MicroCT has emerged as a powerful imaging tool to study biological tissues with high resolution. Imaging soft tissues, such as cardiac samples, requires high molecular weight contrast agents. However, the combination of contrast agents and large sample sizes attenuates x-ray transmission almost completely, compromising the bandwidth of x-ray detection, signal/noise ratio and micro-structural definition.

To define tissue pre-treatment for optimal microCT image quality of large mammalian hearts.

Hearts from male pigs (40-50kg, N=5) were excised and blood rinsed using cardioplegic solution (4°C). The coronaries were cannulated and hearts treated either by:

– hydrated samples with conventional contrast agent (iodine, 0.5%);

– dehydrated in ethanol followed by acetone and air-dried under hexamethyldisilazane (HMDS).

Hearts were imaged at 21.7μm isotropic resolution by microCT (SkyScan 1276, Bruker).

Conventional tissue treatments required imaging samples immersed in water to prevent deformation during long acquisitions (18hours). Yet air-dried samples remained morphologically stable in air. X-ray transmission of hydrated samples was attenuated 93% (Fig 1A), compared to 65% for dry samples: conforming better to the optimum bandwidth of the detector (Fig 1B). Histograms of 3D reconstructions show three populations corresponding to air, immersion solution and tissue with considerable overlap in frequency distribution of pixel intensity (Fig 1C). But for dry samples, two distinct populations were found (Fig 1D). Using histogram minima to define segmentation thresholds, tissue boundaries and intramural structures were not discernible from the background in hydrated samples (Fig 1E), whereas for dry samples, tissue, including myocardial fiber bundles, vessels and fat were segmented (Fig 1F).

Air drying with HMDS vastly improved microCT imaging of large mammalian hearts while preserving micro-structure.