysis of various biomaterials and natural tissues.