Zinc acetate

Native and decellularised tissue samples from each of six different human aortas were fixed in zinc fixative solution (0.5% w/v zinc chloride (VWR Chemicals), 0.5% w/v zinc acetate (VWR Chemicals), in 0.1 M Tris buffer containing 0.05% w/v calcium acetate (Sigma) at pH 7.4) for 48 h, dehydrated, and embedded in paraffin wax using a Leica Tissue Processor (TP1020). Serial sections (6–8 µm) were stained with haematoxylin and eosin (H&E), picrosirius red/Millers elastin (to visualize collagen and elastin) and alcian blue (glycosaminoglycans, GAGs) using standard histological methods. Cell nuclei were visualised using DAPI dye (Sigma Aldrich); briefly, sections of tissue were rehydrated in a graded alcohol series before they were incubated in the dark in 0.1 µg mL⁻¹ DAPI dye in dye buffer (10 mM Tris, 1 mM EDTA, 1 mM sodium chloride in water) for 10 min. The sections were viewed using an upright Zeiss Imager M2 microscope under normal Kohler illumination or under fluorescent illumination using a DAPI filter (λ_ex = 340–380 nm/λ_em = 435–485 nm) and images were captured digitally using Zen Blue Pro (Zeiss).

Brain heart infusion (BHI) broth was obtained from Hopebio, Qingdao, China. Several of the chemicals used in this work were purchased from Sigma-Aldrich, HK, including PCP, 2,4-DCP, 4-CP, and Lead (II) nitrate (≥99%). Zinc acetate and copper (II) sulphate pentahydrate were purchased from VWR Chemicals, Radnor, PA, USA. Resazurin sodium salt (90%) was obtained from J&K Scientific, Beijing, China. Whatman chromatography paper (WCP) was selected from Whatman, Hong Kong.